- Email: [email protected]
The Chromosomal Passenger Complex Takes Center Stage during Mitosis
Developmental Cell, Vol. 7, 145–153, August, 2004, Copyright 2004 by Cell Press
Previews
The Chromosomal Passenger Complex Takes Center Stage during Mitosis The chromosomal passenger complex plays important roles in key mitotic events, including chromosome biorientation, the spindle assembly checkpoint, and cytokinesis. Two groups now report the identification of a novel component of the Incenp/survivin/auroraB complex (Gassmann et al., 2004; Sampath et al., 2004) and show that different subcomplexes may exist during mitosis. Exciting data support the involvement of the passenger complex in yet another key event, the assembly of the mitotic spindle. Microtubules are intrinsically dynamic polymers that perform multiple tasks in the cell. During mitosis, they organize a spindle-shaped apparatus that segregates the chromosomes. The spindle is made of two antiparallel arrays of microtubules with minus-ends focused into the two spindle poles and plus-ends interacting with each other and with the chromosomes. Successful cell division relies on bipolar attachment of each chromosome to the two spindle poles before segregation and on the timely and spatially controlled splitting of the cell during cytokinesis. It is now well established that in organisms ranging from yeast to mammals a complex of Incenp/survivin/auroraB termed the chromosomal passenger complex (CPC) is required for these various mitotic events (Andrews et al., 2003). The name of the complex comes from its characteristic and striking localization pattern. In the early phases of mitosis, the complex associates with chromosome arms and centromeres where it accumulates at metaphase. During anaphase, the complex relocalizes completely to the spindle midzone. The CPC in fact is a sophisticated machinery for targeting and modulating in time and in space the activity of Aurora B, one of the kinases involved in the regulation of the complex series of events required to segregate chromosomes faithfully during mitosis (Andrews et al., 2003). At the kinetochore, fine regulation of Aurora B activity facilitates the turnover of kinetochore-microtubule interactions, permitting error corrections and ultimately generating correct bipolar attachment (Tanaka, 2002). This mechanism most certainly involves MCAK, a microtubule depolymerase also localized to the kinetochore. In vitro, phosphorylation of MCAK by Aurora B downregulates MCAK depolymerase activity (Gorbsky, 2004). In different types of screens to identify novel proteins associated with mitotic chromosomes, two groups, one working with Xenopus (Sampath et al., 2004) and the other with human cells (Gassmann et al., 2004), found proteins displaying the same dynamic localization as chromosomal passenger proteins throughout mitosis. In keeping with Aurora field terminology, the Xenopus
protein was named Dasra A (Sampath et al., 2004) and the human protein Borealin (Gassmann et al., 2004). The sequence of these two proteins is 23% identical. In addition, through sequence homology searches, Sampath et al. identified another related protein in Xenopus that they called Dasra B. This protein is 42% identical to Borealin. Dasra/Borealin coimmunoprecipitate with components of the CPC, and Borealin interacts directly with Incenp and survivin in vitro. In addition, Borealin is phosphorylated by Aurora B. Interestingly, Dasra B/Borealin is required for centromere targeting of the CPC but not for its relocalization to the spindle midzone in anaphase (Gassmann et al., 2004). As expected, therefore, Dasra B/Borealin is required for CPC function in kinetochore attachment error correction. RNA interference experiments also indicate that Borealin is required for bipolar spindle stability (Gassmann et al., 2004), suggesting that the CPC may have another unexpected role during mitosis. This role is revealed by the experiments performed in the Xenopus egg extract system by Sampath et al. Strikingly, depletion of CPC from Xenopus egg extracts results in the inhibition of microtubule assembly around chromatin, therefore preventing spindle assembly. The major catastrophe-promoting factor in the M-phase egg extract is MCAK (also called XKCM1) (Walczak et al., 1996). Its activity is largely responsible for the high turnover of microtubules in the M-phase cytoplasm; in fact, microtubules become so unstable that their existence is unlikely in the absence of active microtubule nucleation. To test whether the lack of microtubule assembly around chromatin in the absence of CPC was due to MCAK activity, Sampath et al. depleted MCAK and Incenp from the egg extract and found that chromatin-induced microtubule assembly was restored. How do these results fit into the current view of the mechanism of spindle assembly? Work done by several labs supports a central role for chromosomes in spindle formation both in the egg extract system and in vivo (Quimby and Dasso, 2003). The presence of RCC1, the GTP exchange factor for Ran, on the chromatin results in the enrichment of the GTP bound form of Ran (RanGTP) around the chromatin (Kalab et al., 2002). RanGTP releases factors required for microtubule nucleation, stabilization, and organization from the inhibitory interaction with importins, thereby promoting microtubule assembly and spindle formation around the chromosomes. The current view is thus that the Ran pathway is the major determinant for chromosome-induced spindle assembly (Karsenti and Vernos, 2001). The results from Sambath et al. suggest that this is an oversimplification. Experiments performed to examine the possible functional interactions between the Ran and the CPC-MCAK pathways indicate that in fact they probably function in parallel. Blocking the production of RanGTP in the M-phase extract by addition of a dominant-negative mutant form of Ran inhibits microtubule assembly around the chromatin whether CPC and MCAK are present or not. Conversely, addition of RanGTP to an M-phase extract triggers microtubule aster formation even in the
Developmental Cell 146
Figure 1. Pathways for Microtubule Assembly around Mitotic Chromatin In M-phase cytoplasm, various factors (SAF, Spindle Assembly Factors) are bound to importins through their NLS and therefore kept inactive. The microtubule depolymerase MCAK is active in its dephosphorylated form and OP18 sequesters tubulin dimers, lowering the concentration of available tubulin. As a result, microtubules do not assemble. The presence of RCC1, the nucleotide exchange factor for Ran, on the chromatin promotes the formation of a RanGTP gradient (Kalab et al., 2002). In the vicinity of the chromatin, RanGTP disrupts the interaction of SAF with importins. One of the SAF, TPX2, promotes microtubule nucleation (1). In a second step (2), several pathways favor microtubule elongation and stabilization: a, some SAF participate in microtubule stabilization; b, the CPC phosphorylates MCAK bound to the chromatin and presumably soluble MCAK in its proximity. This inhibits its microtubule depolymerase activity therefore promoting microtubule elongation; c, OP18 is phosphorylated by the chromosome-associated kinase Polo, releasing tubulin dimers and increasing the critical concentration of tubulin available for microtubule polymerization (Cassimeris, 2002).
absence of CPC. However, microtubules do not assemble in the absence of CPC (therefore presumably with high MCAK microtubule depolymerization activity) even if RanGTP is present around the chromatin. The picture that emerges is the following (Figure 1). M-phase cytoplasm does not support microtubule assembly except in the presence of active microtubule nucleation. Even in this situation, microtubules are short and extremely dynamic. Chromosomes modify the cytoplasm around them to generate a local cytoplasmic state in which microtubules are nucleated and partially stabilized. Previous work and the results from Sambath et al. confirm that microtubule nucleation is under the control of the Ran pathway. RanGTP also promotes microtubule stabilization but the results from Sampath et al. show that this is not sufficient for generating a microtubule population around the chromatin. The CPC plays an essential function, presumably through the inactivation of MCAK microtubules depolymerase activity. A third pathway involves the inhibition of the small tubulin sequestering molecule OP18/stathmin by phosphorylation around the chromatin, increasing the effective concentration of tubulin around the chromosome. The CPC has clearly emerged as a key player for the regulation of microtubule behavior and function throughout mitosis. The exciting new results discussed here
promise further interesting discoveries in the future to help us unravel the complex pathways that lead to the successful division of the cell. Isabelle Vernos EMBL Cell Biology and Biophysics Program Heidelberg 69 117 Germany Selected Reading Andrews, P.D., Knatko, E., Moore, W.J., and Swedlow, J.R. (2003). Curr. Opin. Cell Biol. 15, 672–683. Cassimeris, L. (2002). Curr. Opin. Cell Biol. 14, 18–24. Gassmann, R., Carvalho, A., Henzing, A.J., Ruchaud, S., Hudson, D.F., Honda, R., Nigg, E.A., Gerloff, D.L., and Earnshaw, W.C. (2004). J. Cell Biol. 166, 179–191. Gorbsky, G.J. (2004). Curr. Biol. 14, R346–R348. Kalab, P., Weis, K., and Heald, R. (2002). Science 295, 2452–2456. Karsenti, E., and Vernos, I. (2001). Science 294, 543–547. Quimby, B.B., and Dasso, M. (2003). Curr. Opin. Cell Biol. 15, 338–344. Sampath, C.S., Ohi, R., Leismann, O., Salic, A., Pozniakovski, A., and Funabiki, H. (2004). Cell 118, 187–202. Tanaka, T.U. (2002). Curr. Opin. Cell Biol. 14, 365–371. Walczak, C., Mitchison, T.J., and Desai, A.B. (1996). Cell 84, 37–47.
Previews
The Chromosomal Passenger Complex Takes Center Stage during Mitosis The chromosomal passenger complex plays important roles in key mitotic events, including chromosome biorientation, the spindle assembly checkpoint, and cytokinesis. Two groups now report the identification of a novel component of the Incenp/survivin/auroraB complex (Gassmann et al., 2004; Sampath et al., 2004) and show that different subcomplexes may exist during mitosis. Exciting data support the involvement of the passenger complex in yet another key event, the assembly of the mitotic spindle. Microtubules are intrinsically dynamic polymers that perform multiple tasks in the cell. During mitosis, they organize a spindle-shaped apparatus that segregates the chromosomes. The spindle is made of two antiparallel arrays of microtubules with minus-ends focused into the two spindle poles and plus-ends interacting with each other and with the chromosomes. Successful cell division relies on bipolar attachment of each chromosome to the two spindle poles before segregation and on the timely and spatially controlled splitting of the cell during cytokinesis. It is now well established that in organisms ranging from yeast to mammals a complex of Incenp/survivin/auroraB termed the chromosomal passenger complex (CPC) is required for these various mitotic events (Andrews et al., 2003). The name of the complex comes from its characteristic and striking localization pattern. In the early phases of mitosis, the complex associates with chromosome arms and centromeres where it accumulates at metaphase. During anaphase, the complex relocalizes completely to the spindle midzone. The CPC in fact is a sophisticated machinery for targeting and modulating in time and in space the activity of Aurora B, one of the kinases involved in the regulation of the complex series of events required to segregate chromosomes faithfully during mitosis (Andrews et al., 2003). At the kinetochore, fine regulation of Aurora B activity facilitates the turnover of kinetochore-microtubule interactions, permitting error corrections and ultimately generating correct bipolar attachment (Tanaka, 2002). This mechanism most certainly involves MCAK, a microtubule depolymerase also localized to the kinetochore. In vitro, phosphorylation of MCAK by Aurora B downregulates MCAK depolymerase activity (Gorbsky, 2004). In different types of screens to identify novel proteins associated with mitotic chromosomes, two groups, one working with Xenopus (Sampath et al., 2004) and the other with human cells (Gassmann et al., 2004), found proteins displaying the same dynamic localization as chromosomal passenger proteins throughout mitosis. In keeping with Aurora field terminology, the Xenopus
protein was named Dasra A (Sampath et al., 2004) and the human protein Borealin (Gassmann et al., 2004). The sequence of these two proteins is 23% identical. In addition, through sequence homology searches, Sampath et al. identified another related protein in Xenopus that they called Dasra B. This protein is 42% identical to Borealin. Dasra/Borealin coimmunoprecipitate with components of the CPC, and Borealin interacts directly with Incenp and survivin in vitro. In addition, Borealin is phosphorylated by Aurora B. Interestingly, Dasra B/Borealin is required for centromere targeting of the CPC but not for its relocalization to the spindle midzone in anaphase (Gassmann et al., 2004). As expected, therefore, Dasra B/Borealin is required for CPC function in kinetochore attachment error correction. RNA interference experiments also indicate that Borealin is required for bipolar spindle stability (Gassmann et al., 2004), suggesting that the CPC may have another unexpected role during mitosis. This role is revealed by the experiments performed in the Xenopus egg extract system by Sampath et al. Strikingly, depletion of CPC from Xenopus egg extracts results in the inhibition of microtubule assembly around chromatin, therefore preventing spindle assembly. The major catastrophe-promoting factor in the M-phase egg extract is MCAK (also called XKCM1) (Walczak et al., 1996). Its activity is largely responsible for the high turnover of microtubules in the M-phase cytoplasm; in fact, microtubules become so unstable that their existence is unlikely in the absence of active microtubule nucleation. To test whether the lack of microtubule assembly around chromatin in the absence of CPC was due to MCAK activity, Sampath et al. depleted MCAK and Incenp from the egg extract and found that chromatin-induced microtubule assembly was restored. How do these results fit into the current view of the mechanism of spindle assembly? Work done by several labs supports a central role for chromosomes in spindle formation both in the egg extract system and in vivo (Quimby and Dasso, 2003). The presence of RCC1, the GTP exchange factor for Ran, on the chromatin results in the enrichment of the GTP bound form of Ran (RanGTP) around the chromatin (Kalab et al., 2002). RanGTP releases factors required for microtubule nucleation, stabilization, and organization from the inhibitory interaction with importins, thereby promoting microtubule assembly and spindle formation around the chromosomes. The current view is thus that the Ran pathway is the major determinant for chromosome-induced spindle assembly (Karsenti and Vernos, 2001). The results from Sambath et al. suggest that this is an oversimplification. Experiments performed to examine the possible functional interactions between the Ran and the CPC-MCAK pathways indicate that in fact they probably function in parallel. Blocking the production of RanGTP in the M-phase extract by addition of a dominant-negative mutant form of Ran inhibits microtubule assembly around the chromatin whether CPC and MCAK are present or not. Conversely, addition of RanGTP to an M-phase extract triggers microtubule aster formation even in the
Developmental Cell 146
Figure 1. Pathways for Microtubule Assembly around Mitotic Chromatin In M-phase cytoplasm, various factors (SAF, Spindle Assembly Factors) are bound to importins through their NLS and therefore kept inactive. The microtubule depolymerase MCAK is active in its dephosphorylated form and OP18 sequesters tubulin dimers, lowering the concentration of available tubulin. As a result, microtubules do not assemble. The presence of RCC1, the nucleotide exchange factor for Ran, on the chromatin promotes the formation of a RanGTP gradient (Kalab et al., 2002). In the vicinity of the chromatin, RanGTP disrupts the interaction of SAF with importins. One of the SAF, TPX2, promotes microtubule nucleation (1). In a second step (2), several pathways favor microtubule elongation and stabilization: a, some SAF participate in microtubule stabilization; b, the CPC phosphorylates MCAK bound to the chromatin and presumably soluble MCAK in its proximity. This inhibits its microtubule depolymerase activity therefore promoting microtubule elongation; c, OP18 is phosphorylated by the chromosome-associated kinase Polo, releasing tubulin dimers and increasing the critical concentration of tubulin available for microtubule polymerization (Cassimeris, 2002).
absence of CPC. However, microtubules do not assemble in the absence of CPC (therefore presumably with high MCAK microtubule depolymerization activity) even if RanGTP is present around the chromatin. The picture that emerges is the following (Figure 1). M-phase cytoplasm does not support microtubule assembly except in the presence of active microtubule nucleation. Even in this situation, microtubules are short and extremely dynamic. Chromosomes modify the cytoplasm around them to generate a local cytoplasmic state in which microtubules are nucleated and partially stabilized. Previous work and the results from Sambath et al. confirm that microtubule nucleation is under the control of the Ran pathway. RanGTP also promotes microtubule stabilization but the results from Sampath et al. show that this is not sufficient for generating a microtubule population around the chromatin. The CPC plays an essential function, presumably through the inactivation of MCAK microtubules depolymerase activity. A third pathway involves the inhibition of the small tubulin sequestering molecule OP18/stathmin by phosphorylation around the chromatin, increasing the effective concentration of tubulin around the chromosome. The CPC has clearly emerged as a key player for the regulation of microtubule behavior and function throughout mitosis. The exciting new results discussed here
promise further interesting discoveries in the future to help us unravel the complex pathways that lead to the successful division of the cell. Isabelle Vernos EMBL Cell Biology and Biophysics Program Heidelberg 69 117 Germany Selected Reading Andrews, P.D., Knatko, E., Moore, W.J., and Swedlow, J.R. (2003). Curr. Opin. Cell Biol. 15, 672–683. Cassimeris, L. (2002). Curr. Opin. Cell Biol. 14, 18–24. Gassmann, R., Carvalho, A., Henzing, A.J., Ruchaud, S., Hudson, D.F., Honda, R., Nigg, E.A., Gerloff, D.L., and Earnshaw, W.C. (2004). J. Cell Biol. 166, 179–191. Gorbsky, G.J. (2004). Curr. Biol. 14, R346–R348. Kalab, P., Weis, K., and Heald, R. (2002). Science 295, 2452–2456. Karsenti, E., and Vernos, I. (2001). Science 294, 543–547. Quimby, B.B., and Dasso, M. (2003). Curr. Opin. Cell Biol. 15, 338–344. Sampath, C.S., Ohi, R., Leismann, O., Salic, A., Pozniakovski, A., and Funabiki, H. (2004). Cell 118, 187–202. Tanaka, T.U. (2002). Curr. Opin. Cell Biol. 14, 365–371. Walczak, C., Mitchison, T.J., and Desai, A.B. (1996). Cell 84, 37–47.