0	Survivin mediates targeting of the chromosomal, otherFeat=[]-->, belongsTo=title
1	passenger complex to the centromere and midbody, otherFeat=[]-->, belongsTo=title
2	Gerben Vaderw,Jos J.W. Kauw,Rene? H. Medemaw & Susanne M.A. Lens+w, otherFeat=[]-->, belongsTo=parrnote
3	Division of Molecular Biology, Netherlands Cancer Institute, Amsterdam, The Netherlands, otherFeat=[]-->, belongsTo=title
4	The chromosomal passenger complex (CPC) coordinates chromo-, otherFeat=[]-->, belongsTo=parr
5	somal and cytoskeletal events of mitosis. The enzymatic core of, otherFeat=[]-->, belongsTo=parr
6	this complex (Aurora-B) is guided through the mitotic cell by its, otherFeat=[]-->, belongsTo=parr
7	companion chromosomal passenger proteins, inner centromere, otherFeat=[]-->, belongsTo=parr
8	protein (INCENP), Survivin and Borealin/Dasra-B, thereby, otherFeat=[]-->, belongsTo=parr
9	allowing it to act at the right place at the right time. Here, we, otherFeat=[]-->, belongsTo=parr
10	addressed the individual contributions of INCENP, Survivin and, otherFeat=[]-->, belongsTo=parr
11	Borealin to the proper functioning of this complex. We show that, otherFeat=[]-->, belongsTo=parr
12	INCENP has an important role in stabilizing the complex, and, otherFeat=[]-->, belongsTo=parr
13	that Borealin acts to promote binding of Survivin to INCENP., otherFeat=[]-->, belongsTo=parr
14	Importantly, when Survivin is directly fused to INCENP, this, otherFeat=[]-->, belongsTo=parr
15	hybrid can restore CPC function at the centromeres and midbody,, otherFeat=[]-->, belongsTo=parr
16	even in the absence of Borealin and the centromere-targeting, otherFeat=[]-->, belongsTo=parr
17	domain of INCENP. Thus, Survivin is an important mediator of, otherFeat=[]-->, belongsTo=parr
18	centromere and midbody docking of Aurora-B during mitosis., otherFeat=[]-->, belongsTo=parr
19	Keywords: cytokinesis; spindle checkpoint, otherFeat=[]-->, belongsTo=parr
20	EMBO reports (2006) 7, 85?92. doi:10.1038/sj.embor.7400562, otherFeat=[]-->, belongsTo=parrnote
21	INTRODUCTION, otherFeat=[]-->, belongsTo=title
22	Proper mitosis requires accurate nuclear division, followed by, otherFeat=[]-->, belongsTo=parr
23	cytoplasmic partitioning into two daughter cells during cyto-, otherFeat=[]-->, belongsTo=parr
24	kinesis. Nuclear division is monitored by the spindle assembly, otherFeat=[]-->, belongsTo=parr
25	checkpoint (SAC), which is an intrinsic cell-cycle checkpoint that, otherFeat=[]-->, belongsTo=parr
26	prevents the onset of anaphase until all sister chromatids have, otherFeat=[]-->, belongsTo=parr
27	attained proper attachment to the mitotic spindle (Nicklas, 1997)., otherFeat=[]-->, belongsTo=parr
28	To satisfy this checkpoint, kinetochores of paired sister chromatids, otherFeat=[]-->, belongsTo=parr
29	have to attach in a bipolar manner, creating tension (Tanaka,, otherFeat=[]-->, belongsTo=parr
30	2002). Aurora-B kinase is essential to resolve naturally occurring, otherFeat=[]-->, belongsTo=parr
31	monopolar or syntelic microtubule?kinetochore attachments, otherFeat=[]-->, belongsTo=parr
32	(Tanaka et al, 2002; Lampson et al, 2004), and to communicate, otherFeat=[]-->, belongsTo=parr
33	a lack of bipolarity (that is, tension) to the core SAC (Biggins &, otherFeat=[]-->, belongsTo=parr
34	Murray, 2001; Lens & Medema, 2003). Additionally, in several, otherFeat=[]-->, belongsTo=parr
35	organisms, interference with Aurora-B leads to a failure of, otherFeat=[]-->, belongsTo=parr
36	cytokinesis (Kaitna et al, 2000; Severson et al, 2000; Ditchfield, otherFeat=[]-->, belongsTo=parr
37	et al, 2003; Hauf et al, 2003)., otherFeat=[]-->, belongsTo=parr
38	Aurora-B acts together with Survivin, inner centromere protein, otherFeat=[]-->, belongsTo=parr
39	(INCENP) and Borealin/Dasra-B (hereafter referred to as Borealin)., otherFeat=[]-->, belongsTo=parr
40	These proteins physically interact and show a typical chromo-, otherFeat=[]-->, belongsTo=parr
41	somal passenger localization pattern during mitosis--at the inner, otherFeat=[]-->, belongsTo=parr
42	centromere in (pro)metaphase, on the central spindle during, otherFeat=[]-->, belongsTo=parr
43	anaphase and at the midbody during cytokinesis (Vagnarelli &, otherFeat=[]-->, belongsTo=parr
44	Earnshaw, 2004). Interference with Survivin, INCENP or Borealin, otherFeat=[]-->, belongsTo=parr
45	function disrupts Aurora-B localization and leads to similar mitotic, otherFeat=[]-->, belongsTo=parr
46	defects, as observed after interference with Aurora-B function, otherFeat=[]-->, belongsTo=parr
47	(Vagnarelli & Earnshaw, 2004, and references therein). Aurora-B is, otherFeat=[]-->, belongsTo=parr
48	activated at the G2?M transition and remains active until mitotic, otherFeat=[]-->, belongsTo=parr
49	exit (Yasui et al, 2004). Thus, it is likely that the specific functions, otherFeat=[]-->, belongsTo=parr
50	of Aurora-B are mainly determined by its dynamic localization, otherFeat=[]-->, belongsTo=parr
51	pattern. Until now, INCENP has been proposed to be the targeting, otherFeat=[]-->, belongsTo=parr
52	module of the complex (Ainsztein et al, 1998), but specific, otherFeat=[]-->, belongsTo=parr
53	functions for Survivin and Borealin in the chromosome passenger, otherFeat=[]-->, belongsTo=parr
54	complex (CPC) have yet to be determined., otherFeat=[]-->, belongsTo=parr
55	RESULTS AND DISCUSSION, otherFeat=['U']-->, belongsTo=title
56	An RNAi-complementation to study CPC function, otherFeat=[]-->, belongsTo=title
57	We developed an INCENP RNA interference (RNAi) complemen-, otherFeat=[]-->, belongsTo=parr
58	tation system to study the role of INCENP in the CPC. Endogenous, otherFeat=[]-->, belongsTo=parr
59	INCENP was knocked down by vector-driven short interfering, otherFeat=[]-->, belongsTo=parr
60	RNA (siRNA; Fig 1A; supplementary Fig 1A?C online) and cells, otherFeat=[]-->, belongsTo=parr
61	were reconstituted with an INCENP complementary DNA, otherFeat=[]-->, belongsTo=parr
62	harbouring two silent mutations in the siRNA-targeting region,, otherFeat=[]-->, belongsTo=parr
63	rendering it insensitive to the siRNA (Lens et al, 2003). INCENP-, otherFeat=[]-->, belongsTo=parr
64	depleted cells showed the typical `chromosome passenger, otherFeat=[]-->, belongsTo=parr
65	deficiency' phenotype: defects in chromosome alignment and, otherFeat=[]-->, belongsTo=parr
66	segregation, and a failure to undergo cytokinesis (supplementary, otherFeat=[]-->, belongsTo=parr
67	Fig 1A?C online; Honda et al, 2003). Additionally, Aurora-B and, otherFeat=[]-->, belongsTo=parr
68	BubR1 were mislocalized, and the INCENP-depleted cells failed, otherFeat=[]-->, belongsTo=parr
69	to maintain an active SAC in response to Taxol (supplementary, otherFeat=[]-->, belongsTo=parr
70	Fig 1B,D,E online). All these mitotic defects could be reverted, otherFeat=[]-->, belongsTo=parr
71	by expression of a full-length INCENP containing two silent, otherFeat=[]-->, belongsTo=parr
72	mutations (VSV?INCENP-sil; Figs 1C?E,2F; supplementary, otherFeat=[]-->, belongsTo=parr
73	Fig 2B,C online; for INCENP constructs used, see Fig 4A),, otherFeat=[]-->, belongsTo=parr
74	confirming the specificity of the siRNA and the functionality of, otherFeat=[]-->, belongsTo=parr
75	the complementation approach., otherFeat=[]-->, belongsTo=parr
76	Received 27 May 2005; revised 20 September 2005; accepted 21 September 2005;, otherFeat=[]-->, belongsTo=parrnote
77	published online 21 October 2005, otherFeat=[]-->, belongsTo=parrnote
78	+Corresponding author. Tel: ? 3130 253 9728; Fax: ? 3130 253 8479;, otherFeat=[]-->, belongsTo=parrnote
79	E-mail: s.m.a.lens@med.uu.nl, otherFeat=[]-->, belongsTo=parrnote
80	Division of Molecular Biology, Netherlands Cancer Institute, Plesmanlaan 21, 1066 CX, otherFeat=[]-->, belongsTo=parrnote
81	Amsterdam, The Netherlands, otherFeat=[]-->, belongsTo=parrnote
82	wPresent address: Department of Medical Oncology, University Medical Center,, otherFeat=[]-->, belongsTo=parrnote
83	Str. 2.103 Universiteitsweg 100, 3584 CG Utrecht, The Netherlands, otherFeat=[]-->, belongsTo=parrnote
84	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
85	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
86	scientificreport, otherFeat=[]-->, belongsTo=title
87	scientific report, otherFeat=[]-->, belongsTo=title
88	85, otherFeat=[]-->, belongsTo=nota_cab_pie
89	CPC formation is crucial for protein stability, otherFeat=[]-->, belongsTo=title
90	In line with earlier observations (Honda et al, 2003), total protein, otherFeat=[]-->, belongsTo=parr
91	levels of Aurora-B and Survivin were significantly reduced after, otherFeat=[]-->, belongsTo=parr
92	INCENP depletion (Fig 1A). This reduction was interpreted as an, otherFeat=[]-->, belongsTo=parr
93	indirect consequence of a failed cell division followed by a cell-, otherFeat=[]-->, belongsTo=parr
94	cycle arrest in a tetraploid G1 state (Honda et al, 2003). However,, otherFeat=[]-->, belongsTo=parr
95	we found that INCENP-depleted cells already failed to accumulate, otherFeat=[]-->, belongsTo=parr
96	these proteins during passage through G2 (Fig 1B). Moreover,, otherFeat=[]-->, belongsTo=parr
97	mitotic cells lacking INCENP also showed reduced Aurora-B and, otherFeat=[]-->, belongsTo=parr
98	Survivin levels (Fig 1B,H, lane 2), which indicates that the effect, otherFeat=[]-->, belongsTo=parr
99	of INCENP depletion on protein stability is more direct., otherFeat=[]-->, belongsTo=parr
100	By introducing different INCENP truncation mutants into, otherFeat=[]-->, belongsTo=parr
101	INCENP-depleted cells, we could show that a mutant lacking, otherFeat=[]-->, belongsTo=parr
102	the amino-terminal `centromere-targeting' domain (VSV?, otherFeat=[]-->, belongsTo=parr
103	INCENP47?920-sil; Fig 4A) restored Aurora-B levels, but failed, otherFeat=[]-->, belongsTo=parr
104	to stabilize Survivin (Fig 1G, lane 4). In line with previous, otherFeat=[]-->, belongsTo=parr
105	observations (Ainsztein et al, 1998), this mutant failed to, otherFeat=[]-->, belongsTo=parr
106	concentrate at centromeres during prometaphase and to transfer, otherFeat=[]-->, belongsTo=parr
107	to the midbody following the onset of anaphase (supplementary, otherFeat=[]-->, belongsTo=parr
108	Fig 2B,C online). Consequently, this mutant was unable to restore, otherFeat=[]-->, belongsTo=parr
109	a SAC-mediated arrest in response to Taxol and to overcome the, otherFeat=[]-->, belongsTo=parr
110	cytokinesis defect in INCENP-depleted cells (Fig 1D,E). Because of, otherFeat=[]-->, belongsTo=parr
111	the presence of the carboxy-terminal Aurora-B interaction domain, otherFeat=[]-->, belongsTo=parr
112	(IN-box; Adams et al, 2000), INCENP47?920-sil was able to bind to, otherFeat=[]-->, belongsTo=parr
113	Aurora-B. In contrast, this truncation mutant failed to interact with, otherFeat=[]-->, belongsTo=parr
114	Survivin (Fig 1F, lane 3). Conversely, a construct expressing only, otherFeat=[]-->, belongsTo=parr
115	the first N-terminal 47 amino acids of INCENP (INCENP1?47;, otherFeat=[]-->, belongsTo=note
116	Fig 4A) and thus harbouring the centromere-localization domain, otherFeat=[]-->, belongsTo=parr
117	was able to bind to Survivin (Fig 1F, lane 4), but failed to stabilize, otherFeat=[]-->, belongsTo=parr
118	it (Fig 1G, lane 5). Thus, this domain is both necessary and, otherFeat=[]-->, belongsTo=parr
119	sufficient for interaction with Survivin, but the presence of Aurora-B, otherFeat=[]-->, belongsTo=parr
120	in the complex may be needed for stabilization of Survivin., otherFeat=[]-->, belongsTo=parr
121	Interestingly, Survivin is an in vivo substrate of Aurora-B, otherFeat=[]-->, belongsTo=parr
122	(Wheatley et al, 2004), but whether this phosphorylation affects, otherFeat=[]-->, belongsTo=parr
123	protein stability is unknown. In contrast, the presence of Survivin, otherFeat=[]-->, belongsTo=parr
124	in the complex was not essential to stabilize Aurora-B, because we, otherFeat=[]-->, belongsTo=parr
125	found that a complex of Aurora-B and a truncated INCENP,, otherFeat=[]-->, belongsTo=parr
126	although lacking Survivin, could exist (Fig 1F,G). However,, otherFeat=[]-->, belongsTo=parr
127	whereas Aurora-B protein levels could be restored by expression, otherFeat=[]-->, belongsTo=parr
128	of INCENP47?920-sil, phosphorylation of serine-10 Histone-H3, otherFeat=[]-->, belongsTo=parr
129	in mitotic INCENP-depleted cells remained impaired (Fig 1H,, otherFeat=[]-->, belongsTo=parr
130	lane 4). Thus, stabilization of Aurora-B by INCENP47?920-sil is not, otherFeat=[]-->, belongsTo=parr
131	sufficient to mediate Histone-H3 phosphorylation. This suggests, otherFeat=[]-->, belongsTo=parr
132	that mitotic phosphorylation of Histone-H3 on serine-10 by, otherFeat=[]-->, belongsTo=parr
133	Aurora-B requires either the direct presence of Survivin in the, otherFeat=[]-->, belongsTo=parr
134	complex or centromeric localization of the kinase., otherFeat=[]-->, belongsTo=parr
135	Survivin mediates mitotic localization of the CPC, otherFeat=[]-->, belongsTo=title
136	Although INCENP contains a highly conserved `centromere-, otherFeat=[]-->, belongsTo=parr
137	targeting' domain in its N-terminus (Ainsztein et al, 1998), the, otherFeat=[]-->, belongsTo=parr
138	mechanism by which this domain mediates centromeric localiza-, otherFeat=[]-->, belongsTo=parr
139	tion remains elusive. As INCENP interacts with Survivin by means, otherFeat=[]-->, belongsTo=parr
140	of this N-terminal domain (Fig 1F), it is possible that Survivin, otherFeat=[]-->, belongsTo=parr
141	mediates mitotic localization of the CPC. To test this, a chimeric, otherFeat=[]-->, belongsTo=parr
142	protein that is refractory to both INCENP and Survivin RNAi, otherFeat=[]-->, belongsTo=parr
143	(Fig 2A; data not shown) was generated, in which the centromere-, otherFeat=[]-->, belongsTo=parr
144	targeting domain in INCENP was replaced by Survivin (Survivin-, otherFeat=[]-->, belongsTo=parr
145	INCENP47?920-sil; Fig 4A). This fusion localized normally during, otherFeat=[]-->, belongsTo=parr
146	mitosis, in both an INCENP-depleted and a combined INCENP/, otherFeat=[]-->, belongsTo=parr
147	Survivin-depleted background (Fig 2D,G, arrows). This chimaera, otherFeat=[]-->, belongsTo=parr
148	was also able to localize Aurora-B during mitosis (Fig 2E,F),, otherFeat=[]-->, belongsTo=parr
149	restore serine-10 Histone-H3 phosphorylation (supplementary, otherFeat=[]-->, belongsTo=parr
150	Fig 2D online) and restore CPC function, as expression of the, otherFeat=[]-->, belongsTo=parr
151	chimaera in INCENP-, Survivin- or combined INCENP/Survivin-, otherFeat=[]-->, belongsTo=parr
152	depleted cells rescued SAC function and the proper execution of, otherFeat=[]-->, belongsTo=parr
153	cytokinesis (Fig 2B,C)., otherFeat=[]-->, belongsTo=parr
154	To obtain further evidence that Survivin can serve as a, otherFeat=[]-->, belongsTo=parr
155	determining factor in localizing the CPC, we analysed the location, otherFeat=[]-->, belongsTo=parr
156	of INCENP1?47 in the absence or presence of endogenous, otherFeat=[]-->, belongsTo=parr
157	Survivin. INCENP1?47?GFP (green fluorescent protein) localized, otherFeat=[]-->, belongsTo=parr
158	to centromeres during mitosis in control cells (Fig 2H, upper, otherFeat=[]-->, belongsTo=parr
159	panel, see arrows), but failed to concentrate at centromeres in the, otherFeat=[]-->, belongsTo=parr
160	absence of endogenous Survivin and instead localized diffusely in, otherFeat=[]-->, belongsTo=parr
161	the cytoplasm during mitosis (Fig 2G, lower panel, see arrows),, otherFeat=[]-->, belongsTo=parr
162	indicating that the centromere-targeting domain in INCENP, otherFeat=[]-->, belongsTo=parr
163	requires at least the presence of Survivin to localize to, otherFeat=[]-->, belongsTo=parr
164	centromeres. Taken together, these results show that Survivin is, otherFeat=[]-->, belongsTo=parr
165	an important mediator of mitotic localization of the CPC., otherFeat=[]-->, belongsTo=parr
166	Survivin can target the CPC in the absence of Borealin, otherFeat=[]-->, belongsTo=title
167	Recently, it was suggested that Borealin has a role in determining, otherFeat=[]-->, belongsTo=parr
168	centromeric localization of the CPC (Gassmann et al, 2004). As, otherFeat=[]-->, belongsTo=parr
169	our results suggest that targeting of the Survivin/INCENP/Aurora-B, otherFeat=[]-->, belongsTo=parr
170	complex is mediated by Survivin, we explored whether this, otherFeat=[]-->, belongsTo=parr
171	depended on the presence of Borealin. We depleted Borealin, otherFeat=[]-->, belongsTo=parr
172	using siRNA (Fig 3A?C) and introduced the Survivin?INCENP, otherFeat=[]-->, belongsTo=parr
173	chimaera into these cells. Interestingly, expression of this, otherFeat=[]-->, belongsTo=parr
174	chimaera in Borealin-depleted cells significantly restored CPC, otherFeat=[]-->, belongsTo=parr
175	Fig 1 | Protein?protein interactions within and stability of the chromosomal passenger complex. U2OS cells were transfected with 10 mg mock or inner, otherFeat=[]-->, belongsTo=fig_caption
176	centromere protein (INCENP) short interfering RNA (siRNA) vector combined with pBABE-puro. (A) Puromycin was added to enrich for transfected, otherFeat=[]-->, belongsTo=fig_caption
177	cells, and lysates for western blotting were prepared. (B) Transfected cells were released from a thymidine block and, at indicated time points, lysates, otherFeat=[]-->, belongsTo=fig_caption
178	were prepared for western blotting. N indicates nocodazole treatment for 24 h to enrich for mitotic cells. (C)5 mg of the indicated VSV-tagged plasmids, otherFeat=[]-->, belongsTo=fig_caption
179	was transfected in combination with the siRNA plasmids and pBABE-puro. After puromycin selection, lysates were analysed by western blotting., otherFeat=[]-->, belongsTo=fig_caption
180	(D) Similar to (C), after release from the thymidine block, cells were treated with Taxol for 18 h and collected for fluorescence-activated cell sorting, otherFeat=[]-->, belongsTo=fig_caption
181	(FACS) analysis. Mitotic percentages were determined by staining the cells with the MPM-2 antibody that specifically recognizes mitotic phospho-, otherFeat=[]-->, belongsTo=fig_caption
182	epitopes. (E) Similar to (C), 60 h after transfection, cells were collected for FACS analysis. (F) Human embryonic kidney (HEK) 293 cells were, otherFeat=[]-->, belongsTo=fig_caption
183	transfected with 5 mg of the indicated green fluorescent protein (GFP)-tagged constructs. After 48 h, cells were collected , immunoprecipitations were, otherFeat=[]-->, belongsTo=fig_caption
184	carried out with anti-GFP and samples were analysed by western blotting. The presence of GFP protein (arrowhead) is due to the presence of an, otherFeat=[]-->, belongsTo=fig_caption
185	additional transcription site in the plasmids used. (G) Similar to (C). Note that INCENP1?47 contains both an amino-terminal VSV tag and a carboxy-, otherFeat=[]-->, belongsTo=fig_caption
186	terminal GFP tag. The asterisk denotes an aspecific band recognized by the anti-VSV monoclonal antibody. (H) Similar to (C). At 24 h after puromycin, otherFeat=[]-->, belongsTo=fig_caption
187	selection, nocodazole was added to the cells for 18 h. Mitotic cells were obtained by shake-off and lysates were analysed by western blotting., otherFeat=[]-->, belongsTo=fig_caption
188	c, otherFeat=[]-->, belongsTo=fig_caption
189	CPC targeting mediated by Survivin, otherFeat=[]-->, belongsTo=nota_cab_pie
190	G. Vader et al, otherFeat=[]-->, belongsTo=nota_cab_pie
191	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
192	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
193	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
194	86, otherFeat=[]-->, belongsTo=nota_cab_pie
195	function, as judged by restoration of mitotic arrest in response to, otherFeat=[]-->, belongsTo=parr
196	Taxol and a reduction in the percentage of polyploid cells that, otherFeat=[]-->, belongsTo=parr
197	were observed after depletion of Borealin (Fig 3D,E). Expression of, otherFeat=[]-->, belongsTo=parr
198	neither Survivin nor INCENP individually was sufficient to restore, otherFeat=[]-->, belongsTo=parr
199	CPC function in Borealin-depleted cells (Fig 3E; data not shown)., otherFeat=[]-->, belongsTo=parr
200	In line with these functional data, the chimaera was found to, otherFeat=[]-->, belongsTo=parr
201	localize to centromeres and the midbody in Borealin-depleted, otherFeat=[]-->, belongsTo=parr
202	cells (Fig 3F) and to restore Aurora-B localization (supplementary, otherFeat=[]-->, belongsTo=parr
203	0 9 12 15 18 24 N, otherFeat=[]-->, belongsTo=parrnote
204	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
205	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
206	0 9 12 15 18 24 N, otherFeat=[]-->, belongsTo=parrnote
207	+, otherFeat=[]-->, belongsTo=parrnote
208	INCENP1-47-GFP, otherFeat=[]-->, belongsTo=parrnote
209	-, otherFeat=[]-->, belongsTo=parrnote
210	-, otherFeat=[]-->, belongsTo=parrnote
211	-, otherFeat=[]-->, belongsTo=parrnote
212	INCENP47-920-sil-GFP, otherFeat=[]-->, belongsTo=parrnote
213	+, otherFeat=[]-->, belongsTo=parrnote
214	-, otherFeat=[]-->, belongsTo=parrnote
215	-, otherFeat=[]-->, belongsTo=parrnote
216	-, otherFeat=[]-->, belongsTo=parrnote
217	INCENP-sil-GFP, otherFeat=[]-->, belongsTo=parrnote
218	+, otherFeat=[]-->, belongsTo=parrnote
219	-, otherFeat=[]-->, belongsTo=parrnote
220	-, otherFeat=[]-->, belongsTo=parrnote
221	-, otherFeat=[]-->, belongsTo=parrnote
222	Empty vector, otherFeat=[]-->, belongsTo=parrnote
223	+, otherFeat=[]-->, belongsTo=parrnote
224	-, otherFeat=[]-->, belongsTo=parrnote
225	-, otherFeat=[]-->, belongsTo=parrnote
226	-, otherFeat=[]-->, belongsTo=parrnote
227	Total lysate, otherFeat=[]-->, belongsTo=parrnote
228	IP: Anti-GFP, otherFeat=[]-->, belongsTo=parrnote
229	+, otherFeat=[]-->, belongsTo=parrnote
230	-, otherFeat=[]-->, belongsTo=parrnote
231	-, otherFeat=[]-->, belongsTo=parrnote
232	-, otherFeat=[]-->, belongsTo=parrnote
233	+ -, otherFeat=[]-->, belongsTo=parrnote
234	-, otherFeat=[]-->, belongsTo=parrnote
235	- +, otherFeat=[]-->, belongsTo=parrnote
236	-, otherFeat=[]-->, belongsTo=parrnote
237	-, otherFeat=[]-->, belongsTo=parrnote
238	-, otherFeat=[]-->, belongsTo=parrnote
239	+, otherFeat=[]-->, belongsTo=parrnote
240	-, otherFeat=[]-->, belongsTo=parrnote
241	-, otherFeat=[]-->, belongsTo=parrnote
242	-, otherFeat=[]-->, belongsTo=parrnote
243	Hours, otherFeat=[]-->, belongsTo=parrnote
244	+, otherFeat=[]-->, belongsTo=parrnote
245	+, otherFeat=[]-->, belongsTo=parrnote
246	-, otherFeat=[]-->, belongsTo=parrnote
247	++, otherFeat=[]-->, belongsTo=parrnote
248	++, otherFeat=[]-->, belongsTo=parrnote
249	+, otherFeat=[]-->, belongsTo=parrnote
250	-, otherFeat=[]-->, belongsTo=parrnote
251	--, otherFeat=[]-->, belongsTo=parrnote
252	++, otherFeat=[]-->, belongsTo=parrnote
253	--, otherFeat=[]-->, belongsTo=parrnote
254	+, otherFeat=[]-->, belongsTo=parrnote
255	-, otherFeat=[]-->, belongsTo=parrnote
256	-, otherFeat=[]-->, belongsTo=parrnote
257	-, otherFeat=[]-->, belongsTo=parrnote
258	-, otherFeat=[]-->, belongsTo=parrnote
259	-, otherFeat=[]-->, belongsTo=parrnote
260	-, otherFeat=[]-->, belongsTo=parrnote
261	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
262	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
263	Empty vector, otherFeat=[]-->, belongsTo=parrnote
264	VSV -INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
265	VSV-INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
266	Mitotic shake-off, otherFeat=[]-->, belongsTo=parrnote
267	(nocodazole), otherFeat=[]-->, belongsTo=parrnote
268	+, otherFeat=[]-->, belongsTo=parrnote
269	+, otherFeat=[]-->, belongsTo=parrnote
270	-, otherFeat=[]-->, belongsTo=parrnote
271	++, otherFeat=[]-->, belongsTo=parrnote
272	++, otherFeat=[]-->, belongsTo=parrnote
273	+, otherFeat=[]-->, belongsTo=parrnote
274	-, otherFeat=[]-->, belongsTo=parrnote
275	--, otherFeat=[]-->, belongsTo=parrnote
276	++, otherFeat=[]-->, belongsTo=parrnote
277	--, otherFeat=[]-->, belongsTo=parrnote
278	+, otherFeat=[]-->, belongsTo=parrnote
279	-, otherFeat=[]-->, belongsTo=parrnote
280	-, otherFeat=[]-->, belongsTo=parrnote
281	-, otherFeat=[]-->, belongsTo=parrnote
282	-, otherFeat=[]-->, belongsTo=parrnote
283	-, otherFeat=[]-->, belongsTo=parrnote
284	-, otherFeat=[]-->, belongsTo=parrnote
285	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
286	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
287	Empty vector, otherFeat=[]-->, belongsTo=parrnote
288	VSV-INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
289	VSV-INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
290	+, otherFeat=[]-->, belongsTo=parrnote
291	+, otherFeat=[]-->, belongsTo=parrnote
292	-, otherFeat=[]-->, belongsTo=parrnote
293	-, otherFeat=[]-->, belongsTo=parrnote
294	-, otherFeat=[]-->, belongsTo=parrnote
295	-, otherFeat=[]-->, belongsTo=parrnote
296	--, otherFeat=[]-->, belongsTo=parrnote
297	-, otherFeat=[]-->, belongsTo=parrnote
298	-, otherFeat=[]-->, belongsTo=parrnote
299	VSV-INCENP1-47-GFP, otherFeat=[]-->, belongsTo=parrnote
300	-, otherFeat=[]-->, belongsTo=parrnote
301	INCENPsiRNA, otherFeat=[]-->, belongsTo=parrnote
302	Mock, otherFeat=[]-->, belongsTo=parrnote
303	siRNA, otherFeat=[]-->, belongsTo=parrnote
304	GFP, otherFeat=[]-->, belongsTo=parrnote
305	Mock, otherFeat=[]-->, belongsTo=parrnote
306	siRNA, otherFeat=[]-->, belongsTo=parrnote
307	INCENP, otherFeat=[]-->, belongsTo=parrnote
308	siRNA, otherFeat=[]-->, belongsTo=parrnote
309	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
310	Anti-Cdk6, otherFeat=[]-->, belongsTo=parrnote
311	VSV-INCENP, otherFeat=[]-->, belongsTo=parrnote
312	VSV-INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
313	+, otherFeat=[]-->, belongsTo=parrnote
314	-, otherFeat=[]-->, belongsTo=parrnote
315	+, otherFeat=[]-->, belongsTo=parrnote
316	-, otherFeat=[]-->, belongsTo=parrnote
317	-, otherFeat=[]-->, belongsTo=parrnote
318	+, otherFeat=[]-->, belongsTo=parrnote
319	-, otherFeat=[]-->, belongsTo=parrnote
320	+, otherFeat=[]-->, belongsTo=parrnote
321	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
322	+ VSV-INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
323	4n, otherFeat=[]-->, belongsTo=parrnote
324	2n, otherFeat=[]-->, belongsTo=parrnote
325	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
326	+ VSV-INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
327	8n, otherFeat=[]-->, belongsTo=parrnote
328	VSV, otherFeat=[]-->, belongsTo=parrnote
329	-, otherFeat=[]-->, belongsTo=parrnote
330	INCENP-sil VSV, otherFeat=[]-->, belongsTo=parrnote
331	-, otherFeat=[]-->, belongsTo=parrnote
332	INCENP, otherFeat=[]-->, belongsTo=parrnote
333	47-920, otherFeat=[]-->, belongsTo=parrnote
334	-sil, otherFeat=[]-->, belongsTo=parrnote
335	Emptyvector, otherFeat=[]-->, belongsTo=parrnote
336	Mitotic, otherFeat=[]-->, belongsTo=parrnote
337	cells, otherFeat=[]-->, belongsTo=parrnote
338	(%), otherFeat=[]-->, belongsTo=parrnote
339	40, otherFeat=[]-->, belongsTo=parrnote
340	20, otherFeat=[]-->, belongsTo=parrnote
341	0, otherFeat=[]-->, belongsTo=parrnote
342	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
343	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
344	+Taxol, otherFeat=[]-->, belongsTo=parrnote
345	10.1%, otherFeat=[]-->, belongsTo=parrnote
346	25.8%, otherFeat=[]-->, belongsTo=parrnote
347	12.0%, otherFeat=[]-->, belongsTo=parrnote
348	19.3%, otherFeat=[]-->, belongsTo=parrnote
349	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
350	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
351	12 3 4, otherFeat=[]-->, belongsTo=parrnote
352	5 6, otherFeat=[]-->, belongsTo=parrnote
353	7 8, otherFeat=[]-->, belongsTo=parrnote
354	12, otherFeat=[]-->, belongsTo=parrnote
355	3, otherFeat=[]-->, belongsTo=parrnote
356	4, otherFeat=[]-->, belongsTo=parrnote
357	12345, otherFeat=[]-->, belongsTo=parrnote
358	Anti-INCENP, otherFeat=[]-->, belongsTo=parrnote
359	Anti-Aurora-B, otherFeat=[]-->, belongsTo=parrnote
360	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
361	Anti-Aurora-B, otherFeat=[]-->, belongsTo=parrnote
362	Anti-GFP, otherFeat=[]-->, belongsTo=parrnote
363	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
364	Anti-Aurora-B, otherFeat=[]-->, belongsTo=parrnote
365	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
366	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
367	Anti-Cdk4, otherFeat=[]-->, belongsTo=parrnote
368	Anti-Aurora-B, otherFeat=[]-->, belongsTo=parrnote
369	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
370	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
371	Anti-phospho-Histone H3, otherFeat=[]-->, belongsTo=parrnote
372	Anti-Cdk4, otherFeat=[]-->, belongsTo=parrnote
373	Anti-Aurora-B, otherFeat=[]-->, belongsTo=parrnote
374	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
375	Anti-Actin, otherFeat=[]-->, belongsTo=parrnote
376	Anti-Cdk4, otherFeat=[]-->, belongsTo=parrnote
377	GH, otherFeat=[]-->, belongsTo=parrnote
378	EF, otherFeat=[]-->, belongsTo=parrnote
379	CD, otherFeat=[]-->, belongsTo=parrnote
380	A, otherFeat=[]-->, belongsTo=parrnote
381	B, otherFeat=[]-->, belongsTo=parrnote
382	CPC targeting mediated by Survivin, otherFeat=[]-->, belongsTo=nota_cab_pie
383	G. Vader et al, otherFeat=[]-->, belongsTo=nota_cab_pie
384	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
385	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
386	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
387	87, otherFeat=[]-->, belongsTo=nota_cab_pie
388	Fig 3 online). However, it should be noted that the association, otherFeat=[]-->, belongsTo=parr
389	of the Survivin?INCENP chimaera to centromeres and the central, otherFeat=[]-->, belongsTo=parr
390	spindle was affected in Borealin-depleted cells. Instead of, otherFeat=[]-->, belongsTo=parr
391	concentrating at centromeres and central spindle, a significant, otherFeat=[]-->, belongsTo=parr
392	portion of the chimaera localized to chromatin throughout mitosis, otherFeat=[]-->, belongsTo=parr
393	(Fig 3F, lower panel; data not shown). Taken together, these data, otherFeat=[]-->, belongsTo=parr
394	show that the Survivin?INCENP chimaera is sufficient to restore a, otherFeat=[]-->, belongsTo=parr
395	functional CPC in Borealin-depleted cells, and that Borealin might, otherFeat=[]-->, belongsTo=parr
396	facilitate dissociation of the CPC from chromatin or enhance its, otherFeat=[]-->, belongsTo=parr
397	concentration at centromeres and the central spindle., otherFeat=[]-->, belongsTo=parr
398	Borealin facilitates Survivin?INCENP interaction, otherFeat=[]-->, belongsTo=title
399	As Borealin is essential for localization of INCENP and Survivin, otherFeat=[]-->, belongsTo=parr
400	(Gassmann et al, 2004), but not for a chimaera in which these, otherFeat=[]-->, belongsTo=parr
401	two proteins are covalently linked, we reasoned that Borealin, otherFeat=[]-->, belongsTo=parr
402	might be important for the Survivin?INCENP interaction. To, otherFeat=[]-->, belongsTo=parr
403	further investigate the potential role of Borealin in regulating the, otherFeat=[]-->, belongsTo=parr
404	Survivin?INCENP interaction, immunoprecipitations were carried, otherFeat=[]-->, belongsTo=parr
405	out with GFP?INCENP from Borealin-depleted cells expressing, otherFeat=[]-->, belongsTo=parr
406	either exogenous Survivin or Borealin. In control cells, both, otherFeat=[]-->, belongsTo=parr
407	endogenous and exogenous Survivin could interact with INCENP, otherFeat=[]-->, belongsTo=parr
408	(Fig 3H, lanes 1?3). In contrast, in the absence of endogenous, otherFeat=[]-->, belongsTo=parr
409	Borealin, neither endogenous nor exogenous Survivin was able to, otherFeat=[]-->, belongsTo=parr
410	interact with INCENP (Fig 3H, lanes 4,5). As depletion of Borealin, otherFeat=[]-->, belongsTo=parr
411	also reduced Survivin levels, this probably explains why no, otherFeat=[]-->, belongsTo=parr
412	endogenous Survivin was found to interact with INCENP (Fig 3H,, otherFeat=[]-->, belongsTo=parr
413	lanes 4,10; Gassmann et al, 2004; Sampath et al, 2004). Indeed,, otherFeat=[]-->, belongsTo=parr
414	after expression of exogenous, RNAi-resistant Borealin, endo-, otherFeat=[]-->, belongsTo=parr
415	genous Survivin levels and Survivin interaction with INCENP were, otherFeat=[]-->, belongsTo=parr
416	restored (Fig 3E, lanes 6,12). Importantly, although exogenous, otherFeat=[]-->, belongsTo=parr
417	Survivin levels were not affected by Borealin depletion, Survivin, otherFeat=[]-->, belongsTo=parr
418	failed to interact with INCENP, indicating that, in vivo, an efficient, otherFeat=[]-->, belongsTo=parr
419	interaction of Survivin and INCENP is dependent on the presence, otherFeat=[]-->, belongsTo=parr
420	of Borealin (Fig 3E, lanes 5,11). The observation that Borealin can, otherFeat=[]-->, belongsTo=parr
421	interact with INCENP that lacks the Survivin interaction domain, otherFeat=[]-->, belongsTo=parr
422	(INCENP47?920; Fig 3G, lanes 6,8; supplementary Fig 3 online), in, otherFeat=[]-->, belongsTo=parr
423	combination with previous in vitro data that have shown the direct, otherFeat=[]-->, belongsTo=parr
424	interaction of Survivin and INCENP (Wheatley et al, 2001;, otherFeat=[]-->, belongsTo=parr
425	Romano et al, 2003), rules out a direct bridging function for, otherFeat=[]-->, belongsTo=parr
426	Borealin between Survivin and the centromere-targeting domain, otherFeat=[]-->, belongsTo=parr
427	of INCENP. Hence, similar to CSC-1, its distant homologue in, otherFeat=[]-->, belongsTo=parr
428	Caenorhabditis elegans (Romano et al, 2003), human Borealin has, otherFeat=[]-->, belongsTo=parr
429	a role in facilitating Survivin?INCENP interaction. By generating, otherFeat=[]-->, belongsTo=parr
430	the Survivin?INCENP chimaera, we have circumvented the, otherFeat=[]-->, belongsTo=parr
431	necessity for Borealin in stabilizing Survivin?INCENP interaction,, otherFeat=[]-->, belongsTo=parr
432	which probably explains why a functional CPC can be directed to, otherFeat=[]-->, belongsTo=parr
433	the right mitotic structures in Borealin-depleted cells., otherFeat=[]-->, belongsTo=parr
434	Conclusion, otherFeat=[]-->, belongsTo=title
435	Given the essential functions of Aurora-B during mitosis, explaining, otherFeat=[]-->, belongsTo=parr
436	the mechanisms behind its dynamic localization is of great, otherFeat=[]-->, belongsTo=parr
437	importance. The observation that depletion of one CPC component, otherFeat=[]-->, belongsTo=parr
438	by siRNA results in the destabilization of the entire complex, otherFeat=[]-->, belongsTo=parr
439	instigates the need for more complex experiments to unravel the, otherFeat=[]-->, belongsTo=parr
440	functions of single CPC components. Using RNAi-complementation,, otherFeat=[]-->, belongsTo=parr
441	we have provided evidence that an important function of Survivin, otherFeat=[]-->, belongsTo=parr
442	is to serve as an interface between the centromere/central spindle, otherFeat=[]-->, belongsTo=parr
443	and the CPC, whereas Borealin facilitates the interaction of Survivin, otherFeat=[]-->, belongsTo=parr
444	and INCENP (Fig 4B). Importantly, the targeting function of Survivin, otherFeat=[]-->, belongsTo=parr
445	was revealed only when Survivin was fused to INCENP lacking, otherFeat=[]-->, belongsTo=parr
446	its centromere-targeting domain. This suggests that although, otherFeat=[]-->, belongsTo=parr
447	Survivin can mediate localization of the CPC to its proper mitotic, otherFeat=[]-->, belongsTo=parr
448	structures, the other CPC components are probably needed to, otherFeat=[]-->, belongsTo=parr
449	obtain a stable association (possibly by affecting the on/off rate of, otherFeat=[]-->, belongsTo=parr
450	the complex). Identifying the Survivin interaction partners at the, otherFeat=[]-->, belongsTo=parr
451	centromere and midbody will be an important next step in, otherFeat=[]-->, belongsTo=parr
452	understanding the regulation of the CPC., otherFeat=[]-->, belongsTo=parr
453	METHODS, otherFeat=[]-->, belongsTo=title
454	Antibodies and reagents. The following antibodies were used:, otherFeat=[]-->, belongsTo=parr
455	rabbit anti-INCENP and rabbit anti-Aurora-B (Abcam, Cambridge,, otherFeat=[]-->, belongsTo=parr
456	UK), rabbit anti-Survivin (R&D, Minneapolis, MN, USA), rabbit, otherFeat=[]-->, belongsTo=parr
457	anti-Borealin (gift from Dr H. Funabiki), mouse anti-Aurora-B, otherFeat=[]-->, belongsTo=parr
458	(Transduction, Lexington, KE, USA), mouse anti-VSV and mouse, otherFeat=[]-->, belongsTo=parr
459	anti-a-Tubulin (Sigma, St Louis, MO, USA), rabbit anti-Cdk6, otherFeat=[]-->, belongsTo=parr
460	and rabbit anti-Cdk4 (Santa Cruz, Santa Cruz, CA, USA), sheep, otherFeat=[]-->, belongsTo=parr
461	anti-BubR1 (gift from Dr S. Taylor), rabbit anti-GFP (gift from, otherFeat=[]-->, belongsTo=parr
462	Dr J. Neefjes), rabbit anti-phospho-Ser10-histone-H3 and mouse, otherFeat=[]-->, belongsTo=parr
463	anti-MPM-2 (Upstate, Charlotsville, VA, USA), CREST antiserum, otherFeat=[]-->, belongsTo=parr
464	(Cortex Biochem, San Leandro, CA, USA), peroxidase-conjugated, otherFeat=[]-->, belongsTo=parr
465	goat anti-rabbit and peroxidase-conjugated goat anti-mouse, otherFeat=[]-->, belongsTo=parr
466	(Dako, Glostrup, Denmark) and donkey anti-mouse/Cy5 (Jackson,, otherFeat=[]-->, belongsTo=parr
467	West Grove, PA, USA), goat anti-rabbit/Alexa-568, goat anti-, otherFeat=[]-->, belongsTo=parr
468	rabbit/Alexa-633, goat anti-mouse/Alexa-568 and donkey anti-, otherFeat=[]-->, belongsTo=parr
469	sheep/Alexa-568 (Molecular Probes, Eugene, OR, USA). Reagents, otherFeat=[]-->, belongsTo=parr
470	were from Sigma unless stated otherwise., otherFeat=[]-->, belongsTo=parr
471	Plasmids. RNAi vectors were generated as described (Lens et al,, otherFeat=[]-->, belongsTo=parr
472	2003), and were based on 50-TGACACGGAGATTGCCAAC-30, otherFeat=[]-->, belongsTo=parr
473	(INCENP) and 50-CAGCTGACCTGGATATCAC-30 (Borealin)., otherFeat=[]-->, belongsTo=parr
474	Human INCENP and Borealin were PCR amplified from a T-cell, otherFeat=[]-->, belongsTo=parr
475	Fig 2 | Survivin is a determining factor for mitotic localization of the chromosome passenger complex. (A) U2OS cells were transfected with the, otherFeat=[]-->, belongsTo=fig_caption
476	indicated plasmids. Puromycin was added to enrich for transfected cells. Lysates were analysed by western blotting. (B,C) Cells were transfected with, otherFeat=[]-->, belongsTo=fig_caption
477	the indicated short interfering RNA (siRNA) plasmids combined with the indicated VSV-tagged constructs. (B) After release from a thymidine block,, otherFeat=[]-->, belongsTo=fig_caption
478	cells were treated with Taxol and collected for fluorescence-activated cell sorting (FACS) analysis 18 h later. Mitotic percentages were determinedby, otherFeat=[]-->, belongsTo=fig_caption
479	MPM-2 staining. (C) Cells were collected for FACS analysis 60 h after transfection. (D,E) Cells grown on coverslips were transfected with mock or, otherFeat=[]-->, belongsTo=fig_caption
480	inner centromere protein (INCENP)/Survivin siRNA plasmids, VSV?Survivin?INCENP47?920-sil and H2B?GFP (green fluorescent protein)., otherFeat=[]-->, belongsTo=fig_caption
481	Immunostainings were carried out with the indicated antibodies. DNA (H2B?GFP) is depicted in blue. (F) Cells grown on coverslips were transfected, otherFeat=[]-->, belongsTo=fig_caption
482	with the indicated plasmids and H2B?GFP. Cells were stained with anti-Aurora-B, and the percentage of H2B?GFP-positive prometaphase cells in, otherFeat=[]-->, belongsTo=fig_caption
483	which Aurora-B localized on the centromeres was scored. The number of cells counted per condition is indicated. (G) Time-lapse images of a, otherFeat=[]-->, belongsTo=fig_caption
484	representative cell transfected with INCENP siRNA plasmid, Survivin?INCENP47?920-sil?GFP and H2B?diHcRed. Differential interference contrast, otherFeat=[]-->, belongsTo=fig_caption
485	(DIC), GFP and diHcRed signals are shown. (H) Time-lapse imaging of cells transfected with mock (upper panel) or Survivin siRNA plasmid (lower, otherFeat=[]-->, belongsTo=fig_caption
486	panel), INCENP1?47-GFP and H2B?diHcRed. Scale bars, 5 mm., otherFeat=[]-->, belongsTo=fig_caption
487	c, otherFeat=[]-->, belongsTo=fig_caption
488	CPC targeting mediated by Survivin, otherFeat=[]-->, belongsTo=nota_cab_pie
489	G. Vader et al, otherFeat=[]-->, belongsTo=nota_cab_pie
490	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
491	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
492	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
493	88, otherFeat=[]-->, belongsTo=nota_cab_pie
494	0:00, otherFeat=[]-->, belongsTo=parrnote
495	0:18, otherFeat=[]-->, belongsTo=parrnote
496	0:24, otherFeat=[]-->, belongsTo=parrnote
497	0:30, otherFeat=[]-->, belongsTo=parrnote
498	0:42, otherFeat=[]-->, belongsTo=parrnote
499	0:48, otherFeat=[]-->, belongsTo=parrnote
500	2:18, otherFeat=[]-->, belongsTo=parrnote
501	GFP, otherFeat=[]-->, belongsTo=parrnote
502	H2B-, otherFeat=[]-->, belongsTo=parrnote
503	diHcRed, otherFeat=[]-->, belongsTo=parrnote
504	DIC, otherFeat=[]-->, belongsTo=parrnote
505	INCENP siRNA +, otherFeat=[]-->, belongsTo=parrnote
506	VSV-Survivin-INCENP47-920-GFP, otherFeat=[]-->, belongsTo=parrnote
507	GFP, otherFeat=[]-->, belongsTo=parrnote
508	DIC, otherFeat=[]-->, belongsTo=parrnote
509	H2B -, otherFeat=[]-->, belongsTo=parrnote
510	diHcRed, otherFeat=[]-->, belongsTo=parrnote
511	INCENP1-47 - GFP, otherFeat=[]-->, belongsTo=parrnote
512	+, otherFeat=[]-->, belongsTo=parrnote
513	+, otherFeat=[]-->, belongsTo=parrnote
514	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
515	Anti-Cdk4, otherFeat=[]-->, belongsTo=parrnote
516	-, otherFeat=[]-->, belongsTo=parrnote
517	-, otherFeat=[]-->, belongsTo=parrnote
518	VSV-INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
519	VSV-Survivin-, otherFeat=[]-->, belongsTo=parrnote
520	INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
521	Mock, otherFeat=[]-->, belongsTo=parrnote
522	siRNA, otherFeat=[]-->, belongsTo=parrnote
523	Survivin, otherFeat=[]-->, belongsTo=parrnote
524	siRNA, otherFeat=[]-->, belongsTo=parrnote
525	INCENP, otherFeat=[]-->, belongsTo=parrnote
526	siRNA, otherFeat=[]-->, belongsTo=parrnote
527	H2B-, otherFeat=[]-->, belongsTo=parrnote
528	GFP, otherFeat=[]-->, belongsTo=parrnote
529	Merge, otherFeat=[]-->, belongsTo=parrnote
530	VSV-, otherFeat=[]-->, belongsTo=parrnote
531	Survivin -, otherFeat=[]-->, belongsTo=parrnote
532	INCENP, otherFeat=[]-->, belongsTo=parrnote
533	47 -920-sil, otherFeat=[]-->, belongsTo=parrnote
534	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
535	Survivin siRNA +, otherFeat=[]-->, belongsTo=parrnote
536	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
537	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
538	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
539	Survivin siRNA, otherFeat=[]-->, belongsTo=parrnote
540	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
541	Survivin siRNA +, otherFeat=[]-->, belongsTo=parrnote
542	INCENP siRNA, otherFeat=[]-->, belongsTo=parrnote
543	Empty vector, otherFeat=[]-->, belongsTo=parrnote
544	VSV-Survivin-, otherFeat=[]-->, belongsTo=parrnote
545	INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
546	4n, otherFeat=[]-->, belongsTo=parrnote
547	2n, otherFeat=[]-->, belongsTo=parrnote
548	8n, otherFeat=[]-->, belongsTo=parrnote
549	4n, otherFeat=[]-->, belongsTo=parrnote
550	2 n, otherFeat=[]-->, belongsTo=parrnote
551	8n, otherFeat=[]-->, belongsTo=parrnote
552	5.7%, otherFeat=[]-->, belongsTo=parrnote
553	18.5%, otherFeat=[]-->, belongsTo=parrnote
554	21.9%, otherFeat=[]-->, belongsTo=parrnote
555	6.4%, otherFeat=[]-->, belongsTo=parrnote
556	11.6%, otherFeat=[]-->, belongsTo=parrnote
557	10.2%, otherFeat=[]-->, belongsTo=parrnote
558	7.7%, otherFeat=[]-->, belongsTo=parrnote
559	4 n, otherFeat=[]-->, belongsTo=parrnote
560	2n, otherFeat=[]-->, belongsTo=parrnote
561	8n, otherFeat=[]-->, belongsTo=parrnote
562	VSV-Survivin-sil, otherFeat=[]-->, belongsTo=note
563	7.8%, otherFeat=[]-->, belongsTo=parrnote
564	17.5%, otherFeat=[]-->, belongsTo=parrnote
565	10.4%, otherFeat=[]-->, belongsTo=parrnote
566	18.3%, otherFeat=[]-->, belongsTo=parrnote
567	Empty vector, otherFeat=[]-->, belongsTo=parrnote
568	VSV-INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
569	VSV-Surv-INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
570	VSV-Survivin-sil, otherFeat=[]-->, belongsTo=parrnote
571	Mitotic, otherFeat=[]-->, belongsTo=parrnote
572	cells, otherFeat=[]-->, belongsTo=parrnote
573	(%) 40, otherFeat=[]-->, belongsTo=parrnote
574	20, otherFeat=[]-->, belongsTo=parrnote
575	0, otherFeat=[]-->, belongsTo=parrnote
576	60, otherFeat=[]-->, belongsTo=parrnote
577	Survivin, otherFeat=[]-->, belongsTo=parrnote
578	siRNA, otherFeat=[]-->, belongsTo=parrnote
579	INCENP, otherFeat=[]-->, belongsTo=parrnote
580	siRNA, otherFeat=[]-->, belongsTo=parrnote
581	Mock, otherFeat=[]-->, belongsTo=parrnote
582	siRNA, otherFeat=[]-->, belongsTo=parrnote
583	Survivin, otherFeat=[]-->, belongsTo=parrnote
584	siRNA, otherFeat=[]-->, belongsTo=parrnote
585	+ INCEN, otherFeat=[]-->, belongsTo=parrnote
586	PsiRNA, otherFeat=[]-->, belongsTo=parrnote
587	Merge, otherFeat=[]-->, belongsTo=parrnote
588	DNA, otherFeat=[]-->, belongsTo=parrnote
589	INCENP siRNA +, otherFeat=[]-->, belongsTo=parrnote
590	VSV-Survivin -, otherFeat=[]-->, belongsTo=parrnote
591	INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
592	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
593	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
594	Anti-Aurora-B, otherFeat=[]-->, belongsTo=parrnote
595	Aur, otherFeat=[]-->, belongsTo=parrnote
596	-B-positive, otherFeat=[]-->, belongsTo=parrnote
597	cells, otherFeat=[]-->, belongsTo=parrnote
598	(%), otherFeat=[]-->, belongsTo=parrnote
599	0, otherFeat=[]-->, belongsTo=parrnote
600	40, otherFeat=[]-->, belongsTo=parrnote
601	60, otherFeat=[]-->, belongsTo=parrnote
602	80, otherFeat=[]-->, belongsTo=parrnote
603	100, otherFeat=[]-->, belongsTo=parrnote
604	20, otherFeat=[]-->, belongsTo=parrnote
605	Mock, otherFeat=[]-->, belongsTo=parrnote
606	siRNA, otherFeat=[]-->, belongsTo=parrnote
607	INCENP, otherFeat=[]-->, belongsTo=parrnote
608	siRNA, otherFeat=[]-->, belongsTo=parrnote
609	INCENP, otherFeat=[]-->, belongsTo=parrnote
610	siRNA, otherFeat=[]-->, belongsTo=parrnote
611	+, otherFeat=[]-->, belongsTo=parrnote
612	VSV, otherFeat=[]-->, belongsTo=parrnote
613	-INCENP-silINCENP, otherFeat=[]-->, belongsTo=parrnote
614	siRNA, otherFeat=[]-->, belongsTo=parrnote
615	+, otherFeat=[]-->, belongsTo=parrnote
616	VSV, otherFeat=[]-->, belongsTo=parrnote
617	-Surv, otherFeat=[]-->, belongsTo=parrnote
618	-INCENP, otherFeat=[]-->, belongsTo=parrnote
619	47- 920, otherFeat=[]-->, belongsTo=parrnote
620	-sil, otherFeat=[]-->, belongsTo=parrnote
621	n=47, otherFeat=[]-->, belongsTo=parrnote
622	n=96, otherFeat=[]-->, belongsTo=parrnote
623	n=87, otherFeat=[]-->, belongsTo=parrnote
624	n=62, otherFeat=[]-->, belongsTo=parrnote
625	+ Taxol, otherFeat=[]-->, belongsTo=parrnote
626	G, otherFeat=[]-->, belongsTo=parrnote
627	H, otherFeat=[]-->, belongsTo=parrnote
628	F, otherFeat=[]-->, belongsTo=parrnote
629	D, otherFeat=[]-->, belongsTo=parrnote
630	C, otherFeat=[]-->, belongsTo=parrnote
631	E, otherFeat=[]-->, belongsTo=parrnote
632	AB, otherFeat=[]-->, belongsTo=parrnote
633	23.9%, otherFeat=[]-->, belongsTo=parrnote
634	CPC targeting mediated by Survivin, otherFeat=[]-->, belongsTo=nota_cab_pie
635	G. Vader et al, otherFeat=[]-->, belongsTo=nota_cab_pie
636	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
637	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
638	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
639	89, otherFeat=[]-->, belongsTo=nota_cab_pie
640	Borealin siRNA, otherFeat=[]-->, belongsTo=parrnote
641	+, otherFeat=[]-->, belongsTo=parrnote
642	-, otherFeat=[]-->, belongsTo=parrnote
643	VSV?Borealin, otherFeat=[]-->, belongsTo=parrnote
644	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
645	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
646	Anti-Tubulin, otherFeat=[]-->, belongsTo=parrnote
647	+, otherFeat=[]-->, belongsTo=parrnote
648	-, otherFeat=[]-->, belongsTo=parrnote
649	Mock siRNA, otherFeat=[]-->, belongsTo=parrnote
650	+, otherFeat=[]-->, belongsTo=parrnote
651	+, otherFeat=[]-->, belongsTo=parrnote
652	4 n, otherFeat=[]-->, belongsTo=parrnote
653	2n, otherFeat=[]-->, belongsTo=parrnote
654	8 n, otherFeat=[]-->, belongsTo=parrnote
655	4n, otherFeat=[]-->, belongsTo=parrnote
656	2n, otherFeat=[]-->, belongsTo=parrnote
657	8 n, otherFeat=[]-->, belongsTo=parrnote
658	Mock, otherFeat=[]-->, belongsTo=parrnote
659	siRNA, otherFeat=[]-->, belongsTo=parrnote
660	Borealin, otherFeat=[]-->, belongsTo=parrnote
661	siRNA, otherFeat=[]-->, belongsTo=parrnote
662	Empty vector, otherFeat=[]-->, belongsTo=parrnote
663	VSV-Survivin-, otherFeat=[]-->, belongsTo=parrnote
664	INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
665	4.3%, otherFeat=[]-->, belongsTo=parrnote
666	8.7%, otherFeat=[]-->, belongsTo=parrnote
667	9.4%, otherFeat=[]-->, belongsTo=parrnote
668	26.7%, otherFeat=[]-->, belongsTo=parrnote
669	D, otherFeat=[]-->, belongsTo=parrnote
670	F, otherFeat=[]-->, belongsTo=parrnote
671	E, otherFeat=[]-->, belongsTo=parrnote
672	40, otherFeat=[]-->, belongsTo=parrnote
673	20, otherFeat=[]-->, belongsTo=parrnote
674	0, otherFeat=[]-->, belongsTo=parrnote
675	INCENP, otherFeat=[]-->, belongsTo=parrnote
676	siRNA, otherFeat=[]-->, belongsTo=parrnote
677	Mock, otherFeat=[]-->, belongsTo=parrnote
678	siRNA, otherFeat=[]-->, belongsTo=parrnote
679	Bor, otherFeat=[]-->, belongsTo=parrnote
680	ealin, otherFeat=[]-->, belongsTo=parrnote
681	siRNA, otherFeat=[]-->, belongsTo=parrnote
682	+Taxol, otherFeat=[]-->, belongsTo=parrnote
683	Mitotic, otherFeat=[]-->, belongsTo=parrnote
684	cells, otherFeat=[]-->, belongsTo=parrnote
685	(%), otherFeat=[]-->, belongsTo=parrnote
686	Empty vector, otherFeat=[]-->, belongsTo=parrnote
687	VSV-INCENP-sil, otherFeat=[]-->, belongsTo=parrnote
688	VSV-Surv-INCENP47-920-sil, otherFeat=[]-->, belongsTo=parrnote
689	Mock, otherFeat=[]-->, belongsTo=parrnote
690	siRNA, otherFeat=[]-->, belongsTo=parrnote
691	Borealin, otherFeat=[]-->, belongsTo=parrnote
692	siRNA, otherFeat=[]-->, belongsTo=parrnote
693	Mock, otherFeat=[]-->, belongsTo=parrnote
694	siRNA, otherFeat=[]-->, belongsTo=parrnote
695	Borealin, otherFeat=[]-->, belongsTo=parrnote
696	siRNA, otherFeat=[]-->, belongsTo=parrnote
697	VSV-Survivin-sil, otherFeat=[]-->, belongsTo=parrnote
698	VSV-Borealin-sil, otherFeat=[]-->, belongsTo=parrnote
699	Anti-GFP, otherFeat=[]-->, belongsTo=parrnote
700	Anti-Survivin, otherFeat=[]-->, belongsTo=parrnote
701	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
702	(Borealin), otherFeat=[]-->, belongsTo=parrnote
703	Empty vector, otherFeat=[]-->, belongsTo=parrnote
704	+ -- + --, otherFeat=[]-->, belongsTo=parrnote
705	+ -- +, otherFeat=[]-->, belongsTo=parrnote
706	--, otherFeat=[]-->, belongsTo=parrnote
707	- + -- + -- + -- + -, otherFeat=[]-->, belongsTo=parrnote
708	-- +, otherFeat=[]-->, belongsTo=parrnote
709	-- +, otherFeat=[]-->, belongsTo=parrnote
710	-- + -- +, otherFeat=[]-->, belongsTo=parrnote
711	Exog., otherFeat=[]-->, belongsTo=parrnote
712	Endog., otherFeat=[]-->, belongsTo=parrnote
713	Total lysate, otherFeat=[]-->, belongsTo=parrnote
714	12, otherFeat=[]-->, belongsTo=parrnote
715	3 4 5 6, otherFeat=[]-->, belongsTo=parrnote
716	78 9 10 11 12, otherFeat=[]-->, belongsTo=parrnote
717	GFP-INCENP, otherFeat=[]-->, belongsTo=parrnote
718	++ + + + +, otherFeat=[]-->, belongsTo=parrnote
719	+ + + +, otherFeat=[]-->, belongsTo=parrnote
720	+ +, otherFeat=[]-->, belongsTo=parrnote
721	4 n, otherFeat=[]-->, belongsTo=parrnote
722	2n, otherFeat=[]-->, belongsTo=parrnote
723	8 n, otherFeat=[]-->, belongsTo=parrnote
724	5.4%, otherFeat=[]-->, belongsTo=parrnote
725	35.7%, otherFeat=[]-->, belongsTo=parrnote
726	6.9%, otherFeat=[]-->, belongsTo=parrnote
727	Mock, otherFeat=[]-->, belongsTo=parrnote
728	siRNA, otherFeat=[]-->, belongsTo=parrnote
729	Borealin, otherFeat=[]-->, belongsTo=parrnote
730	siRNA, otherFeat=[]-->, belongsTo=parrnote
731	Borealin, otherFeat=[]-->, belongsTo=parrnote
732	siRNA +, otherFeat=[]-->, belongsTo=parrnote
733	VSV -Borealin-sil, otherFeat=[]-->, belongsTo=parrnote
734	Mock, otherFeat=[]-->, belongsTo=parrnote
735	siRNA, otherFeat=[]-->, belongsTo=parrnote
736	Borealin, otherFeat=[]-->, belongsTo=parrnote
737	siRNA, otherFeat=[]-->, belongsTo=parrnote
738	Anti-Borealin, otherFeat=[]-->, belongsTo=parrnote
739	H2B-GFP, otherFeat=[]-->, belongsTo=parrnote
740	Merge, otherFeat=[]-->, belongsTo=parrnote
741	Mock, otherFeat=[]-->, belongsTo=parrnote
742	siRNA, otherFeat=[]-->, belongsTo=parrnote
743	Borealin, otherFeat=[]-->, belongsTo=parrnote
744	siRNA, otherFeat=[]-->, belongsTo=parrnote
745	0, otherFeat=[]-->, belongsTo=parrnote
746	20, otherFeat=[]-->, belongsTo=parrnote
747	40, otherFeat=[]-->, belongsTo=parrnote
748	60, otherFeat=[]-->, belongsTo=parrnote
749	80, otherFeat=[]-->, belongsTo=parrnote
750	100 n=18, otherFeat=[]-->, belongsTo=parrnote
751	n=127, otherFeat=[]-->, belongsTo=parrnote
752	Bor, otherFeat=[]-->, belongsTo=parrnote
753	ealin-positive cells, otherFeat=[]-->, belongsTo=parrnote
754	(%), otherFeat=[]-->, belongsTo=parrnote
755	GFP, otherFeat=[]-->, belongsTo=parrnote
756	Anti-Borealin, otherFeat=[]-->, belongsTo=parrnote
757	Anti-Borealin, otherFeat=[]-->, belongsTo=parrnote
758	CREST, otherFeat=[]-->, belongsTo=parrnote
759	Merge, otherFeat=[]-->, belongsTo=parrnote
760	GFP, otherFeat=[]-->, belongsTo=parrnote
761	DNA, otherFeat=[]-->, belongsTo=parrnote
762	Merge, otherFeat=[]-->, belongsTo=parrnote
763	Borealin siRNA +, otherFeat=[]-->, belongsTo=parrnote
764	VSV -Survivin-INCENP47-920-sil-GFP, otherFeat=[]-->, belongsTo=parrnote
765	Total lysate, otherFeat=[]-->, belongsTo=parrnote
766	IP: Anti-GFP, otherFeat=[]-->, belongsTo=parrnote
767	IP: Anti-GFP, otherFeat=[]-->, belongsTo=parrnote
768	GFP-INCENP, otherFeat=[]-->, belongsTo=parrnote
769	-, otherFeat=[]-->, belongsTo=parrnote
770	+, otherFeat=[]-->, belongsTo=parrnote
771	+, otherFeat=[]-->, belongsTo=parrnote
772	+, otherFeat=[]-->, belongsTo=parrnote
773	--, otherFeat=[]-->, belongsTo=parrnote
774	++ -, otherFeat=[]-->, belongsTo=parrnote
775	-, otherFeat=[]-->, belongsTo=parrnote
776	++, otherFeat=[]-->, belongsTo=parrnote
777	--, otherFeat=[]-->, belongsTo=parrnote
778	++ -, otherFeat=[]-->, belongsTo=parrnote
779	-, otherFeat=[]-->, belongsTo=parrnote
780	+, otherFeat=[]-->, belongsTo=parrnote
781	-, otherFeat=[]-->, belongsTo=parrnote
782	+, otherFeat=[]-->, belongsTo=parrnote
783	-, otherFeat=[]-->, belongsTo=parrnote
784	+, otherFeat=[]-->, belongsTo=parrnote
785	-, otherFeat=[]-->, belongsTo=parrnote
786	+ VSV-Borealin, otherFeat=[]-->, belongsTo=parrnote
787	Anti-GFP, otherFeat=[]-->, belongsTo=parrnote
788	Anti-VSV, otherFeat=[]-->, belongsTo=parrnote
789	GFP-INCENP47-920, otherFeat=[]-->, belongsTo=parrnote
790	123 4 5 678, otherFeat=[]-->, belongsTo=parrnote
791	GFP, otherFeat=[]-->, belongsTo=parrnote
792	CREST, otherFeat=[]-->, belongsTo=parrnote
793	DNA, otherFeat=[]-->, belongsTo=parrnote
794	Merge w/DNA, otherFeat=[]-->, belongsTo=parrnote
795	GFP, otherFeat=[]-->, belongsTo=parrnote
796	Borealin, otherFeat=[]-->, belongsTo=parrnote
797	CREST, otherFeat=[]-->, belongsTo=parrnote
798	IgG, otherFeat=[]-->, belongsTo=parrnote
799	AB, otherFeat=[]-->, belongsTo=parrnote
800	C, otherFeat=[]-->, belongsTo=parrnote
801	G, otherFeat=[]-->, belongsTo=parrnote
802	H, otherFeat=[]-->, belongsTo=parrnote
803	CPC targeting mediated by Survivin, otherFeat=[]-->, belongsTo=nota_cab_pie
804	G. Vader et al, otherFeat=[]-->, belongsTo=nota_cab_pie
805	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
806	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
807	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
808	90, otherFeat=[]-->, belongsTo=nota_cab_pie
809	cDNA library using a 50 primer containing a VSV tag and cloned, otherFeat=[]-->, belongsTo=parr
810	into pcDNA3.1 (INCENP) or into pCR3 containing an N-terminal, otherFeat=[]-->, belongsTo=parr
811	VSV tag (Borealin). RNAi-resistant INCENP and Borealin con-, otherFeat=[]-->, belongsTo=parr
812	structs were generated using the QuickChanges Mutagenesis kit, otherFeat=[]-->, belongsTo=parr
813	(Stratagene, La Jolla, CA, USA). For INCENP47?920, INCENP was, otherFeat=[]-->, belongsTo=parr
814	digested with EcoRI/XhoI. This fragment was cloned into pCR3, otherFeat=[]-->, belongsTo=parr
815	containing an N-terminal VSV tag. For INCENP1?47, INCENP was, otherFeat=[]-->, belongsTo=parr
816	digested with HindIII/EcoRI, and the product was cloned into, otherFeat=[]-->, belongsTo=parr
817	pcDNA3.1. For Survivin?INCENP47?920, Survivin was ligated to, otherFeat=[]-->, belongsTo=parr
818	the N terminus of INCENP47?920 after EcoRI digestion. For the, otherFeat=[]-->, belongsTo=parr
819	C-terminal GFP constructs, inserts were cloned into various, otherFeat=[]-->, belongsTo=parr
820	peGFP-N-vectors (Clontech, Mountain View, CA, USA). Note, otherFeat=[]-->, belongsTo=parr
821	that the INCENP?GFP constructs also contain an N-terminal, otherFeat=[]-->, belongsTo=parr
822	VSV tag. Histone H2B?GFP, Histone H2B?diHcRed (gifts from, otherFeat=[]-->, belongsTo=parr
823	Dr J. Ellenberg), Survivin siRNA, VSV?Survivin, pBABE-puro and, otherFeat=[]-->, belongsTo=parr
824	Spectrin?GFP have been described previously (Lens et al, 2003)., otherFeat=[]-->, belongsTo=parr
825	Cell culture and transfection. Human osteosarcoma U2OS and, otherFeat=[]-->, belongsTo=parr
826	human embryonic kidney 293 cells were cultured and synchro-, otherFeat=[]-->, belongsTo=parr
827	nized, as described (Lens et al, 2003). Cells were transfected using, otherFeat=[]-->, belongsTo=parr
828	the standard calcium phosphate protocol., otherFeat=[]-->, belongsTo=parr
829	Fig 3 | Survivin can target a functional chromosome passenger complex in the absence of Borealin, but interaction with inner centromere protein, otherFeat=[]-->, belongsTo=fig_caption
830	(INCENP) depends on Borealin. (A) U2OS cells were transfected with the indicated plasmids, puromycin was added to enrich for transfected cells and, otherFeat=[]-->, belongsTo=fig_caption
831	lysates were analysed by western blotting. (B?E) Cells were transfected with the indicated plasmids. (C) Cells grown on coverslips were transfected, otherFeat=[]-->, belongsTo=fig_caption
832	with mock or Borealin short interfering RNA (siRNA) plasmid and H2B?GFP (green fluorescent protein). Immunostainings were carried out with, otherFeat=[]-->, belongsTo=fig_caption
833	anti-Borealin, and the percentage of H2B?GFP-positive prometaphase cells in which Borealin localized on centromeres was scored. The number of cells, otherFeat=[]-->, belongsTo=fig_caption
834	counted per condition is indicated. (B,D) Cells were grown asynchronously for 60 h and collected for fluorescence-activated cell sorting (FACS), otherFeat=[]-->, belongsTo=fig_caption
835	analysis. (E) Taxol was added after thymidine release, and cells were collected for FACS analysis 18 h later. Mitotic percentages were determined, otherFeat=[]-->, belongsTo=fig_caption
836	by MPM-2 staining. (F) Cells grown on coverslips were transfected with Borealin siRNA plasmid and VSV?Survivin?INCENP47?920-sil?GFP., otherFeat=[]-->, belongsTo=fig_caption
837	Immunostainings were carried out with the indicated antibodies, and DNA was stained with 4,6-diamidino-2-phenylindole. Different colours used for, otherFeat=[]-->, belongsTo=fig_caption
838	the two merge pictures are indicated. (G) Human embryonic kidney 293 cells were transfected with 5 mg of the indicated constructs. After 48 h, cells, otherFeat=[]-->, belongsTo=fig_caption
839	were collected, immunoprecipitations were carried out with anti-GFP and samples were analysed by western blotting. (H) U2OS cells were transfected, otherFeat=[]-->, belongsTo=fig_caption
840	with the indicated plasmids. At 18 h after transfection, puromycin was added and 36 h later, cells were collected . Immunoprecipitations were carried, otherFeat=[]-->, belongsTo=fig_caption
841	out with anti-GFP and western blots were probed with the indicated antibodies. Scale bars, 5 mm., otherFeat=[]-->, belongsTo=fig_caption
842	b, otherFeat=[]-->, belongsTo=nota_cab_pie
843	Indirect centromere-, otherFeat=[]-->, belongsTo=??
844	targeting domain, otherFeat=[]-->, belongsTo=??
845	IN-box, otherFeat=[]-->, belongsTo=??
846	147, otherFeat=[]-->, belongsTo=??
847	920, otherFeat=[]-->, belongsTo=??
848	INCENP, otherFeat=[]-->, belongsTo=??
849	INCENP-sil, otherFeat=[]-->, belongsTo=??
850	47, otherFeat=[]-->, belongsTo=??
851	920, otherFeat=[]-->, belongsTo=??
852	INCENP, otherFeat=[]-->, belongsTo=??
853	INCENP47-920-sil, otherFeat=[]-->, belongsTo=??
854	47, otherFeat=[]-->, belongsTo=??
855	920, otherFeat=[]-->, belongsTo=??
856	Survivin, otherFeat=[]-->, belongsTo=??
857	INCENP, otherFeat=[]-->, belongsTo=??
858	Survivin-INCENP47-920-sil, otherFeat=[]-->, belongsTo=??
859	147, otherFeat=[]-->, belongsTo=??
860	INCENP1-47, otherFeat=[]-->, belongsTo=??
861	* *, otherFeat=[]-->, belongsTo=??
862	* *, otherFeat=[]-->, belongsTo=??
863	* *, otherFeat=[]-->, belongsTo=??
864	Indirect centromere-, otherFeat=[]-->, belongsTo=??
865	targeting domain, otherFeat=[]-->, belongsTo=??
866	IN-box, otherFeat=[]-->, belongsTo=??
867	147, otherFeat=[]-->, belongsTo=??
868	920, otherFeat=[]-->, belongsTo=??
869	INCENP, otherFeat=[]-->, belongsTo=??
870	Survivin, otherFeat=[]-->, belongsTo=??
871	Anaphase, otherFeat=[]-->, belongsTo=??
872	(central spindle), otherFeat=[]-->, belongsTo=??
873	Prometaphase, otherFeat=[]-->, belongsTo=??
874	(centromeres), otherFeat=[]-->, belongsTo=??
875	Localization, otherFeat=[]-->, belongsTo=??
876	Interaction,, otherFeat=[]-->, belongsTo=??
877	stabilization, otherFeat=[]-->, belongsTo=??
878	RNAi, otherFeat=[]-->, belongsTo=??
879	Aurora-B, otherFeat=[]-->, belongsTo=??
880	Interaction,, otherFeat=[]-->, belongsTo=??
881	stabilization, otherFeat=[]-->, belongsTo=??
882	Borealin, otherFeat=[]-->, belongsTo=??
883	Cytokinesis, otherFeat=[]-->, belongsTo=??
884	(midbody), otherFeat=[]-->, belongsTo=??
885	A, otherFeat=[]-->, belongsTo=??
886	B, otherFeat=[]-->, belongsTo=??
887	Fig 4 | Overview of inner centromere protein constructs and model. (A) Schematic representation of inner centromere protein (INCENP) constructs., otherFeat=[]-->, belongsTo=fig_caption
888	Survivin and Aurora-B binding domains are depicted. Introduced RNA interference (RNAi) silent mutations are shown. (B) Model showing the, otherFeat=[]-->, belongsTo=fig_caption
889	proposed upstream targeting role of Survivin in the chromosome passenger complex., otherFeat=[]-->, belongsTo=fig_caption
890	CPC targeting mediated by Survivin, otherFeat=[]-->, belongsTo=nota_cab_pie
891	G. Vader et al, otherFeat=[]-->, belongsTo=nota_cab_pie
892	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION, otherFeat=[]-->, belongsTo=nota_cab_pie
893	EMBO reports VOL 7 | NO 1 | 2006, otherFeat=[]-->, belongsTo=nota_cab_pie
894	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
895	91, otherFeat=[]-->, belongsTo=nota_cab_pie
896	Flow cytometry and time-lapse microscopy. The percentage of, otherFeat=[]-->, belongsTo=parr
897	mitotic cells (MPM-2 positivity) and cell cycle distribution of, otherFeat=[]-->, belongsTo=parr
898	spectrin?GFP-transfected cells were determined by flow cytometry,, otherFeat=[]-->, belongsTo=parr
899	as described (Smits et al, 2000). H2B?GFP-expressing cells were, otherFeat=[]-->, belongsTo=parr
900	followed by time-lapse microscopy, as described (Lens et al, 2003)., otherFeat=[]-->, belongsTo=parr
901	Immunoblotting, immunoprecipitation and immunofluorescence., otherFeat=[]-->, belongsTo=parr
902	Immunoblotting was carried out as described (Smits et al, 2000)., otherFeat=[]-->, belongsTo=parr
903	For immunoprecipitation, cells were lysed in E1A lysis buffer with, otherFeat=[]-->, belongsTo=parr
904	protease inhibitors (Completes, Roche, Indianapolis, IN, USA) for, otherFeat=[]-->, belongsTo=parr
905	30 min at 4 1C (Smits et al, 2000). GFP-tagged proteins were, otherFeat=[]-->, belongsTo=parr
906	immunoprecipitated with 3 mg anti-GFP precoupled to protein G?, otherFeat=[]-->, belongsTo=parr
907	Sepharose (Amersham Biosciences, Pittsburg, CA, USA). Immuno-, otherFeat=[]-->, belongsTo=parr
908	fluorescence was carried out as described (Lens et al, 2003). DNA, otherFeat=[]-->, belongsTo=parr
909	was stained with 4,6-diamidino-2-phenylindole., otherFeat=[]-->, belongsTo=parr
910	Supplementary information is available at EMBO reports online, otherFeat=[]-->, belongsTo=parr
911	(http://www.emboreports.org)., otherFeat=[]-->, belongsTo=parr
912	ACKNOWLEDGEMENTS, otherFeat=[]-->, belongsTo=parrnote
913	We thank B.v.d. Weerdt, M.v. Vugt and G. Kops for comments on, otherFeat=[]-->, belongsTo=parrnote
914	the manuscript, T.v. Harn for technical support and S. Taylor and, otherFeat=[]-->, belongsTo=parrnote
915	H. Funabiki for the gift of reagents. This work was supported by, otherFeat=[]-->, belongsTo=parrnote
916	the Dutch Cancer Society (NKI 2002-2764)., otherFeat=[]-->, belongsTo=parrnote
917	REFERENCES, otherFeat=[]-->, belongsTo=parrnote
918	Adams RR, Wheatley SP, Gouldsworthy AM, Kandels-Lewis SE, Carmena M,, otherFeat=[]-->, belongsTo=parrnote
919	Smythe C, Gerloff DL, Earnshaw WC (2000) INCENP binds the Aurora-, otherFeat=[]-->, belongsTo=parrnote
920	related kinase AIRK2 and is required to target it to chromosomes, the, otherFeat=[]-->, belongsTo=parrnote
921	central spindle and cleavage furrow. Curr Biol 10: 1075?1078, otherFeat=[]-->, belongsTo=parrnote
922	Ainsztein AM, Kandels-Lewis SE, Mackay AM, Earnshaw WC (1998) INCENP, otherFeat=[]-->, belongsTo=parrnote
923	centromere and spindle targeting: identification of essential conserved, otherFeat=[]-->, belongsTo=parrnote
924	motifs and involvement of heterochromatin protein HP1. J Cell Biol 143:, otherFeat=[]-->, belongsTo=parrnote
925	1763?1774, otherFeat=[]-->, belongsTo=parrnote
926	Biggins S, Murray AW (2001) The budding yeast protein kinase Ipl1/Aurora, otherFeat=[]-->, belongsTo=parrnote
927	allows the absence of tension to activate the spindle checkpoint. Genes, otherFeat=[]-->, belongsTo=parrnote
928	Dev 15: 3118?3129, otherFeat=[]-->, belongsTo=parrnote
929	Ditchfield C, Johnson VL, Tighe A, Ellston R, Haworth C, Johnson T, Mortlock, otherFeat=[]-->, belongsTo=parrnote
930	A, Keen N, Taylor SS (2003) Aurora B couples chromosome alignment, otherFeat=[]-->, belongsTo=parrnote
931	with anaphase by targeting BubR1, Mad2, and Cenp-E to kinetochores., otherFeat=[]-->, belongsTo=parrnote
932	J Cell Biol 161: 267?280, otherFeat=[]-->, belongsTo=parrnote
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999	scientific report, otherFeat=[]-->, belongsTo=nota_cab_pie
1000	92, otherFeat=[]-->, belongsTo=nota_cab_pie
==============================
0	Survivin mediates targeting of the chromosomal passenger complex to the centromere and midbody-->id=0, page=0, size=33, fam=Times, col=#000000, type=title, textLines=2--->[]--->font_title	Survivin mediates ta>>>ntromere and midbody
1	Gerben Vaderw,Jos J.W. Kauw,Rene´ H. Medemaw & Susanne M.A. Lens+w-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	Gerben Vaderw,Jos J.>>> Susanne M.A. Lens+w
2	Division of Molecular Biology, Netherlands Cancer Institute, Amsterdam, The Netherlands-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_note	Division of Molecula>>>dam, The Netherlands
3	The chromosomal passenger complex (CPC) coordinates chromosomal and cytoskeletal events of mitosis. The enzymatic core of this complex (Aurora-B) is guided through the mitotic cell by its companion chromosomal passenger proteins, inner centromere protein (INCENP), Survivin and Borealin/Dasra-B, thereby allowing it to act at the right place at the right time. Here, we addressed the individual contributions of INCENP, Survivin and Borealin to the proper functioning of this complex. We show that INCENP has an important role in stabilizing the complex, and that Borealin acts to promote binding of Survivin to INCENP. Importantly, when Survivin is directly fused to INCENP, this hybrid can restore CPC function at the centromeres and midbody, even in the absence of Borealin and the centromere-targeting domain of INCENP. Thus, Survivin is an important mediator of centromere and midbody docking of Aurora-B during mitosis.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	The chromosomal pass>>>ra-B during mitosis.
4	Keywords: cytokinesis; spindle checkpoint-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Keywords: cytokinesi>>>; spindle checkpoint
5	EMBO reports (2006) 7, 85­92. doi:10.1038/sj.embor.7400562-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	EMBO reports (2006) >>>038/sj.embor.7400562
6	INTRODUCTION-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	INTRODUCTION>>>INTRODUCTION
7	Proper mitosis requires accurate nuclear division, followed by cytoplasmic partitioning into two daughter cells during cytokinesis. Nuclear division is monitored by the spindle assembly checkpoint (SAC), which is an intrinsic cell-cycle checkpoint that prevents the onset of anaphase until all sister chromatids have attained proper attachment to the mitotic spindle (Nicklas, 1997). To satisfy this checkpoint, kinetochores of paired sister chromatids have to attach in a bipolar manner, creating tension (Tanaka, 2002). Aurora-B kinase is essential to resolve naturally occurring monopolar or syntelic microtubule­kinetochore attachments (Tanaka et al, 2002; Lampson et al, 2004), and to communicate a lack of bipolarity (that is, tension) to the core SAC (Biggins & Murray, 2001; Lens & Medema, 2003). Additionally, in several organisms, interference with Aurora-B leads to a failure of cytokinesis (Kaitna et al, 2000; Severson et al, 2000; Ditchfield et al, 2003; Hauf et al, 2003).-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Proper mitosis requi>>>; Hauf et al, 2003).
8	Aurora-B acts together with Survivin, inner centromere protein (INCENP) and Borealin/Dasra-B (hereafter referred to as Borealin). These proteins physically interact and show a typical chromosomal passenger localization pattern during mitosis--at the inner centromere in (pro)metaphase, on the central spindle during anaphase and at the midbody during cytokinesis (Vagnarelli & Earnshaw, 2004). Interference with Survivin, INCENP or Borealin function disrupts Aurora-B localization and leads to similar mitotic defects, as observed after interference with Aurora-B function (Vagnarelli & Earnshaw, 2004, and references therein). Aurora-B is activated at the G2­M transition and remains active until mitotic exit (Yasui et al, 2004). Thus, it is likely that the specific functions of Aurora-B are mainly determined by its dynamic localization pattern. Until now, INCENP has been proposed to be the targeting module of the complex (Ainsztein et al, 1998), but specific functions for Survivin and Borealin in the chromosome passenger complex (CPC) have yet to be determined.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Aurora-B acts togeth>>>et to be determined.
9	RESULTS AND DISCUSSION-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->['U']--->font_title	RESULTS AND DISCUSSI>>>SULTS AND DISCUSSION
10	An RNAi-complementation to study CPC function-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	An RNAi-complementat>>>o study CPC function
11	We developed an INCENP RNA interference (RNAi) complementation system to study the role of INCENP in the CPC. Endogenous INCENP was knocked down by vector-driven short interfering RNA (siRNA; Fig 1A; supplementary Fig 1A­C online) and cells were reconstituted with an INCENP complementary DNA harbouring two silent mutations in the siRNA-targeting region, rendering it insensitive to the siRNA (Lens et al, 2003). INCENPdepleted cells showed the typical `chromosome passenger deficiency' phenotype: defects in chromosome alignment and segregation, and a failure to undergo cytokinesis (supplementary Fig 1A­C online; Honda et al, 2003). Additionally, Aurora-B and BubR1 were mislocalized, and the INCENP-depleted cells failed to maintain an active SAC in response to Taxol (supplementary Fig 1B,D,E online). All these mitotic defects could be reverted by expression of a full-length INCENP containing two silent mutations (VSV­INCENP-sil; Figs 1C­E,2F; supplementary Fig 2B,C online; for INCENP constructs used, see Fig 4A), confirming the specificity of the siRNA and the functionality of the complementation approach.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	We developed an INCE>>>ementation approach.
12	Received 27 May 2005; revised 20 September 2005; accepted 21 September 2005; published online 21 October 2005-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Received 27 May 2005>>>line 21 October 2005
13	+Corresponding author. Tel: þ 3130 253 9728; Fax: þ 3130 253 8479; E-mail: s.m.a.lens@med.uu.nl-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+Corresponding autho>>>s.m.a.lens@med.uu.nl
14	Division of Molecular Biology, Netherlands Cancer Institute, Plesmanlaan 21, 1066 CX Amsterdam, The Netherlands wPresent address: Department of Medical Oncology, University Medical Center, Str. 2.103 Universiteitsweg 100, 3584 CG Utrecht, The Netherlands-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Division of Molecula>>>cht, The Netherlands
15	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
16	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
17	scientificreport-->id=8, page=0, size=29, fam=Times, col=#dedede, type=title, textLines=1--->[]--->font_note	scientificreport>>>scientificreport
18	scientific report-->id=9, page=0, size=72, fam=Times, col=#008a33, type=title, textLines=1--->[]--->font_note	scientific report>>>scientific report
19	85-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	85>>>85
20	CPC formation is crucial for protein stability-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	CPC formation is cru>>>or protein stability
21	In line with earlier observations (Honda et al, 2003), total protein levels of Aurora-B and Survivin were significantly reduced after INCENP depletion (Fig 1A). This reduction was interpreted as an indirect consequence of a failed cell division followed by a cellcycle arrest in a tetraploid G1 state (Honda et al, 2003). However, we found that INCENP-depleted cells already failed to accumulate these proteins during passage through G2 (Fig 1B). Moreover, mitotic cells lacking INCENP also showed reduced Aurora-B and Survivin levels (Fig 1B,H, lane 2), which indicates that the effect of INCENP depletion on protein stability is more direct.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	In line with earlier>>>lity is more direct.
22	By introducing different INCENP truncation mutants into INCENP-depleted cells, we could show that a mutant lacking the amino-terminal `centromere-targeting' domain (VSV­ INCENP47­920-sil; Fig 4A) restored Aurora-B levels, but failed to stabilize Survivin (Fig 1G, lane 4). In line with previous observations (Ainsztein et al, 1998), this mutant failed to concentrate at centromeres during prometaphase and to transfer to the midbody following the onset of anaphase (supplementary Fig 2B,C online). Consequently, this mutant was unable to restore a SAC-mediated arrest in response to Taxol and to overcome the cytokinesis defect in INCENP-depleted cells (Fig 1D,E). Because of the presence of the carboxy-terminal Aurora-B interaction domain (IN-box; Adams et al, 2000), INCENP47­920-sil was able to bind to Aurora-B. In contrast, this truncation mutant failed to interact with Survivin (Fig 1F, lane 3). Conversely, a construct expressing only-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	By introducing diffe>>>ruct expressing only
23	the first N-terminal 47 amino acids of INCENP (INCENP1­47;-->id=785, page=1, size=6, fam=Times, col=#000000, type=note, textLines=1--->[]--->font_note	the first N-terminal>>> INCENP (INCENP1­47;
24	Fig 4A) and thus harbouring the centromere-localization domain was able to bind to Survivin (Fig 1F, lane 4), but failed to stabilize it (Fig 1G, lane 5). Thus, this domain is both necessary and sufficient for interaction with Survivin, but the presence of Aurora-B in the complex may be needed for stabilization of Survivin. Interestingly, Survivin is an in vivo substrate of Aurora-B (Wheatley et al, 2004), but whether this phosphorylation affects protein stability is unknown. In contrast, the presence of Survivin in the complex was not essential to stabilize Aurora-B, because we found that a complex of Aurora-B and a truncated INCENP, although lacking Survivin, could exist (Fig 1F,G). However, whereas Aurora-B protein levels could be restored by expression of INCENP47­920-sil, phosphorylation of serine-10 Histone-H3 in mitotic INCENP-depleted cells remained impaired (Fig 1H, lane 4). Thus, stabilization of Aurora-B by INCENP47­920-sil is not sufficient to mediate Histone-H3 phosphorylation. This suggests that mitotic phosphorylation of Histone-H3 on serine-10 by Aurora-B requires either the direct presence of Survivin in the complex or centromeric localization of the kinase.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Fig 4A) and thus har>>>ation of the kinase.
25	Survivin mediates mitotic localization of the CPC-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	Survivin mediates mi>>>alization of the CPC
26	Although INCENP contains a highly conserved `centromeretargeting' domain in its N-terminus (Ainsztein et al, 1998), the mechanism by which this domain mediates centromeric localization remains elusive. As INCENP interacts with Survivin by means of this N-terminal domain (Fig 1F), it is possible that Survivin mediates mitotic localization of the CPC. To test this, a chimeric protein that is refractory to both INCENP and Survivin RNAi (Fig 2A; data not shown) was generated, in which the centromeretargeting domain in INCENP was replaced by Survivin (SurvivinINCENP47­920-sil; Fig 4A). This fusion localized normally during mitosis, in both an INCENP-depleted and a combined INCENP/ Survivin-depleted background (Fig 2D,G, arrows). This chimaera was also able to localize Aurora-B during mitosis (Fig 2E,F), restore serine-10 Histone-H3 phosphorylation (supplementary Fig 2D online) and restore CPC function, as expression of the chimaera in INCENP-, Survivin- or combined INCENP/Survivindepleted cells rescued SAC function and the proper execution of cytokinesis (Fig 2B,C).-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Although INCENP cont>>>okinesis (Fig 2B,C).
27	To obtain further evidence that Survivin can serve as a determining factor in localizing the CPC, we analysed the location of INCENP1­47 in the absence or presence of endogenous Survivin. INCENP1­47­GFP (green fluorescent protein) localized to centromeres during mitosis in control cells (Fig 2H, upper panel, see arrows), but failed to concentrate at centromeres in the absence of endogenous Survivin and instead localized diffusely in the cytoplasm during mitosis (Fig 2G, lower panel, see arrows), indicating that the centromere-targeting domain in INCENP requires at least the presence of Survivin to localize to centromeres. Taken together, these results show that Survivin is an important mediator of mitotic localization of the CPC.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	To obtain further ev>>>lization of the CPC.
28	Survivin can target the CPC in the absence of Borealin-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	Survivin can target >>> absence of Borealin
29	Recently, it was suggested that Borealin has a role in determining centromeric localization of the CPC (Gassmann et al, 2004). As our results suggest that targeting of the Survivin/INCENP/Aurora-B complex is mediated by Survivin, we explored whether this depended on the presence of Borealin. We depleted Borealin using siRNA (Fig 3A­C) and introduced the Survivin­INCENP chimaera into these cells. Interestingly, expression of this chimaera in Borealin-depleted cells significantly restored CPC-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Recently, it was sug>>>icantly restored CPC
30	Fig 1 | Protein­protein interactions within and stability of the chromosomal passenger complex. U2OS cells were transfected with 10 mg mock or inner centromere protein (INCENP) short interfering RNA (siRNA) vector combined with pBABE-puro. (A) Puromycin was added to enrich for transfected cells, and lysates for western blotting were prepared. (B) Transfected cells were released from a thymidine block and, at indicated time points, lysates were prepared for western blotting. N indicates nocodazole treatment for 24 h to enrich for mitotic cells. (C)5 mg of the indicated VSV-tagged plasmids was transfected in combination with the siRNA plasmids and pBABE-puro. After puromycin selection, lysates were analysed by western blotting. (D) Similar to (C), after release from the thymidine block, cells were treated with Taxol for 18 h and collected for fluorescence-activated cell sorting (FACS) analysis. Mitotic percentages were determined by staining the cells with the MPM-2 antibody that specifically recognizes mitotic phosphoepitopes. (E) Similar to (C), 60 h after transfection, cells were collected for FACS analysis. (F) Human embryonic kidney (HEK) 293 cells were transfected with 5 mg of the indicated green fluorescent protein (GFP)-tagged constructs. After 48 h, cells were collected , immunoprecipitations were carried out with anti-GFP and samples were analysed by western blotting. The presence of GFP protein (arrowhead) is due to the presence of an additional transcription site in the plasmids used. (G) Similar to (C). Note that INCENP1­47 contains both an amino-terminal VSV tag and a carboxyterminal GFP tag. The asterisk denotes an aspecific band recognized by the anti-VSV monoclonal antibody. (H) Similar to (C). At 24 h after puromycin selection, nocodazole was added to the cells for 18 h. Mitotic cells were obtained by shake-off and lysates were analysed by western blotting.-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_capfig	Fig 1 | Protein­prot>>>by western blotting.
31	c-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_capfig	c>>>c
32	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
33	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
34	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
35	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
36	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
37	86-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	86>>>86
38	function, as judged by restoration of mitotic arrest in response to Taxol and a reduction in the percentage of polyploid cells that were observed after depletion of Borealin (Fig 3D,E). Expression of neither Survivin nor INCENP individually was sufficient to restore CPC function in Borealin-depleted cells (Fig 3E; data not shown). In line with these functional data, the chimaera was found to localize to centromeres and the midbody in Borealin-depleted cells (Fig 3F) and to restore Aurora-B localization (supplementary-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	function, as judged >>>ation (supplementary
39	0 9 12 15 18 24 N-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0 9 12 15 18 24 N>>>0 9 12 15 18 24 N
40	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
41	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
42	0 9 12 15 18 24 N-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0 9 12 15 18 24 N>>>0 9 12 15 18 24 N
43	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
44	INCENP1-47-GFP-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP1-47-GFP>>>INCENP1-47-GFP
45	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
46	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
47	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
48	INCENP47-920-sil-GFP-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP47-920-sil-GFP>>>INCENP47-920-sil-GFP
49	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
50	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
51	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
52	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
53	INCENP-sil-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP-sil-GFP>>>INCENP-sil-GFP
54	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
55	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
56	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
57	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
58	Empty vector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Empty vector>>>Empty vector
59	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
60	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
61	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
62	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
63	Total lysate-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Total lysate>>>Total lysate
64	IP: Anti-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	IP: Anti-GFP>>>IP: Anti-GFP
65	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
66	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
67	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
68	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
69	+ --->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ ->>>+ -
70	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
71	- +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	- +>>>- +
72	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
73	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
74	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
75	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
76	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
77	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
78	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
79	Hours-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Hours>>>Hours
80	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
81	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
82	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
83	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
84	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
85	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
86	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
87	---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-->>>--
88	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
89	---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-->>>--
90	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
91	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
92	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
93	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
94	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
95	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
96	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
97	Mock siRNA INCENP siRNA Empty vector VSV -INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA INCENP si>>>ctor VSV -INCENP-sil
98	VSV-INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	VSV-INCENP47-920-sil>>>VSV-INCENP47-920-sil
99	Mitotic shake-off (nocodazole)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mitotic shake-off (n>>>ake-off (nocodazole)
100	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
101	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
102	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
103	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
104	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
105	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
106	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
107	---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-->>>--
108	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
109	-+ --->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-+ ->>>-+ -
110	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
111	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
112	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
113	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
114	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
115	Mock siRNA INCENP siRNA Empty vector VSV-INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA INCENP si>>>ector VSV-INCENP-sil
116	VSV-INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	VSV-INCENP47-920-sil>>>VSV-INCENP47-920-sil
117	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
118	+ --->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ ->>>+ -
119	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
120	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
121	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
122	---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-->>>--
123	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
124	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
125	VSV-INCENP1-47-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-INCENP1-47-GFP>>>VSV-INCENP1-47-GFP
126	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
127	INCENPsiRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENPsiRNA>>>INCENPsiRNA
128	Mock-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock>>>Mock
129	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
130	GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP>>>GFP
131	Mock siRNA INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA INCENP si>>>k siRNA INCENP siRNA
132	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
133	Anti-Cdk6-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Cdk6>>>Anti-Cdk6
134	VSV-INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-INCENP>>>VSV-INCENP
135	VSV-INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-INCENP-sil>>>VSV-INCENP-sil
136	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
137	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
138	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
139	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
140	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
141	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
142	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
143	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
144	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
145	+ VSV-INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ VSV-INCENP-sil>>>+ VSV-INCENP-sil
146	4n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4n>>>4n
147	2n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2n>>>2n
148	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
149	+ VSV-INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	+ VSV-INCENP47-920-s>>>VSV-INCENP47-920-sil
150	8n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8n>>>8n
151	VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV>>>VSV
152	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
153	INCENP-sil VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP-sil VSV>>>INCENP-sil VSV
154	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
155	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP>>>INCENP
156	47-920-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	47-920>>>47-920
157	-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-sil>>>-sil
158	Emptyvector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Emptyvector>>>Emptyvector
159	Mitotic-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mitotic>>>Mitotic
160	cells-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	cells>>>cells
161	(%)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	(%)>>>(%)
162	40-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	40>>>40
163	20-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	20>>>20
164	0-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0>>>0
165	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
166	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
167	+Taxol-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+Taxol>>>+Taxol
168	10.1%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	10.1%>>>10.1%
169	25.8%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	25.8%>>>25.8%
170	12.0%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	12.0%>>>12.0%
171	19.3%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	19.3%>>>19.3%
172	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
173	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
174	12 3 4-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	12 3 4>>>12 3 4
175	5 6-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	5 6>>>5 6
176	7 8-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	7 8>>>7 8
177	12-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	12>>>12
178	3-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	3>>>3
179	4-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4>>>4
180	12345-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	12345>>>12345
181	Anti-INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-INCENP>>>Anti-INCENP
182	Anti-Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Aurora-B>>>Anti-Aurora-B
183	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
184	Anti-Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Aurora-B>>>Anti-Aurora-B
185	Anti-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-GFP>>>Anti-GFP
186	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
187	Anti-Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Aurora-B>>>Anti-Aurora-B
188	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
189	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
190	Anti-Cdk4-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Cdk4>>>Anti-Cdk4
191	Anti-Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Aurora-B>>>Anti-Aurora-B
192	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
193	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
194	Anti-phospho-Histone H3-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-phospho-Histone>>>i-phospho-Histone H3
195	Anti-Cdk4-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Cdk4>>>Anti-Cdk4
196	Anti-Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Aurora-B>>>Anti-Aurora-B
197	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
198	Anti-Actin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Actin>>>Anti-Actin
199	Anti-Cdk4-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Cdk4>>>Anti-Cdk4
200	GH-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	GH>>>GH
201	EF-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	EF>>>EF
202	CD-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	CD>>>CD
203	A-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	A>>>A
204	B-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	B>>>B
205	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
206	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
207	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
208	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
209	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
210	87-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	87>>>87
211	Fig 3 online). However, it should be noted that the association of the Survivin­INCENP chimaera to centromeres and the central spindle was affected in Borealin-depleted cells. Instead of concentrating at centromeres and central spindle, a significant portion of the chimaera localized to chromatin throughout mitosis (Fig 3F, lower panel; data not shown). Taken together, these data show that the Survivin­INCENP chimaera is sufficient to restore a functional CPC in Borealin-depleted cells, and that Borealin might facilitate dissociation of the CPC from chromatin or enhance its concentration at centromeres and the central spindle.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Fig 3 online). Howev>>>the central spindle.
212	Borealin facilitates Survivin­INCENP interaction-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	Borealin facilitates>>>n­INCENP interaction
213	As Borealin is essential for localization of INCENP and Survivin (Gassmann et al, 2004), but not for a chimaera in which these two proteins are covalently linked, we reasoned that Borealin might be important for the Survivin­INCENP interaction. To further investigate the potential role of Borealin in regulating the Survivin­INCENP interaction, immunoprecipitations were carried out with GFP­INCENP from Borealin-depleted cells expressing either exogenous Survivin or Borealin. In control cells, both endogenous and exogenous Survivin could interact with INCENP (Fig 3H, lanes 1­3). In contrast, in the absence of endogenous Borealin, neither endogenous nor exogenous Survivin was able to interact with INCENP (Fig 3H, lanes 4,5). As depletion of Borealin also reduced Survivin levels, this probably explains why no endogenous Survivin was found to interact with INCENP (Fig 3H, lanes 4,10; Gassmann et al, 2004; Sampath et al, 2004). Indeed, after expression of exogenous, RNAi-resistant Borealin, endogenous Survivin levels and Survivin interaction with INCENP were restored (Fig 3E, lanes 6,12). Importantly, although exogenous Survivin levels were not affected by Borealin depletion, Survivin failed to interact with INCENP, indicating that, in vivo, an efficient interaction of Survivin and INCENP is dependent on the presence of Borealin (Fig 3E, lanes 5,11). The observation that Borealin can interact with INCENP that lacks the Survivin interaction domain (INCENP47­920; Fig 3G, lanes 6,8; supplementary Fig 3 online), in combination with previous in vitro data that have shown the direct interaction of Survivin and INCENP (Wheatley et al, 2001; Romano et al, 2003), rules out a direct bridging function for Borealin between Survivin and the centromere-targeting domain of INCENP. Hence, similar to CSC-1, its distant homologue in Caenorhabditis elegans (Romano et al, 2003), human Borealin has a role in facilitating Survivin­INCENP interaction. By generating the Survivin­INCENP chimaera, we have circumvented the necessity for Borealin in stabilizing Survivin­INCENP interaction, which probably explains why a functional CPC can be directed to the right mitotic structures in Borealin-depleted cells.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	As Borealin is essen>>>alin-depleted cells.
214	Conclusion-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	Conclusion>>>Conclusion
215	Given the essential functions of Aurora-B during mitosis, explaining the mechanisms behind its dynamic localization is of great importance. The observation that depletion of one CPC component by siRNA results in the destabilization of the entire complex instigates the need for more complex experiments to unravel the functions of single CPC components. Using RNAi-complementation, we have provided evidence that an important function of Survivin is to serve as an interface between the centromere/central spindle and the CPC, whereas Borealin facilitates the interaction of Survivin and INCENP (Fig 4B). Importantly, the targeting function of Survivin was revealed only when Survivin was fused to INCENP lacking its centromere-targeting domain. This suggests that although Survivin can mediate localization of the CPC to its proper mitotic structures, the other CPC components are probably needed to obtain a stable association (possibly by affecting the on/off rate of the complex). Identifying the Survivin interaction partners at the centromere and midbody will be an important next step in understanding the regulation of the CPC.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Given the essential >>>gulation of the CPC.
216	METHODS-->id=4, page=0, size=12, fam=Times, col=#000000, type=title, textLines=10--->[]--->font_title	METHODS>>>METHODS
217	Antibodies and reagents. The following antibodies were used: rabbit anti-INCENP and rabbit anti-Aurora-B (Abcam, Cambridge, UK), rabbit anti-Survivin (R&D, Minneapolis, MN, USA), rabbit anti-Borealin (gift from Dr H. Funabiki), mouse anti-Aurora-B (Transduction, Lexington, KE, USA), mouse anti-VSV and mouse anti-a-Tubulin (Sigma, St Louis, MO, USA), rabbit anti-Cdk6 and rabbit anti-Cdk4 (Santa Cruz, Santa Cruz, CA, USA), sheep anti-BubR1 (gift from Dr S. Taylor), rabbit anti-GFP (gift from Dr J. Neefjes), rabbit anti-phospho-Ser10-histone-H3 and mouse anti-MPM-2 (Upstate, Charlotsville, VA, USA), CREST antiserum (Cortex Biochem, San Leandro, CA, USA), peroxidase-conjugated goat anti-rabbit and peroxidase-conjugated goat anti-mouse (Dako, Glostrup, Denmark) and donkey anti-mouse/Cy5 (Jackson, West Grove, PA, USA), goat anti-rabbit/Alexa-568, goat antirabbit/Alexa-633, goat anti-mouse/Alexa-568 and donkey antisheep/Alexa-568 (Molecular Probes, Eugene, OR, USA). Reagents were from Sigma unless stated otherwise.-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Antibodies and reage>>>ss stated otherwise.
218	Plasmids. RNAi vectors were generated as described (Lens et al, 2003), and were based on 50-TGACACGGAGATTGCCAAC-30 (INCENP) and 50-CAGCTGACCTGGATATCAC-30 (Borealin). Human INCENP and Borealin were PCR amplified from a T-cell 0:00 Borealin siRNA cDNA library using a 50 primer containing a VSV tag and cloned into pcDNA3.1 (INCENP) or into pCR3 containing an N-terminal VSV tag (Borealin). RNAi-resistant INCENP and Borealin constructs were generated using the QuickChanges Mutagenesis kit (Stratagene, La Jolla, CA, USA). For INCENP47­920, INCENP was digested with EcoRI/XhoI. This fragment was cloned into pCR3 containing an N-terminal VSV tag. For INCENP1­47, INCENP was digested with HindIII/EcoRI, and the product was cloned into pcDNA3.1. For Survivin­INCENP47­920, Survivin was ligated to the N terminus of INCENP47­920 after EcoRI digestion. For the Flow cytometry and time-lapse microscopy. The percentage of mitotic cells (MPM-2 positivity) and cell cycle distribution of spectrin­GFP-transfected cells were determined by flow cytometry, as described (Smits et al, 2000). H2B­GFP-expressing cells were followed by time-lapse microscopy, as described (Lens et al, 2003). Immunoblotting, immunoprecipitation and immunofluorescence. Immunoblotting was carried out as described (Smits et al, 2000). For immunoprecipitation, cells were lysed in E1A lysis buffer with protease inhibitors (Completes, Roche, Indianapolis, IN, USA) for 30 min at 4 1C (Smits et al, 2000). GFP-tagged proteins were immunoprecipitated with 3 mg anti-GFP precoupled to protein G­ Sepharose (Amersham Biosciences, Pittsburg, CA, USA). Immunofluorescence was carried out as described (Lens et al, 2003). DNA was stained with 4,6-diamidino-2-phenylindole. Supplementary information is available at EMBO reports online (http://www.emboreports.org).-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_parr	Plasmids. RNAi vecto>>>ww.emboreports.org).
219	Fig 2 | Survivin is a determining factor for mitotic localization of the chromosome passenger complex. (A) U2OS cells were transfected with the indicated plasmids. Puromycin was added to enrich for transfected cells. Lysates were analysed by western blotting. (B,C) Cells were transfected with the indicated short interfering RNA (siRNA) plasmids combined with the indicated VSV-tagged constructs. (B) After release from a thymidine block, cells were treated with Taxol and collected for fluorescence-activated cell sorting (FACS) analysis 18 h later. Mitotic percentages were determinedby MPM-2 staining. (C) Cells were collected for FACS analysis 60 h after transfection. (D,E) Cells grown on coverslips were transfected with mock or inner centromere protein (INCENP)/Survivin siRNA plasmids, VSV­Survivin­INCENP47­920-sil and H2B­GFP (green fluorescent protein). Immunostainings were carried out with the indicated antibodies. DNA (H2B­GFP) is depicted in blue. (F) Cells grown on coverslips were transfected with the indicated plasmids and H2B­GFP. Cells were stained with anti-Aurora-B, and the percentage of H2B­GFP-positive prometaphase cells in which Aurora-B localized on the centromeres was scored. The number of cells counted per condition is indicated. (G) Time-lapse images of a representative cell transfected with INCENP siRNA plasmid, Survivin­INCENP47­920-sil­GFP and H2B­diHcRed. Differential interference contrast (DIC), GFP and diHcRed signals are shown. (H) Time-lapse imaging of cells transfected with mock (upper panel) or Survivin siRNA plasmid (lower panel), INCENP1­47-GFP and H2B­diHcRed. Scale bars, 5 mm.-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_capfig	Fig 2 | Survivin is >>>d. Scale bars, 5 mm.
220	c-->id=5, page=0, size=11, fam=Times, col=#000000, type=parr, textLines=294--->[]--->font_capfig	c>>>c
221	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
222	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
223	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
224	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
225	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
226	88-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	88>>>88
227	0:18-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0:18>>>0:18
228	0:24-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0:24>>>0:24
229	0:30-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0:30>>>0:30
230	0:42-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0:42>>>0:42
231	0:48-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0:48>>>0:48
232	2:18-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2:18>>>2:18
233	GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP>>>GFP
234	H2B--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	H2B->>>H2B-
235	diHcRed-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	diHcRed>>>diHcRed
236	DIC-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	DIC>>>DIC
237	INCENP siRNA +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA +>>>INCENP siRNA +
238	VSV-Survivin-INCENP47-920-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-Survivin-INCENP4>>>vin-INCENP47-920-GFP
239	GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP>>>GFP
240	DIC-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	DIC>>>DIC
241	H2B diHcRed-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	H2B diHcRed>>>H2B diHcRed
242	INCENP1-47 - GFP-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP1-47 - GFP>>>INCENP1-47 - GFP
243	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
244	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
245	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
246	Anti-Cdk4-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Cdk4>>>Anti-Cdk4
247	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
248	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
249	VSV-INCENP-sil VSV-Survivin--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-INCENP-sil VSV-S>>>NP-sil VSV-Survivin-
250	INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP47-920-sil>>>INCENP47-920-sil
251	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
252	Survivin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Survivin siRNA>>>Survivin siRNA
253	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
254	H2BGFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	H2BGFP>>>H2BGFP
255	Merge-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Merge>>>Merge
256	VSVSurvivin INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSVSurvivin INCENP>>>VSVSurvivin INCENP
257	47 -920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	47 -920-sil>>>47 -920-sil
258	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
259	Survivin siRNA + INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Survivin siRNA + INC>>>siRNA + INCENP siRNA
260	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
261	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
262	Survivin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Survivin siRNA>>>Survivin siRNA
263	INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA>>>INCENP siRNA
264	Survivin siRNA + INCENP siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Survivin siRNA + INC>>>siRNA + INCENP siRNA
265	Empty vector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Empty vector>>>Empty vector
266	VSV-Survivin--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-Survivin->>>VSV-Survivin-
267	INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP47-920-sil>>>INCENP47-920-sil
268	4n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4n>>>4n
269	2n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2n>>>2n
270	8n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8n>>>8n
271	4n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4n>>>4n
272	2 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2 n>>>2 n
273	8n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8n>>>8n
274	5.7%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	5.7%>>>5.7%
275	18.5%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	18.5%>>>18.5%
276	21.9%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	21.9%>>>21.9%
277	6.4%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	6.4%>>>6.4%
278	11.6%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	11.6%>>>11.6%
279	10.2%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	10.2%>>>10.2%
280	7.7%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	7.7%>>>7.7%
281	4 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4 n>>>4 n
282	2n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2n>>>2n
283	8n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8n>>>8n
284	VSV-Survivin-sil-->id=2146, page=3, size=7, fam=Times, col=#000000, type=note, textLines=2--->[]--->font_note	VSV-Survivin-sil>>>VSV-Survivin-sil
285	7.8%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	7.8%>>>7.8%
286	17.5%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	17.5%>>>17.5%
287	10.4%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	10.4%>>>10.4%
288	18.3%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	18.3%>>>18.3%
289	Empty vector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Empty vector>>>Empty vector
290	VSV-INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-INCENP-sil>>>VSV-INCENP-sil
291	VSV-Surv-INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	VSV-Surv-INCENP47-92>>>urv-INCENP47-920-sil
292	VSV-Survivin-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-Survivin-sil>>>VSV-Survivin-sil
293	Mitotic-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mitotic>>>Mitotic
294	cells-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	cells>>>cells
295	(%) 40-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	(%) 40>>>(%) 40
296	20-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	20>>>20
297	0-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0>>>0
298	60-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	60>>>60
299	Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Survivin>>>Survivin
300	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
301	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP>>>INCENP
302	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
303	Mock-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock>>>Mock
304	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
305	Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Survivin>>>Survivin
306	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
307	+ INCEN-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ INCEN>>>+ INCEN
308	PsiRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	PsiRNA>>>PsiRNA
309	Merge-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Merge>>>Merge
310	DNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	DNA>>>DNA
311	INCENP siRNA + VSV-Survivin --->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP siRNA + VSV-S>>>RNA + VSV-Survivin -
312	INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP47-920-sil>>>INCENP47-920-sil
313	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
314	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
315	Anti-Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Aurora-B>>>Anti-Aurora-B
316	Aur-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Aur>>>Aur
317	-B-positive-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-B-positive>>>-B-positive
318	cells-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	cells>>>cells
319	(%)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	(%)>>>(%)
320	0-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0>>>0
321	40-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	40>>>40
322	60-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	60>>>60
323	80-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	80>>>80
324	100-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	100>>>100
325	20-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	20>>>20
326	Mock-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock>>>Mock
327	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
328	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP>>>INCENP
329	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
330	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP>>>INCENP
331	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
332	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
333	VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV>>>VSV
334	-INCENP-silINCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-INCENP-silINCENP>>>-INCENP-silINCENP
335	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
336	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
337	VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV>>>VSV
338	-Surv-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-Surv>>>-Surv
339	-INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-INCENP>>>-INCENP
340	47- 920-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	47- 920>>>47- 920
341	-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-sil>>>-sil
342	n=47-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	n=47>>>n=47
343	n=96-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	n=96>>>n=96
344	n=87-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	n=87>>>n=87
345	n=62-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	n=62>>>n=62
346	+ Taxol-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ Taxol>>>+ Taxol
347	G-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	G>>>G
348	H-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	H>>>H
349	F-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	F>>>F
350	D-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	D>>>D
351	C-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	C>>>C
352	E-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	E>>>E
353	AB-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	AB>>>AB
354	23.9% Borealin siRNA cDNA library using a 50 primer containing a VSV tag and cloned into pcDNA3.1 (INCENP) or into pCR3 containing an N-terminal VSV tag (Borealin). RNAi-resistant INCENP and Borealin constructs were generated using the QuickChanges Mutagenesis kit (Stratagene, La Jolla, CA, USA). For INCENP47­920, INCENP was digested with EcoRI/XhoI. This fragment was cloned into pCR3 containing an N-terminal VSV tag. For INCENP1­47, INCENP was digested with HindIII/EcoRI, and the product was cloned into pcDNA3.1. For Survivin­INCENP47­920, Survivin was ligated to the N terminus of INCENP47­920 after EcoRI digestion. For the Flow cytometry and time-lapse microscopy. The percentage of mitotic cells (MPM-2 positivity) and cell cycle distribution of spectrin­GFP-transfected cells were determined by flow cytometry, as described (Smits et al, 2000). H2B­GFP-expressing cells were followed by time-lapse microscopy, as described (Lens et al, 2003). Immunoblotting, immunoprecipitation and immunofluorescence. Immunoblotting was carried out as described (Smits et al, 2000). For immunoprecipitation, cells were lysed in E1A lysis buffer with protease inhibitors (Completes, Roche, Indianapolis, IN, USA) for 30 min at 4 1C (Smits et al, 2000). GFP-tagged proteins were immunoprecipitated with 3 mg anti-GFP precoupled to protein G­ Sepharose (Amersham Biosciences, Pittsburg, CA, USA). Immunofluorescence was carried out as described (Lens et al, 2003). DNA was stained with 4,6-diamidino-2-phenylindole. Supplementary information is available at EMBO reports online (http://www.emboreports.org).-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	23.9% Borealin siRNA>>>ww.emboreports.org).
355	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
356	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
357	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
358	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
359	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
360	89-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	89>>>89
361	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
362	VSV­Borealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV­Borealin>>>VSV­Borealin
363	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
364	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
365	Anti-Tubulin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Tubulin>>>Anti-Tubulin
366	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
367	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
368	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
369	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
370	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
371	4 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4 n>>>4 n
372	2n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2n>>>2n
373	8 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8 n>>>8 n
374	4n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4n>>>4n
375	2n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2n>>>2n
376	8 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8 n>>>8 n
377	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
378	Borealin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Borealin siRNA>>>Borealin siRNA
379	Empty vector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Empty vector>>>Empty vector
380	VSV-Survivin--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-Survivin->>>VSV-Survivin-
381	INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	INCENP47-920-sil>>>INCENP47-920-sil
382	4.3%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4.3%>>>4.3%
383	8.7%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8.7%>>>8.7%
384	9.4%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	9.4%>>>9.4%
385	26.7%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	26.7%>>>26.7%
386	D-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	D>>>D
387	F-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	F>>>F
388	E-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	E>>>E
389	40-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	40>>>40
390	20-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	20>>>20
391	0-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0>>>0
392	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	INCENP>>>INCENP
393	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
394	Mock-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock>>>Mock
395	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
396	Bor-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Bor>>>Bor
397	ealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	ealin>>>ealin
398	siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	siRNA>>>siRNA
399	+Taxol-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+Taxol>>>+Taxol
400	Mitotic-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mitotic>>>Mitotic
401	cells-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	cells>>>cells
402	(%)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	(%)>>>(%)
403	Empty vector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Empty vector>>>Empty vector
404	VSV-INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-INCENP-sil>>>VSV-INCENP-sil
405	VSV-Surv-INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	VSV-Surv-INCENP47-92>>>urv-INCENP47-920-sil
406	Mock siRNA Borealin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA Borealin >>>siRNA Borealin siRNA
407	Mock siRNA Borealin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA Borealin >>>siRNA Borealin siRNA
408	VSV-Survivin-sil VSV-Borealin-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	VSV-Survivin-sil VSV>>>sil VSV-Borealin-sil
409	Anti-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-GFP>>>Anti-GFP
410	Anti-Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Survivin>>>Anti-Survivin
411	Anti-VSV (Borealin)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV (Borealin)>>>Anti-VSV (Borealin)
412	Empty vector-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Empty vector>>>Empty vector
413	+ -- + ---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ -- + -->>>+ -- + --
414	+ -- +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ -- +>>>+ -- +
415	-- + -- + -- + -- + -- + -- + -- + -- +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-- + -- + -- + -- + >>> -- + -- + -- + -- +
416	Exog. Endog.-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Exog. Endog.>>>Exog. Endog.
417	Total lysate-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Total lysate>>>Total lysate
418	12-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	12>>>12
419	3 4 5 6-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	3 4 5 6>>>3 4 5 6
420	78 9 10 11 12-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	78 9 10 11 12>>>78 9 10 11 12
421	GFP-INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP-INCENP>>>GFP-INCENP
422	++ + + + +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++ + + + +>>>++ + + + +
423	+ + + +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ + + +>>>+ + + +
424	+ +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ +>>>+ +
425	4 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	4 n>>>4 n
426	2n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	2n>>>2n
427	8 n-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	8 n>>>8 n
428	5.4%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	5.4%>>>5.4%
429	35.7%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	35.7%>>>35.7%
430	6.9%-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	6.9%>>>6.9%
431	Mock siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA>>>Mock siRNA
432	Borealin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Borealin siRNA>>>Borealin siRNA
433	Borealin siRNA + VSV -Borealin-sil Mock siRNA Borealin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Borealin siRNA + VSV>>>siRNA Borealin siRNA
434	Anti-Borealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Borealin>>>Anti-Borealin
435	H2B-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	H2B-GFP>>>H2B-GFP
436	Merge-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Merge>>>Merge
437	Mock siRNA Borealin siRNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Mock siRNA Borealin >>>siRNA Borealin siRNA
438	0-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	0>>>0
439	20-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	20>>>20
440	40-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	40>>>40
441	60-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	60>>>60
442	80-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	80>>>80
443	100 n=18-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	100 n=18>>>100 n=18
444	n=127-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	n=127>>>n=127
445	Bor-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Bor>>>Bor
446	ealin-positive cells-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	ealin-positive cells>>>ealin-positive cells
447	(%)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	(%)>>>(%)
448	GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP>>>GFP
449	Anti-Borealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Borealin>>>Anti-Borealin
450	Anti-Borealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-Borealin>>>Anti-Borealin
451	CREST-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	CREST>>>CREST
452	Merge-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Merge>>>Merge
453	GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP>>>GFP
454	DNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	DNA>>>DNA
455	Merge-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Merge>>>Merge
456	Borealin siRNA +-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Borealin siRNA +>>>Borealin siRNA +
457	VSV -Survivin-INCENP47-920-sil-GFP-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	VSV -Survivin-INCENP>>>INCENP47-920-sil-GFP
458	Total lysate-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Total lysate>>>Total lysate
459	IP: Anti-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	IP: Anti-GFP>>>IP: Anti-GFP
460	IP: Anti-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	IP: Anti-GFP>>>IP: Anti-GFP
461	GFP-INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP-INCENP>>>GFP-INCENP
462	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
463	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
464	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
465	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
466	---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-->>>--
467	++ --->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++ ->>>++ -
468	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
469	++-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++>>>++
470	---->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	-->>>--
471	++ --->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	++ ->>>++ -
472	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
473	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
474	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
475	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
476	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
477	+-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+>>>+
478	--->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	->>>-
479	+ VSV-Borealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	+ VSV-Borealin>>>+ VSV-Borealin
480	Anti-GFP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-GFP>>>Anti-GFP
481	Anti-VSV-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Anti-VSV>>>Anti-VSV
482	GFP-INCENP47-920-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_parr	GFP-INCENP47-920>>>GFP-INCENP47-920
483	123 4 5 678-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	123 4 5 678>>>123 4 5 678
484	GFP CREST DNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP CREST DNA>>>GFP CREST DNA
485	Merge w/DNA-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	Merge w/DNA>>>Merge w/DNA
486	GFP Borealin CREST-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	GFP Borealin CREST>>>GFP Borealin CREST
487	IgG-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_parr	IgG>>>IgG
488	AB-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	AB>>>AB
489	C-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	C>>>C
490	G-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	G>>>G
491	H cDNA library using a 50 primer containing a VSV tag and cloned into pcDNA3.1 (INCENP) or into pCR3 containing an N-terminal VSV tag (Borealin). RNAi-resistant INCENP and Borealin constructs were generated using the QuickChanges Mutagenesis kit (Stratagene, La Jolla, CA, USA). For INCENP47­920, INCENP was digested with EcoRI/XhoI. This fragment was cloned into pCR3 containing an N-terminal VSV tag. For INCENP1­47, INCENP was digested with HindIII/EcoRI, and the product was cloned into pcDNA3.1. For Survivin­INCENP47­920, Survivin was ligated to the N terminus of INCENP47­920 after EcoRI digestion. For the Flow cytometry and time-lapse microscopy. The percentage of mitotic cells (MPM-2 positivity) and cell cycle distribution of spectrin­GFP-transfected cells were determined by flow cytometry, as described (Smits et al, 2000). H2B­GFP-expressing cells were followed by time-lapse microscopy, as described (Lens et al, 2003). Immunoblotting, immunoprecipitation and immunofluorescence. Immunoblotting was carried out as described (Smits et al, 2000). For immunoprecipitation, cells were lysed in E1A lysis buffer with protease inhibitors (Completes, Roche, Indianapolis, IN, USA) for 30 min at 4 1C (Smits et al, 2000). GFP-tagged proteins were immunoprecipitated with 3 mg anti-GFP precoupled to protein G­ Sepharose (Amersham Biosciences, Pittsburg, CA, USA). Immunofluorescence was carried out as described (Lens et al, 2003). DNA was stained with 4,6-diamidino-2-phenylindole. Supplementary information is available at EMBO reports online (http://www.emboreports.org).-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_parr	H cDNA library using>>>ww.emboreports.org).
492	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
493	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
494	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
495	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
496	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
497	90-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	90>>>90
498	Fig 3 | Survivin can target a functional chromosome passenger complex in the absence of Borealin, but interaction with inner centromere protein (INCENP) depends on Borealin. (A) U2OS cells were transfected with the indicated plasmids, puromycin was added to enrich for transfected cells and lysates were analysed by western blotting. (B­E) Cells were transfected with the indicated plasmids. (C) Cells grown on coverslips were transfected with mock or Borealin short interfering RNA (siRNA) plasmid and H2B­GFP (green fluorescent protein). Immunostainings were carried out with anti-Borealin, and the percentage of H2B­GFP-positive prometaphase cells in which Borealin localized on centromeres was scored. The number of cells counted per condition is indicated. (B,D) Cells were grown asynchronously for 60 h and collected for fluorescence-activated cell sorting (FACS) analysis. (E) Taxol was added after thymidine release, and cells were collected for FACS analysis 18 h later. Mitotic percentages were determined by MPM-2 staining. (F) Cells grown on coverslips were transfected with Borealin siRNA plasmid and VSV­Survivin­INCENP47­920-sil­GFP. Immunostainings were carried out with the indicated antibodies, and DNA was stained with 4,6-diamidino-2-phenylindole. Different colours used for the two merge pictures are indicated. (G) Human embryonic kidney 293 cells were transfected with 5 mg of the indicated constructs. After 48 h, cells were collected, immunoprecipitations were carried out with anti-GFP and samples were analysed by western blotting. (H) U2OS cells were transfected with the indicated plasmids. At 18 h after transfection, puromycin was added and 36 h later, cells were collected . Immunoprecipitations were carried out with anti-GFP and western blots were probed with the indicated antibodies. Scale bars, 5 mm.-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_capfig	Fig 3 | Survivin can>>>s. Scale bars, 5 mm.
499	Indirect centromeretargeting domain IN-box-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Indirect centromeret>>>geting domain IN-box
500	147-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	147>>>147
501	920-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	920>>>920
502	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	INCENP>>>INCENP
503	INCENP-sil-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	INCENP-sil>>>INCENP-sil
504	47-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	47>>>47
505	920-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	920>>>920
506	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	INCENP>>>INCENP
507	INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_note	INCENP47-920-sil>>>INCENP47-920-sil
508	47-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	47>>>47
509	920-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	920>>>920
510	Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Survivin>>>Survivin
511	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	INCENP>>>INCENP
512	Survivin-INCENP47-920-sil-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_note	Survivin-INCENP47-92>>>vin-INCENP47-920-sil
513	147-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	147>>>147
514	INCENP1-47-->id=786, page=1, size=5, fam=Times, col=#000000, type=parrnote, textLines=23--->[]--->font_note	INCENP1-47>>>INCENP1-47
515	* *-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	* *>>>* *
516	* *-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	* *>>>* *
517	* *-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	* *>>>* *
518	Indirect centromeretargeting domain IN-box-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Indirect centromeret>>>geting domain IN-box
519	147-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	147>>>147
520	920-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	920>>>920
521	INCENP-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	INCENP>>>INCENP
522	Survivin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Survivin>>>Survivin
523	Anaphase-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Anaphase>>>Anaphase
524	(central spindle)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	(central spindle)>>>(central spindle)
525	Prometaphase (centromeres)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Prometaphase (centro>>>aphase (centromeres)
526	Localization-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Localization>>>Localization
527	Interaction, stabilization-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Interaction, stabili>>>ction, stabilization
528	RNAi-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	RNAi>>>RNAi
529	Aurora-B-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Aurora-B>>>Aurora-B
530	Interaction, stabilization-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Interaction, stabili>>>ction, stabilization
531	Borealin-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Borealin>>>Borealin
532	Cytokinesis (midbody)-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	Cytokinesis (midbody>>>ytokinesis (midbody)
533	A-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_note	A>>>A
534	B-->id=2, page=0, size=10, fam=Times, col=#000000, type=parrnote, textLines=21--->[]--->font_note	B>>>B
535	Fig 4 | Overview of inner centromere protein constructs and model. (A) Schematic representation of inner centromere protein (INCENP) constructs. Survivin and Aurora-B binding domains are depicted. Introduced RNA interference (RNAi) silent mutations are shown. (B) Model showing the proposed upstream targeting role of Survivin in the chromosome passenger complex.-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_capfig	Fig 4 | Overview of >>>e passenger complex.
536	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
537	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
538	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
539	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
540	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
541	91-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	91>>>91
542	ACKNOWLEDGEMENTS We thank B.v.d. Weerdt, M.v. Vugt and G. Kops for comments on the manuscript, T.v. Harn for technical support and S. Taylor and H. Funabiki for the gift of reagents. This work was supported by the Dutch Cancer Society (NKI 2002-2764).-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	ACKNOWLEDGEMENTS We >>>ety (NKI 2002-2764).
543	Ainsztein AM, Kandels-Lewis SE, Mackay AM, Earnshaw WC (1998) INCENP centromere and spindle targeting: identification of essential conserved motifs and involvement of heterochromatin protein HP1. J Cell Biol 143: 1763­1774-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Ainsztein AM, Kandel>>> Biol 143: 1763­1774
544	Biggins S, Murray AW (2001) The budding yeast protein kinase Ipl1/Aurora allows the absence of tension to activate the spindle checkpoint. Genes Dev 15: 3118­3129-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Biggins S, Murray AW>>>es Dev 15: 3118­3129
545	Gassmann R, Carvalho A, Henzing AJ, Ruchaud S, Hudson DF, Honda R, Nigg EA, Gerloff DL, Earnshaw WC (2004) Borealin: a novel chromosomal passenger required for stability of the bipolar mitotic spindle. J Cell Biol 166: 179­191-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Gassmann R, Carvalho>>>ll Biol 166: 179­191
546	Hauf S, Cole RW, LaTerra S, Zimmer C, Schnapp G, Walter R, Heckel A, van Meel J, Rieder CL, Peters JM (2003) The small molecule Hesperadin reveals a role for Aurora B in correcting kinetochore­microtubule attachment and in maintaining the spindle assembly checkpoint. J Cell Biol 161: 281­294 Honda R, Korner R, Nigg EA (2003) Exploring the functional interactions between Aurora B, INCENP, and survivin in mitosis. Mol Biol Cell 14: 3325­3341-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Hauf S, Cole RW, LaT>>>l Cell 14: 3325­3341
547	Kaitna S, Mendoza M, Jantsch-Plunger V, Glotzer M (2000) Incenp and an aurora-like kinase form a complex essential for chromosome segregation and efficient completion of cytokinesis. Curr Biol 10: 1172­1181-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Kaitna S, Mendoza M,>>>r Biol 10: 1172­1181
548	Lampson MA, Renduchitala K, Khodjakov A, Kapoor TM (2004) Correcting improper chromosome­spindle attachments during cell division. Nat Cell Biol 6: 232­237-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Lampson MA, Renduchi>>>Cell Biol 6: 232­237
549	Lens SM, Medema RH (2003) The survivin/Aurora B complex: its role in coordinating tension and attachment. Cell Cycle 2: 507­510-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Lens SM, Medema RH (>>>ell Cycle 2: 507­510
550	Lens SM, Wolthuis RM, Klompmaker R, Kauw J, Agami R, Brummelkamp T, Kops G, Medema RH (2003) Survivin is required for a sustained spindle checkpoint arrest in response to lack of tension. EMBO J 22: 2934­2947-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Lens SM, Wolthuis RM>>>EMBO J 22: 2934­2947
551	Nicklas RB (1997) How cells get the right chromosomes. Science 275: 632­637-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_parr	Nicklas RB (1997) Ho>>>Science 275: 632­637
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562	CPC targeting mediated by Survivin-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	CPC targeting mediat>>>mediated by Survivin
563	G. Vader et al-->id=3, page=0, size=9, fam=Times, col=#000000, type=parrnote, textLines=143--->[]--->font_note	G. Vader et al>>>G. Vader et al
564	EMBO reports VOL 7 | NO 1 | 2006-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	EMBO reports VOL 7 |>>> VOL 7 | NO 1 | 2006
565	&2006 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	&2006 EUROPEAN MOLEC>>>BIOLOGY ORGANIZATION
566	scientific report-->id=787, page=1, size=36, fam=Times, col=#008a33, type=title, textLines=7--->[]--->font_note	scientific report>>>scientific report
567	92-->id=6, page=0, size=8, fam=Times, col=#000000, type=parrnote, textLines=624--->[]--->font_note	92>>>92