A critical event during the eukaryotic cell cycle is the duplication of the centrosome to create the two spindle poles necessary for proper spindle formation and chromosome segregation. If centrosome duplication is not completed in time for chromosome segregation, aberrant spindles are formed causing chromosome nondisjunction and genomic instability. These are general features of cancer cells, therefore an important question is how the centrosome is duplicated faithfully in every cell cycle to ensure genomic stability. The functional equivalent of the mammalian centrosome is the spindle pole body (SPB) in the budding yeast Saccharomyces cerevisiae. The experiments described in this proposal are aimed at elucidating the role of MPS2 and its bypass suppressors in SPB duplication, and possibly nuclear pore complex (NPC) function/assembly. The MPS2 gene product (Mps2p) has been localized to both the SPB and the NPC, thus nuclear transport and NPC assembly assays will be performed to determine if NPC function/assembly is normal in mps2 mutants. The SPB and NPC-specific functions will be separated by constructing new mutant alleles of MPS2. Yeast harboring a null allele of MPS2 are marginally alive; a bypass suppressor screen with a yeast genomic library in a high copy number vector will be utilized to identify yeast genes which keep the MPS2 null marginally alive. High copy suppressors of the MPS2 null mutant will be characterized extensively at the genetic and phenotypic levels to determine if they perform a specific role in SPB duplication and/or NPC function/assembly. Ultimately, the knowledge gained studying the yeast SPB will yield insights into how the centrosome is duplicated and assembled to ensure proper chromosome segregation and genetic stability.
