During meiosis, sequential release of sister chromatid cohesion (SSC) during two successive nuclear divisions allows the production of haploid gametes from diploid progenitor cells. Release of SSC along chromosome arms allows first a reductional segregation of homologs, and, subsequently, release of centromeric cohesion at anaphase II allows the segregation of chromatids. The Shugoshin (SGO) protein family plays a major role in the protection of centromeric cohesion in Drosophila and yeast. A maize mutant was isolated that displays premature loss of centromeric cohesion at anaphase I. This phenotype is due to the absence of ZmSGO1 protein, a maize shugoshin homolog. ZmSGO1 is localized to the centromeres. The ZmSGO1 protein is not found on mitotic chromosomes and has no obvious mitotic function. On the basis of these results, it is proposed that ZmSGO1 specifically maintains centromeric cohesion during meiosis I and therefore it is suggested that SGO1 core functions during meiosis are conserved across kingdoms and in large-genome species. However, in contrast to other Shugoshins, an early and REC8-dependent recruitment of ZmSGO1 is observed in maize, suggesting that control of SGO1 recruitment to chromosomes is different in plants than in other model organisms (Hamant, 2005).
Meiosis comprises a pair of specialized nuclear divisions that produce haploid germ cells. To accomplish this, sister chromatids must segregate together during the first meiotic division (meiosis I), which requires that sister chromatid cohesion persists at centromeres. The factors that protect centromeric cohesion during meiosis I have remained elusive. This study identifies Sgo1 (shugoshin), a protector of the centromeric cohesin Rec8 in fission yeast. A homologue of Sgo1 was also identified in budding yeast. Evidence is provided that shugoshin is widely conserved among eukaryotes. Moreover, Sgo2, a paralogue of shugoshin, was identified in fission yeast that is required for faithful mitotic chromosome segregation. Localization of Sgo1 and Sgo2 at centromeres requires the kinase Bub1 (see Drosophila Bub1), identifying shugoshin as a crucial target for the kinetochore function of Bub1. These findings provide insights into the evolution of meiosis and kinetochore regulation during mitosis and meiosis (Kitajima, 2004).
The halving of chromosome number that occurs during meiosis depends on three factors: (1) homologs must pair and recombine; (2) sister centromeres must attach to microtubules that emanate from the same spindle pole, which ensures that homologous maternal and paternal pairs can be pulled in opposite directions (called homolog biorientation); (3) cohesion between sister centromeres must persist after the first meiotic division to enable their biorientation at the second. A screen performed in fission yeast to identify meiotic chromosome missegregation mutants has identified a conserved protein called Sgo1 that is required to maintain sister chromatid cohesion after the first meiotic division. An orthologous protein is described in the budding yeast S. cerevisiae (Sc), which has not only meiotic but also mitotic chromosome segregation functions. Deletion of Sc SGO1 not only causes frequent homolog nondisjunction at meiosis I but also random segregation of sister centromeres at meiosis II. Meiotic cohesion fails to persist at centromeres after the first meiotic division, and sister centromeres frequently separate precociously. Sgo1 is a kinetochore-associated protein whose abundance declines at anaphase I but, nevertheless, persists on chromatin until anaphase II. The finding that Sgo1 is localized to the centromere at the time of the first division suggests that it may play a direct role in preventing the removal of centromeric cohesin. The similarity in sequence composition, chromosomal location, and mutant phenotypes of sgo1 mutants in two distant yeasts with that of MEI-S332 in Drosophila suggests that these proteins define an orthologous family conserved in most eukaryotic lineages (Katis, 2004).
Meiosis produces haploid gametes from diploid progenitor cells. This reduction is achieved by two successive nuclear divisions after one round of DNA replication. Correct chromosome segregation during the first division depends on sister kinetochores being oriented toward the same spindle pole while homologous kinetochores must face opposite poles. Segregation during the second division depends on retention of sister chromatid cohesion between centromeres until the onset of anaphase II, which in Drosophila melanogaster depends on a protein called Mei-S332 that binds to centromeres. Two homologs of Mei-S332 have been identified in fission yeast using a knockout screen. Together with their fly ortholog they define a protein family conserved from fungi to mammals. The two identified genes, sgo1 and sgo2, are required for retention of sister centromere cohesion between meiotic divisions and kinetochore orientation during meiosis I, respectively. The amount of meiotic cohesin's Rec8 subunit retained at centromeres after meiosis I is reduced in Deltasgo1, but not in Deltasgo2, cells, and Sgo1 appears to regulate cleavage of Rec8 by separase. Both Sgo1 and Sgo2 proteins localize to centromere regions. The abundance of Sgo1 protein normally declines after the first meiotic division, but extending its expression by altering its 3'UTR sequences does not greatly affect meiosis II. Its mere presence within the cell might therefore be insufficient to protect centromeric cohesion. In conclusion, a conserved protein family based on Mei-S332 has been identified. The two fission yeast homologs are implicated in meiosis I kinetochore orientation and retention of centromeric sister chromatid cohesion until meiosis II (Rabitsch, 2004).
During meiosis, two chromosome segregation phases follow a single round of DNA replication. Factors that are required to establish this specialized cell cycle have been identified by examining meiotic chromosome segregation in a collection of yeast strains lacking all nonessential genes. This analysis revealed Sgo1, Chl4, and Iml3 to be important for retaining centromeric cohesin until the onset of anaphase II. Consistent with this role, Sgo1 localizes to centromeric regions but dissociates at the onset of anaphase II. The screen described in this study provides a comprehensive analysis of the genes required for the meiotic cell cycle and identifies three factors important for the stepwise loss of sister chromatid cohesion (Marston, 1994).
Homologue segregation during the first meiotic division requires the proper spatial regulation of sister chromatid cohesion and its dissolution along chromosome arms, but its protection at centromeric regions. This protection requires the conserved MEI-S332/Sgo1 proteins that localize to centromeric regions and also recruit the PP2A phosphatase by binding its regulatory subunit, Rts1. Centromeric Rts1/PP2A then locally prevents cohesion dissolution possibly by dephosphorylating the protein complex cohesin. This study shows that Aurora B kinase in Saccharomyces cerevisiae (Ipl1) is also essential for the protection of meiotic centromeric cohesion. Coupled with a previous study in Drosophila, this meiotic function of Aurora B kinase appears to be conserved among eukaryotes. Furthermore, Sgo1 recruits Ipl1 to centromeric regions. In the absence of Ipl1, Rts1 can initially bind to centromeric regions but disappears from these regions after anaphase I onset. It is suggested that centromeric Ipl1 ensures the continued centromeric presence of active Rts1/PP2A, which in turn locally protects cohesin and cohesion (Yu, 2007).
Cohesion between sister chromatids is essential for their bi-orientation on mitotic spindles. It is mediated by a multisubunit complex called cohesin. In yeast, proteolytic cleavage of cohesin's alpha kleisin subunit at the onset of anaphase removes cohesin from both centromeres and chromosome arms and thus triggers sister chromatid separation. In animal cells, most cohesin is removed from chromosome arms during prophase via a separase-independent pathway involving phosphorylation of its Scc3-SA1/2 subunits. Cohesin at centromeres is refractory to this process and persists until metaphase, whereupon its alpha kleisin subunit is cleaved by separase, which is thought to trigger anaphase. What protects centromeric cohesin from the prophase pathway? Potential candidates are proteins, known as shugoshins, that are homologous to Drosophila Mei-S332 and yeast Sgo1 proteins, which prevent removal of meiotic cohesin complexes from centromeres at the first meiotic division. A vertebrate shugoshin-like protein associates with centromeres during prophase and disappears at the onset of anaphase. Its depletion by RNA interference causes HeLa cells to arrest in mitosis. Most chromosomes bi-orient on a metaphase plate, but precocious loss of centromeric cohesin from chromosomes is accompanied by loss of all sister chromatid cohesion, the departure of individual chromatids from the metaphase plate, and a permanent cell cycle arrest, presumably due to activation of the spindle checkpoint. Remarkably, expression of a version of Scc3-SA2 whose mitotic phosphorylation sites have been mutated to alanine alleviates the precocious loss of sister chromatid cohesion and the mitotic arrest of cells lacking shugoshin. These data suggest that shugoshin, and by inference its Drosophila homolog Mei-S332, prevents phosphorylation of cohesin's Scc3-SA2 subunit at centromeres during mitosis. This ensures that cohesin persists at centromeres until activation of separase causes cleavage of its alpha kleisin subunit. Centromeric cohesion is one of the hallmarks of mitotic chromosomes. These results imply that it is not an intrinsically stable property, because it can easily be destroyed by mitotic kinases, which are kept in check by shugoshin (Marston, 2004).
Drosophila MEI-S332 and fungal Sgo1 genes are essential for sister centromere cohesion in meiosis I. The related vertebrate Sgo localizes to kinetochores and is required to prevent premature sister centromere separation in mitosis, thus providing an explanation for the differential cohesion observed between the arms and the centromeres of mitotic sister chromatids. Sgo is degraded by the anaphase-promoting complex, allowing the separation of sister centromeres in anaphase. Intriguingly, Sgo interacts strongly with microtubules in vitro and it regulates kinetochore microtubule stability in vivo, consistent with a direct microtubule interaction. Sgo is thus critical for mitotic progression and chromosome segregation and provides an unexpected link between sister centromere cohesion and microtubule interactions at kinetochores (Salic, 2004).
Sister chromatids in mammalian cells remain attached mostly at their centromeres at metaphase because of the loss of cohesion along chromosome arms in prophase. Bub1 retains centromeric cohesion in mitosis of human cells. Depletion of Bub1 or Shugoshin (Sgo1) in HeLa cells by RNA interference causes massive missegregation of sister chromatids that originates at centromeres. Surprisingly, loss of chromatid cohesion in Bub1 and Sgo1 RNA-interference cells does not appear to require the full activation of separase but, instead, triggers a mitotic arrest that depends on Mad2 and Aurora B. Bub1 maintains the steady-state levels and centromeric localization of Sgo1. Therefore, Bub1 protects centromeric cohesion through Shugoshin in mitosis (Tang, 2004).
Shugoshin (Sgo) proteins constitute a conserved protein family defined as centromeric protectors of Rec8-containing cohesin complexes in meiosis. In vertebrate mitosis, Scc1/Rad21-containing cohesin complexes (see Drosophila Rad21) are also protected at centromeres because arm cohesin, but not centromeric cohesin, is largely dissociated in prophase and prometaphase. The dissociation process is dependent on the activity of polo-like kinase (Plk1) and partly dependent on Aurora B. Recently, it has been demonstrated that vertebrate shugoshin is required for preserving centromeric cohesion during mitosis; however, whether human shugoshin protects cohesin itself was not addressed. The persistence of human Scc1 at centromeres in mitosis is indeed dependent on human Sgo1. In fission yeast, Sgo localization depends on Bub1, a conserved spindle checkpoint protein, which is enigmatically also required for chromosome congression during prometaphase in vertebrate cells. Human Sgo1 fails to localize at centromeres in Bub1-repressed cells, and centromeric cohesion is significantly loosened. Remarkably, in these cells, Sgo1 relocates to chromosomes all along their length and provokes ectopic protection from dissociation of Scc1 on chromosome arms. These results reveal a hitherto concealed role for human Bub1 in defining the persistent cohesion site of mitotic chromosomes (Kitajima, 2005).
Fission yeast shugoshin Sgo1 is meiosis specific and cooperates with protein phosphatase 2A to protect centromeric cohesin at meiosis I. The other shugoshin-like protein Sgo2, which requires the heterochromatin protein Swi6/HP1 for full viability, plays a crucial role for proper chromosome segregation at both mitosis and meiosis; however, the underlying mechanisms are totally elusive. This study demonstrates that, unlike Sgo1, Sgo2 is dispensable for centromeric protection of cohesin. Instead, Sgo2 interacts with Bir1/Survivin and promotes Aurora kinase complex localization to the pericentromeric region, to correct erroneous attachment of kinetochores and thereby enable tension-generating attachment. Forced localization of Bir1 to centromeres partly restored the defects of sgo2Delta. This newly identified interaction of shugoshin with Survivin is conserved between mitosis and meiosis and presumably across eukaryotes. It is proposed that ensuring bipolar attachment of kinetochores is the primary role of shugoshin and the role of cohesion protection might have codeveloped to facilitate this process (Kawashima, 2007).
This study demonstrates that human shugoshin hSgo1 associates with Survivin and Aurora and requires these components for its centromeric localization. Together with the recent finding in Drosophila that the Aurora kinase complex is required for centromeric localization of Sgo/Mei-S332 (Resnick, 2006), these studies suggest that the linkage between shugoshin and Aurora kinase complex is conserved among eukaryotes. Studies in human cells present the strongest data to date indicating the existence of a complex including shugoshin and Survivin in vivo; hSgo1 could coprecipitate with Survivin better than Aurora in extracts prepared from chromatin fraction. This result fits with the immunoprecipitation using a cross-linker in fission yeast and with genetic results indicating that Sgo2 closely interacts with Bir1/Survivin for the centromeric localization. Although the linkage between shugoshin and the Aurora kinase complex is conserved across species, the precise manner of interaction has apparently diverged. The centromeric localization of Drosophila Mei-S332 reportedly requires phosphorylation by Aurora (Resnick, 2006); however, fission yeast Sgo2 does not require it, albeit Sgo2, like Mei-S332, is a good substrate of Ark1 in vitro. Whereas fission yeast shugoshin (Sgo2) is required for the localization and function of Aurora kinase complex at centromeres, Drosophila Mei-S332 as well as human Sgo1 is not required for the localization of the Aurora kinase complex (Resnick, 2006), albeit the centromeric function of the Aurora kinase complex might nevertheless be regulated by Mei-S332 (Kawashima, 2007).
The sole shugoshin protein in budding yeast seems to play dual roles in protecting centromeric cohesin at meiosis I (but not at mitosis) as well as in establishing tension-generating attachment at mitosis. Drosophila SGO/MEI-S332 mutants show nondisjunction of homologs at meiosis I and a reduced ratio of meta/anaphase (but only slight or little defect in cohesion) in mitosis. Therefore, it is suggested that Mei-S332, the sole shugoshin of Drosophila, is also required for establishing tension-generating attachment, like fission yeast Sgo2. The localization of the Aurora kinase complex does not depend on Mei-S332; however, it is tenable that the activation of centromeric Aurora kinase complex may somewhat depend on Mei-S332 since they physically interact in vitro (Resnick, 2006). Similarly, fission yeast Sgo2 might play an additional role in activating centromeric Aurora rather than merely promoting its localization. Given that hSgo1 associates with Survivin (and Aurora) in HeLa cells, a similar functional link is conceivable also in human cells (Kawashima, 2007).
Studies in fission yeast enabled definition of two distinct shugoshin functions or pathways that are carried out by two diverged shugoshins, Sgo1 and Sgo2; the former interacts with PP2A to protect cohesin, but the latter interacts with the Aurora kinase complex to facilitate centromeric Aurora function. It is speculated that the ancestral shugoshin molecule played dual roles at kinetochores like in budding yeast or Drosophila; fission yeast shugoshin might have divided the labor to Sgo1 and Sgo2. Thus, these findings of a functional link between Sgo2 and the Aurora kinase complex open a new view that shugoshin in general may play a role in facilitating Aurora function at centromeres, thereby ensuring tension-generating kinetochore microtubule attachment. At the centromere, microtubule attachment is ensured by tension across centromeres, which is generated depending on the cohesion between sister chromatids. Therefore, cohesion and tension are two sides of a 'coin' ensuring bipolar attachment of kinetochores. It is suggested that the original role of shugoshin was to guarantee bipolar attachment rather than to protect cohesin, because fission yeast and presumably budding yeast, two primitive eukaryotes, exhibit this role only during mitosis. The protection role, once acquired, might facilitate the generation of tension by counteracting the spindle force, improving the fidelity of chromosome segregation. This function might have been modified to evolve meiosis, in which the requirement for centromeric protection is more essential and therefore has been preserved in all eukaryotes. Whatever the validity of this view, the finding of how Sgo2 acts will contribute to understand the fundamental regulation of eukaryotic chromosome segregation (Kawashima, 2007).
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