Regulation and Function of the Drosophila Shugoshin, MEI-S332. Belinda Pinto, Cristina Nogueira, Terry Orr-Weaver. Whitehead Inst, Cambridge, MA.

   Accurate chromosome segregation requires the step-wise release of chromatid cohesion from the arms and centromeres. This pattern of release is facilitated by the Shugoshin family of proteins that protect centromeric cohesion. To execute this protective role, Shugoshins interact with the B associated form of the PP2A phosphatase (PP2A-B). The role of PP2A-B during metazoan mitosis has been elucidated, but the role during metazoan meiosis is unclear. The function of Shugoshins in protecting centromeric cohesion is linked to centromeric localization from prometaphase to the metaphase-anaphase transition. A few proteins that regulate Shugoshin localization have been identified, but other as yet unidentified factors are likely to regulate the spatial and temporal localization of these proteins. Here, we examined the role and regulation of Shugoshins in centromeric cohesion through studies of the Drosophila homolog, MEI-S332. First, genetic interaction studies indicate that two B phosphatase subunits, Wdb and dPP2A-B, make redundant contributions to the role of MEI-S332 in centromeric cohesion. Second, we developed a model using Drosophila mitotic cells to identify factors that regulate the centromeric localization of MEI-S332. We performed a high-throughput RNAi screen in Drosophila cell culture and identified a number of candidates that affect MEI-S332 localization at metaphase and anaphase. Validation of one these candidates uncovered a role for the proteasome in delocalizing MEI-S332 at anaphase. Studies to investigate the mechanism by which the proteasome regulates MEI-S332 localization are underway. Taken together, data from our studies of MEI-S332 will be valuable in understanding the mechanism of action of Shugoshins during meiotic and mitotic chromosome segregation.