- Email: [email protected]
Chromosome Segregation: Learning to Let Go
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an mTORC1 substrate that negatively regulates insulin signaling. Science 332, 1322–1326. 16. Chung, J., Kuo, C.J., Crabtree, G.R., and Blenis, J. (1992). Rapamycin-FKBP specifically blocks growth-dependent activation of and signaling by the 70 kd S6 protein kinases. Cell 69, 1227–1236. 17. Zhang, Y., and Zheng, X.F. (2012). mTOR-independent 4E-BP1 phosphorylation is associated with cancer resistance to mTOR kinase inhibitors. Cell Cycle 11, 594–603. 18. Ducker, G.S., Atreya, C.E., Simko, J.P., Hom, Y.K., Matli, M.R., Benes, C.H., Hann, B., Nakakura, E.K., Bergsland, E.K., Donner, D.B., et al. (2013). Incomplete inhibition of phosphorylation of 4E-BP1 as a mechanism of primary resistance to ATP-competitive mTOR
inhibitors. Oncogene http://dx.doi.org/10.1038/ onc.2013.92. 19. She, Q.B., Halilovic, E., Ye, Q., Zhen, W., Shirasawa, S., Sasazuki, T., Solit, D.B., and Rosen, N. (2010). 4E-BP1 is a key effector of the oncogenic activation of the AKT and ERK signaling pathways that integrates their function in tumors. Cancer Cell 18, 39–51. 20. Shin, S., Wolgamott, L., Tcherkezian, J., Vallabhapurapu, S., Yu, Y., Roux, P.P., and Yoon, S.O. (2013). Glycogen synthase kinase-3beta positively regulates protein synthesis and cell proliferation through the regulation of translation initiation factor 4E-binding protein 1. Oncogene http:// dx.doi.org/10.1038/onc.2013.113.
Chromosome Segregation: Learning to Let Go To ensure accurate chromosome segregation, cohesion between sister chromatids must be released in a controlled manner during mitosis. A new study reveals how distinct centromere populations of the cohesin protector Sgo1 are regulated by microtubule attachments, cyclin-dependent kinases, and the kinetochore kinase Bub1. Jonathan M.G. Higgins Dividing cells must convey the correct complement of chromosomes to their offspring. Eukaryotes accomplish this by maintaining cohesion between replicated sister chromatids until chromosomes are bi-oriented on the mitotic spindle. Only once this has been accomplished are the attachments between chromatids released, allowing them to be sorted accurately to opposite poles of the dividing cell. Clearly then, although sister chromatids may be inseparable at first, they must learn to let go when the time comes. A report from Liu, Jia and Yu in this issue of Current Biology [1] provides new insight into this process that may have broader implications for our understanding of inner centromere function. Cohesion between sister chromatids is maintained by cohesin complexes, together with regulators such as Sororin [2]. In vertebrate mitosis, cohesin is removed from chromosomes in two steps. In prophase, a mechanism involving phosphorylation of cohesin and Sororin by mitotic kinases removes the bulk of cohesin from chromosome arms (Figure 1). Cohesin at centromeres, however, is protected by Sgo1–PP2A phosphatase complexes that counteract phosphorylation of cohesin
and Sororin [3–5]. To fully separate chromatids at anaphase, the remaining cohesin is cleaved by the protease Separase [2]. This raises the question of how cleavage of centromeric cohesin is limited to anaphase. A simple possibility is that Separase only becomes active at anaphase, and that Sgo1 does not protect cohesin from cleavage in mitosis. However, it has been reported that Sgo1, when inappropriately maintained at inner centromeres, prevents Separase-mediated cohesin cleavage [6]. Also, at least in budding yeast, Sgo1–PP2A complexes may inhibit Separase more directly [7]. Therefore, it is important to understand how the localization and activity of Sgo1 are regulated. During prophase in mammalian cells, Sgo1 is found at inner centromeres (defined here as the area between the chromatin regions that contain centromeric histone CENP-A; Figure 1). As chromosomes become bi-oriented, Sgo1 appears to move outwards, relocating to two regions roughly coinciding with CENP-A-containing chromatin underlying kinetochores [1,6,8]. This movement of Sgo1 away from cohesin complexes located at inner centromeres might render cohesin susceptible to cleavage by Separase, and would provide a way to make removal of cohesin favorable
1Department of Cancer and Cell Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA. 2Institute for Research in Immunology and Cancer (IRIC), Universite´ de Montre´al, Montreal, Quebec H3C 3J7, Canada. 3Department of Pathology and Cell Biology, Faculty of Medicine, Universite´ de Montre´al, Montreal, Quebec, H3C 3J7, Canada. E-mail: [email protected], philippe. [email protected]
http://dx.doi.org/10.1016/j.cub.2013.08.030
only when chromosomes are correctly bi-oriented and microtubules exert tension across sister kinetochores [6]. How this relocation of Sgo1 is controlled, however, has been unknown. A number of ways to recruit Sgo1 to centromeres have been reported, but the relative contributions of these pathways are debated. It is widely accepted that Sgo1 is brought to centromeres when histone H2A is phosphorylated at Thr-120 (H2AT120ph) by the kinetochore kinase Bub1 [9,10], though the structural basis for this recruitment is unknown. Sgo1 can also bind to the heterochromatin protein HP1, which itself binds chromatin by recognizing histone H3 trimethylated on Lys-9 (H3K9me3) [11]. Although most HP1 is removed from chromosomes during mitosis, a small population remains at inner centromeres that could recruit Sgo1. However, other studies have found that key H3K9 methyltransferases are not required for HP1 or Sgo1 localization in mitosis [12,13], and that HP1 binds to mitotic centromeres via the chromosomal passenger complex (CPC) in a manner that excludes HP1 binding to Sgo1 [14]. An alternative potential contribution to inner centromere Sgo1 localization is binding to cohesin itself, an interaction that depends on phosphorylation of Sgo1 at Thr-346 by cyclin-dependent kinases (Cdk) [5]. How do these proposed mechanisms act together to control Sgo1 function? Although the dependency of Sgo1 localization on Bub1 activity is largely unquestioned, the reason that centromeric cohesion depends on Bub1 is less clear [15,16]. Bub1 is a mitotic checkpoint protein, and lowering Bub1 levels might lead to
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Figure 1. Model for regulation of Sgo1 localization during mitosis. During prophase, Sgo1 (green) is phosphorylated at Thr-346 and binds to cohesin complexes (red) at the inner centromere between sister chromatids (pale blue). This inner centromeric accumulation of Sgo1 requires Bub1 and binding to H2AT120ph in an as yet undetermined manner (curved arrows). Cohesin on chromosome arms is released through the action of mitotic kinases, but Sgo1 protects inner centromere cohesin. Once the chromosomes become bi-oriented, Sgo1 is dephosphosphorylated at Thr-346 and Sgo1 no longer binds to cohesin. Instead, Sgo1 redistributes towards H2AT120ph (dark blue), in regions approximately coinciding with centromeric chromatin containing CENP-A (yellow). Once the mitotic checkpoint is satisfied, Separase is activated and cleaves the now unprotected cohesin at inner centromeres. In anaphase, H2AT120ph begins to decline, and Sgo1 is eventually released.
cohesion loss because the checkpoint is compromised and anaphase is initiated, rather than because Bub1 and H2AT120ph are required for Sgo1 localization [15]. In their new study, Liu et al. acknowledge that inactivation of Bub1 causes a weaker cohesion phenotype than loss of Sgo1 but argue that centromeric cohesion is flawed when Bub1 is depleted, even when the checkpoint remains active [1]. However, they also find that Sgo1 does not always co-localize with H2AT120ph, particularly on chromosomes that lack microtubule attachments. On such chromosomes, H2AT120ph largely overlaps with CENP-A-containing chromatin at kinetochores whereas Sgo1 is found at inner centromeres (Figure 1). These results are consistent with an additional contribution to Sgo1 localization and function beyond that of the Bub1–H2AT120ph pathway. To determine the relative roles of the Bub1–H2AT120ph and cohesin-dependent pathways, Liu et al. examined separation-of-function mutants of Sgo1. A mutant (K492A) that could not co-immunoprecipitate H2AT120ph, but still bound cohesin, was no longer enriched at centromeres. Instead, it was found on chromosome arms, consistent with the effects of depleting Bub1. In contrast, a mutant (T346A) that could interact with
H2AT120ph but was unable to bind cohesin was found at kinetochores, but was unable to localize to inner centromeres. Therefore, H2AT120ph binding appears important for all centromeric enrichment of Sgo1, while cohesin binding is important specifically for the accumulation of Sgo1 at inner centromeres. Notably, Sgo1-T346A (which cannot bind cohesin) was unable to restore cohesion in Sgo1-depleted cells. In contrast, Sgo1-K492A (which retains cohesin binding) was largely, though not fully, able to support cohesion. The authors propose that these two different binding modes underlie the redistribution of Sgo1 observed during mitosis. When microtubules were depolymerized with nocodazole, the inner centromere localization and phosphorylation of Sgo1 at Thr-346 were increased, and Sgo1 interaction with H2AT120ph was decreased. Furthermore, a phospho-mimicking Sgo1-T346D mutant was partially retained at inner centromeres, even when chromosomes were bi-oriented. Cells expressing this mutant had increased numbers of lagging chromosomes in anaphase, consistent with failure to fully remove cohesin from centromeres. These results led to a model in which Cdk-dependent phosphorylation at Thr-346 in prophase allows Sgo1
to bind and protect cohesin at inner centromeres. Bi-orientation of chromosomes in metaphase leads to dephosphorylation of Thr-346, loss of cohesin binding, and redistribution of Sgo1 toward H2AT120ph at inner kinetochores, where it cannot prevent cleavage of inner centromeric cohesin by Separase (Figure 1). Thus, microtubule attachment imposes an orchestrated change in the phosphorylation and binding partners of Sgo1 to bring about its relocalization and to regulate cohesion. The findings raise a number of questions. The model provides a mechanism for Sgo1 regulation by tension across bi-oriented chromosomes, but is it really tension that triggers Sgo1 relocation, or is stable microtubule attachment to kinetochores sufficient? What makes Cdk-dependent phosphorylation of Sgo1 responsive to attachment status and could kinetochore-bound cyclin B [6,17] play a role? Do these studies imply that HP1 has no role in Sgo1 recruitment? Not necessarily. One possibility is that HP1 is important for Sgo1 localization prior to, but not during, mitosis [13,14]. Alternatively, ongoing work suggests that Sgo1 can be retained at inner centromeres in mitosis by HP1, but that this system is compromised in a wide range of cancer cells (Y. Tanno and Y. Watanabe, personal communication). The possibility that commonly studied cell lines are defective in certain aspects of cohesion regulation could underlie other conflicting observations in the field, including those regarding the role of Bub1 in cohesion regulation. A significant unresolved issue is why inner centromeric localization of Sgo1 depends on the Bub1–H2AT120ph pathway. Recruitment by H2AT120ph might increase the local concentration of Sgo1 and make binding to nearby cohesin (or HP1) more likely. However, Liu et al. find that Sgo1 does not interact detectably with H2AT120ph in nocodazole-treated cells even though, based on results with the K492A mutant, the ability to interact with H2AT120ph is required for Sgo1 to accumulate at inner centromeres in similar conditions [1]. Perhaps transient association with H2AT120ph allows Sgo1 to pick up a binding partner or modification (such as Thr-346 phosphorylation) that is needed to then bind at inner centromeres.
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Bub1 appears to be a major conduit for ‘outside-in’ signals from the kinetochore to the inner centromere [18], so these studies are likely to have implications beyond cohesion. In particular, H2AT120ph generated by Bub1 co-operates with another histone modification, H3T3ph generated by Haspin, to specify the inner centromere localization of the CPC [10,19]. The mechanism of this co-operation, however, is incompletely defined. The new results from Liu et al. imply that Bub1 and H2AT120ph indirectly enhance Sgo1 binding to inner centromeres. Inner centromeric Sgo1 might then provide direct binding sites for the CPC and/or protect cohesin to provide binding sites for Haspin [10], and could therefore help make CPC localization sensitive to kinetochore–microtubule attachments [20]. Further work to fully understand how Bub1 activity enhances the inner centromeric localization of Sgo1 is likely to provide insight into multiple aspects of inner centromere function and chromosome segregation in mitosis. References 1. Liu, H., Jia, L., and Yu, H. (2013). Phospho-H2A and cohesin specify distinct tension-regulated Sgo1 pools at kinetochores and inner centromeres. Curr. Biol. 23, 1927–1933. 2. Nasmyth, K., and Haering, C.H. (2009). Cohesin: its roles and mechanisms. Annu. Rev. Genet. 43, 525–558. 3. Riedel, C.G., Katis, V.L., Katou, Y., Mori, S., Itoh, T., Helmhart, W., Galova, M., Petronczki, M., Gregan, J., Cetin, B., et al.
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(2006). Protein phosphatase 2A protects centromeric sister chromatid cohesion during meiosis I. Nature 441, 53–61. Kitajima, T.S., Sakuno, T., Ishiguro, K., Iemura, S., Natsume, T., Kawashima, S.A., and Watanabe, Y. (2006). Shugoshin collaborates with protein phosphatase 2A to protect cohesin. Nature 441, 46–52. Liu, H., Rankin, S., and Yu, H. (2013). Phosphorylation-enabled binding of SGO1-PP2A to cohesin protects sororin and centromeric cohesion during mitosis. Nat. Cell Biol. 15, 40–49. Lee, J., Kitajima, T.S., Tanno, Y., Yoshida, K., Morita, T., Miyano, T., Miyake, M., and Watanabe, Y. (2008). Unified mode of centromeric protection by shugoshin in mammalian oocytes and somatic cells. Nat. Cell Biol. 10, 42–52. Clift, D., Bizzari, F., and Marston, A.L. (2009). Shugoshin prevents cohesin cleavage by PP2A(Cdc55)-dependent inhibition of separase. Genes Dev. 23, 766–780. McGuinness, B.E., Hirota, T., Kudo, N.R., Peters, J.M., and Nasmyth, K. (2005). Shugoshin prevents dissociation of cohesin from centromeres during mitosis in vertebrate cells. PLoS Biol. 3, e86. Kawashima, S.A., Yamagishi, Y., Honda, T., Ishiguro, K., and Watanabe, Y. (2010). Phosphorylation of H2A by Bub1 prevents chromosomal instability through localizing shugoshin. Science 327, 172–177. Yamagishi, Y., Honda, T., Tanno, Y., and Watanabe, Y. (2010). Two histone marks establish the inner centromere and chromosome bi-orientation. Science 330, 239–243. Yamagishi, Y., Sakuno, T., Shimura, M., and Watanabe, Y. (2008). Heterochromatin links to centromeric protection by recruiting shugoshin. Nature 455, 251–255. Koch, B., Kueng, S., Ruckenbauer, C., Wendt, K.S., and Peters, J.M. (2008). The Suv39h-HP1 histone methylation pathway is dispensable for enrichment and protection of cohesin at centromeres in mammalian cells. Chromosoma 117, 199–210. Perera, D., and Taylor, S.S. (2010). Sgo1 establishes the centromeric cohesion
Evolution: Sperm, Cryptic Choice, and the Origin of Species In two fruit fly species, in vivo observations of competing sperm reveal how differences in sperm size, female behavior and reproductive architecture promote retention of same-species sperm. Sexual selection continues after mating and may play an important role in speciation. Adam K. Chippindale Populations may diverge into separate species when they become physically isolated, each adapting to different environments and genetically drifting apart for long periods of time. But when there isn’t complete physical isolation, the probability of speciation will be greater if there are mechanisms that inhibit gene flow between diverging populations. Differences in
habitat use, the timing of reproduction and mating preferences that favour like breeding with like are factors that may promote speciation. In some species, a female can successfully mate and produce offspring with a male from her own species (a ‘conspecific’ male) or with a male from a closely related species (a ‘heterospecific’ male). If she were to mate with both types of male within a short time period, their sperm would compete for fertilization
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protection mechanism in G2 before subsequent Bub1-dependent recruitment in mitosis. J. Cell Sci. 123, 653–659. Kang, J., Chaudhary, J., Dong, H., Kim, S., Brautigam, C.A., and Yu, H. (2011). Mitotic centromeric targeting of HP1 and its binding to Sgo1 are dispensable for sister-chromatid cohesion in human cells. Mol. Biol. Cell 22, 1181–1190. Perera, D., Tilston, V., Hopwood, J.A., Barchi, M., Boot-Handford, R.P., and Taylor, S.S. (2007). Bub1 maintains centromeric cohesion by activation of the spindle checkpoint. Dev. Cell 13, 566–579. Ricke, R.M., Jeganathan, K.B., Malureanu, L., Harrison, A.M., and van Deursen, J.M. (2012). Bub1 kinase activity drives error correction and mitotic checkpoint control but not tumor suppression. J. Cell Biol. 199, 931–949. Bentley, A.M., Normand, G., Hoyt, J., and King, R.W. (2007). Distinct sequence elements of cyclin B1 promote localization to chromatin, centrosomes, and kinetochores during mitosis. Mol. Biol. Cell 18, 4847–4858. Boyarchuk, Y., Salic, A., Dasso, M., and Arnaoutov, A. (2007). Bub1 is essential for assembly of the functional inner centromere. J. Cell Biol. 176, 919–928. Wang, F., Ulyanova, N.P., van der Waal, M.S., Patnaik, D., Lens, S.M.A., and Higgins, J.M.G. (2011). A positive feedback loop involving Haspin and Aurora B promotes CPC accumulation at centromeres in mitosis. Curr. Biol. 21, 1061–1069. Salimian, K.J., Ballister, E.R., Smoak, E.M., Wood, S., Panchenko, T., Lampson, M.A., and Black, B.E. (2011). Feedback control in sensing chromosome biorientation by the Aurora B kinase. Curr. Biol. 21, 1158–1165.
Division of Rheumatology, Immunology and Allergy, Brigham and Women’s Hospital, Harvard Medical School, Smith Building Room 538A, 1 Jimmy Fund Way, Boston, MA 02115, USA. E-mail: [email protected] http://dx.doi.org/10.1016/j.cub.2013.08.026
opportunities inside her reproductive tract. In sperm competition, the conspecific male tends to hold a fertilization advantage, irrespective of mating order, whereas in sperm competition between two conspecific males, mating order matters. This home court advantage in the interspecific love triangle, called conspecific sperm precedence, suggests a complicated interaction between the two different males’ ejaculates and the female reproductive tract in which they compete. Such postcopulatory sexual selection is among the most cryptic of biological processes known, yet is important because it influences paternity and can promote the evolution of isolation, driving populations towards new species [1]. In this issue of Current Biology, Mollie Manier, Scott
an mTORC1 substrate that negatively regulates insulin signaling. Science 332, 1322–1326. 16. Chung, J., Kuo, C.J., Crabtree, G.R., and Blenis, J. (1992). Rapamycin-FKBP specifically blocks growth-dependent activation of and signaling by the 70 kd S6 protein kinases. Cell 69, 1227–1236. 17. Zhang, Y., and Zheng, X.F. (2012). mTOR-independent 4E-BP1 phosphorylation is associated with cancer resistance to mTOR kinase inhibitors. Cell Cycle 11, 594–603. 18. Ducker, G.S., Atreya, C.E., Simko, J.P., Hom, Y.K., Matli, M.R., Benes, C.H., Hann, B., Nakakura, E.K., Bergsland, E.K., Donner, D.B., et al. (2013). Incomplete inhibition of phosphorylation of 4E-BP1 as a mechanism of primary resistance to ATP-competitive mTOR
inhibitors. Oncogene http://dx.doi.org/10.1038/ onc.2013.92. 19. She, Q.B., Halilovic, E., Ye, Q., Zhen, W., Shirasawa, S., Sasazuki, T., Solit, D.B., and Rosen, N. (2010). 4E-BP1 is a key effector of the oncogenic activation of the AKT and ERK signaling pathways that integrates their function in tumors. Cancer Cell 18, 39–51. 20. Shin, S., Wolgamott, L., Tcherkezian, J., Vallabhapurapu, S., Yu, Y., Roux, P.P., and Yoon, S.O. (2013). Glycogen synthase kinase-3beta positively regulates protein synthesis and cell proliferation through the regulation of translation initiation factor 4E-binding protein 1. Oncogene http:// dx.doi.org/10.1038/onc.2013.113.
Chromosome Segregation: Learning to Let Go To ensure accurate chromosome segregation, cohesion between sister chromatids must be released in a controlled manner during mitosis. A new study reveals how distinct centromere populations of the cohesin protector Sgo1 are regulated by microtubule attachments, cyclin-dependent kinases, and the kinetochore kinase Bub1. Jonathan M.G. Higgins Dividing cells must convey the correct complement of chromosomes to their offspring. Eukaryotes accomplish this by maintaining cohesion between replicated sister chromatids until chromosomes are bi-oriented on the mitotic spindle. Only once this has been accomplished are the attachments between chromatids released, allowing them to be sorted accurately to opposite poles of the dividing cell. Clearly then, although sister chromatids may be inseparable at first, they must learn to let go when the time comes. A report from Liu, Jia and Yu in this issue of Current Biology [1] provides new insight into this process that may have broader implications for our understanding of inner centromere function. Cohesion between sister chromatids is maintained by cohesin complexes, together with regulators such as Sororin [2]. In vertebrate mitosis, cohesin is removed from chromosomes in two steps. In prophase, a mechanism involving phosphorylation of cohesin and Sororin by mitotic kinases removes the bulk of cohesin from chromosome arms (Figure 1). Cohesin at centromeres, however, is protected by Sgo1–PP2A phosphatase complexes that counteract phosphorylation of cohesin
and Sororin [3–5]. To fully separate chromatids at anaphase, the remaining cohesin is cleaved by the protease Separase [2]. This raises the question of how cleavage of centromeric cohesin is limited to anaphase. A simple possibility is that Separase only becomes active at anaphase, and that Sgo1 does not protect cohesin from cleavage in mitosis. However, it has been reported that Sgo1, when inappropriately maintained at inner centromeres, prevents Separase-mediated cohesin cleavage [6]. Also, at least in budding yeast, Sgo1–PP2A complexes may inhibit Separase more directly [7]. Therefore, it is important to understand how the localization and activity of Sgo1 are regulated. During prophase in mammalian cells, Sgo1 is found at inner centromeres (defined here as the area between the chromatin regions that contain centromeric histone CENP-A; Figure 1). As chromosomes become bi-oriented, Sgo1 appears to move outwards, relocating to two regions roughly coinciding with CENP-A-containing chromatin underlying kinetochores [1,6,8]. This movement of Sgo1 away from cohesin complexes located at inner centromeres might render cohesin susceptible to cleavage by Separase, and would provide a way to make removal of cohesin favorable
1Department of Cancer and Cell Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA. 2Institute for Research in Immunology and Cancer (IRIC), Universite´ de Montre´al, Montreal, Quebec H3C 3J7, Canada. 3Department of Pathology and Cell Biology, Faculty of Medicine, Universite´ de Montre´al, Montreal, Quebec, H3C 3J7, Canada. E-mail: [email protected], philippe. [email protected]
http://dx.doi.org/10.1016/j.cub.2013.08.030
only when chromosomes are correctly bi-oriented and microtubules exert tension across sister kinetochores [6]. How this relocation of Sgo1 is controlled, however, has been unknown. A number of ways to recruit Sgo1 to centromeres have been reported, but the relative contributions of these pathways are debated. It is widely accepted that Sgo1 is brought to centromeres when histone H2A is phosphorylated at Thr-120 (H2AT120ph) by the kinetochore kinase Bub1 [9,10], though the structural basis for this recruitment is unknown. Sgo1 can also bind to the heterochromatin protein HP1, which itself binds chromatin by recognizing histone H3 trimethylated on Lys-9 (H3K9me3) [11]. Although most HP1 is removed from chromosomes during mitosis, a small population remains at inner centromeres that could recruit Sgo1. However, other studies have found that key H3K9 methyltransferases are not required for HP1 or Sgo1 localization in mitosis [12,13], and that HP1 binds to mitotic centromeres via the chromosomal passenger complex (CPC) in a manner that excludes HP1 binding to Sgo1 [14]. An alternative potential contribution to inner centromere Sgo1 localization is binding to cohesin itself, an interaction that depends on phosphorylation of Sgo1 at Thr-346 by cyclin-dependent kinases (Cdk) [5]. How do these proposed mechanisms act together to control Sgo1 function? Although the dependency of Sgo1 localization on Bub1 activity is largely unquestioned, the reason that centromeric cohesion depends on Bub1 is less clear [15,16]. Bub1 is a mitotic checkpoint protein, and lowering Bub1 levels might lead to
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Figure 1. Model for regulation of Sgo1 localization during mitosis. During prophase, Sgo1 (green) is phosphorylated at Thr-346 and binds to cohesin complexes (red) at the inner centromere between sister chromatids (pale blue). This inner centromeric accumulation of Sgo1 requires Bub1 and binding to H2AT120ph in an as yet undetermined manner (curved arrows). Cohesin on chromosome arms is released through the action of mitotic kinases, but Sgo1 protects inner centromere cohesin. Once the chromosomes become bi-oriented, Sgo1 is dephosphosphorylated at Thr-346 and Sgo1 no longer binds to cohesin. Instead, Sgo1 redistributes towards H2AT120ph (dark blue), in regions approximately coinciding with centromeric chromatin containing CENP-A (yellow). Once the mitotic checkpoint is satisfied, Separase is activated and cleaves the now unprotected cohesin at inner centromeres. In anaphase, H2AT120ph begins to decline, and Sgo1 is eventually released.
cohesion loss because the checkpoint is compromised and anaphase is initiated, rather than because Bub1 and H2AT120ph are required for Sgo1 localization [15]. In their new study, Liu et al. acknowledge that inactivation of Bub1 causes a weaker cohesion phenotype than loss of Sgo1 but argue that centromeric cohesion is flawed when Bub1 is depleted, even when the checkpoint remains active [1]. However, they also find that Sgo1 does not always co-localize with H2AT120ph, particularly on chromosomes that lack microtubule attachments. On such chromosomes, H2AT120ph largely overlaps with CENP-A-containing chromatin at kinetochores whereas Sgo1 is found at inner centromeres (Figure 1). These results are consistent with an additional contribution to Sgo1 localization and function beyond that of the Bub1–H2AT120ph pathway. To determine the relative roles of the Bub1–H2AT120ph and cohesin-dependent pathways, Liu et al. examined separation-of-function mutants of Sgo1. A mutant (K492A) that could not co-immunoprecipitate H2AT120ph, but still bound cohesin, was no longer enriched at centromeres. Instead, it was found on chromosome arms, consistent with the effects of depleting Bub1. In contrast, a mutant (T346A) that could interact with
H2AT120ph but was unable to bind cohesin was found at kinetochores, but was unable to localize to inner centromeres. Therefore, H2AT120ph binding appears important for all centromeric enrichment of Sgo1, while cohesin binding is important specifically for the accumulation of Sgo1 at inner centromeres. Notably, Sgo1-T346A (which cannot bind cohesin) was unable to restore cohesion in Sgo1-depleted cells. In contrast, Sgo1-K492A (which retains cohesin binding) was largely, though not fully, able to support cohesion. The authors propose that these two different binding modes underlie the redistribution of Sgo1 observed during mitosis. When microtubules were depolymerized with nocodazole, the inner centromere localization and phosphorylation of Sgo1 at Thr-346 were increased, and Sgo1 interaction with H2AT120ph was decreased. Furthermore, a phospho-mimicking Sgo1-T346D mutant was partially retained at inner centromeres, even when chromosomes were bi-oriented. Cells expressing this mutant had increased numbers of lagging chromosomes in anaphase, consistent with failure to fully remove cohesin from centromeres. These results led to a model in which Cdk-dependent phosphorylation at Thr-346 in prophase allows Sgo1
to bind and protect cohesin at inner centromeres. Bi-orientation of chromosomes in metaphase leads to dephosphorylation of Thr-346, loss of cohesin binding, and redistribution of Sgo1 toward H2AT120ph at inner kinetochores, where it cannot prevent cleavage of inner centromeric cohesin by Separase (Figure 1). Thus, microtubule attachment imposes an orchestrated change in the phosphorylation and binding partners of Sgo1 to bring about its relocalization and to regulate cohesion. The findings raise a number of questions. The model provides a mechanism for Sgo1 regulation by tension across bi-oriented chromosomes, but is it really tension that triggers Sgo1 relocation, or is stable microtubule attachment to kinetochores sufficient? What makes Cdk-dependent phosphorylation of Sgo1 responsive to attachment status and could kinetochore-bound cyclin B [6,17] play a role? Do these studies imply that HP1 has no role in Sgo1 recruitment? Not necessarily. One possibility is that HP1 is important for Sgo1 localization prior to, but not during, mitosis [13,14]. Alternatively, ongoing work suggests that Sgo1 can be retained at inner centromeres in mitosis by HP1, but that this system is compromised in a wide range of cancer cells (Y. Tanno and Y. Watanabe, personal communication). The possibility that commonly studied cell lines are defective in certain aspects of cohesion regulation could underlie other conflicting observations in the field, including those regarding the role of Bub1 in cohesion regulation. A significant unresolved issue is why inner centromeric localization of Sgo1 depends on the Bub1–H2AT120ph pathway. Recruitment by H2AT120ph might increase the local concentration of Sgo1 and make binding to nearby cohesin (or HP1) more likely. However, Liu et al. find that Sgo1 does not interact detectably with H2AT120ph in nocodazole-treated cells even though, based on results with the K492A mutant, the ability to interact with H2AT120ph is required for Sgo1 to accumulate at inner centromeres in similar conditions [1]. Perhaps transient association with H2AT120ph allows Sgo1 to pick up a binding partner or modification (such as Thr-346 phosphorylation) that is needed to then bind at inner centromeres.
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Bub1 appears to be a major conduit for ‘outside-in’ signals from the kinetochore to the inner centromere [18], so these studies are likely to have implications beyond cohesion. In particular, H2AT120ph generated by Bub1 co-operates with another histone modification, H3T3ph generated by Haspin, to specify the inner centromere localization of the CPC [10,19]. The mechanism of this co-operation, however, is incompletely defined. The new results from Liu et al. imply that Bub1 and H2AT120ph indirectly enhance Sgo1 binding to inner centromeres. Inner centromeric Sgo1 might then provide direct binding sites for the CPC and/or protect cohesin to provide binding sites for Haspin [10], and could therefore help make CPC localization sensitive to kinetochore–microtubule attachments [20]. Further work to fully understand how Bub1 activity enhances the inner centromeric localization of Sgo1 is likely to provide insight into multiple aspects of inner centromere function and chromosome segregation in mitosis. References 1. Liu, H., Jia, L., and Yu, H. (2013). Phospho-H2A and cohesin specify distinct tension-regulated Sgo1 pools at kinetochores and inner centromeres. Curr. Biol. 23, 1927–1933. 2. Nasmyth, K., and Haering, C.H. (2009). Cohesin: its roles and mechanisms. Annu. Rev. Genet. 43, 525–558. 3. Riedel, C.G., Katis, V.L., Katou, Y., Mori, S., Itoh, T., Helmhart, W., Galova, M., Petronczki, M., Gregan, J., Cetin, B., et al.
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Evolution: Sperm, Cryptic Choice, and the Origin of Species In two fruit fly species, in vivo observations of competing sperm reveal how differences in sperm size, female behavior and reproductive architecture promote retention of same-species sperm. Sexual selection continues after mating and may play an important role in speciation. Adam K. Chippindale Populations may diverge into separate species when they become physically isolated, each adapting to different environments and genetically drifting apart for long periods of time. But when there isn’t complete physical isolation, the probability of speciation will be greater if there are mechanisms that inhibit gene flow between diverging populations. Differences in
habitat use, the timing of reproduction and mating preferences that favour like breeding with like are factors that may promote speciation. In some species, a female can successfully mate and produce offspring with a male from her own species (a ‘conspecific’ male) or with a male from a closely related species (a ‘heterospecific’ male). If she were to mate with both types of male within a short time period, their sperm would compete for fertilization
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protection mechanism in G2 before subsequent Bub1-dependent recruitment in mitosis. J. Cell Sci. 123, 653–659. Kang, J., Chaudhary, J., Dong, H., Kim, S., Brautigam, C.A., and Yu, H. (2011). Mitotic centromeric targeting of HP1 and its binding to Sgo1 are dispensable for sister-chromatid cohesion in human cells. Mol. Biol. Cell 22, 1181–1190. Perera, D., Tilston, V., Hopwood, J.A., Barchi, M., Boot-Handford, R.P., and Taylor, S.S. (2007). Bub1 maintains centromeric cohesion by activation of the spindle checkpoint. Dev. Cell 13, 566–579. Ricke, R.M., Jeganathan, K.B., Malureanu, L., Harrison, A.M., and van Deursen, J.M. (2012). Bub1 kinase activity drives error correction and mitotic checkpoint control but not tumor suppression. J. Cell Biol. 199, 931–949. Bentley, A.M., Normand, G., Hoyt, J., and King, R.W. (2007). Distinct sequence elements of cyclin B1 promote localization to chromatin, centrosomes, and kinetochores during mitosis. Mol. Biol. Cell 18, 4847–4858. Boyarchuk, Y., Salic, A., Dasso, M., and Arnaoutov, A. (2007). Bub1 is essential for assembly of the functional inner centromere. J. Cell Biol. 176, 919–928. Wang, F., Ulyanova, N.P., van der Waal, M.S., Patnaik, D., Lens, S.M.A., and Higgins, J.M.G. (2011). A positive feedback loop involving Haspin and Aurora B promotes CPC accumulation at centromeres in mitosis. Curr. Biol. 21, 1061–1069. Salimian, K.J., Ballister, E.R., Smoak, E.M., Wood, S., Panchenko, T., Lampson, M.A., and Black, B.E. (2011). Feedback control in sensing chromosome biorientation by the Aurora B kinase. Curr. Biol. 21, 1158–1165.
Division of Rheumatology, Immunology and Allergy, Brigham and Women’s Hospital, Harvard Medical School, Smith Building Room 538A, 1 Jimmy Fund Way, Boston, MA 02115, USA. E-mail: [email protected] http://dx.doi.org/10.1016/j.cub.2013.08.026
opportunities inside her reproductive tract. In sperm competition, the conspecific male tends to hold a fertilization advantage, irrespective of mating order, whereas in sperm competition between two conspecific males, mating order matters. This home court advantage in the interspecific love triangle, called conspecific sperm precedence, suggests a complicated interaction between the two different males’ ejaculates and the female reproductive tract in which they compete. Such postcopulatory sexual selection is among the most cryptic of biological processes known, yet is important because it influences paternity and can promote the evolution of isolation, driving populations towards new species [1]. In this issue of Current Biology, Mollie Manier, Scott