During mitosis, chromosomes segregate with exquisite fidelity. This is achieved by the attachment of paired sister chromatids to the bipolar spindle and by the presence of regulatory mechanisms, called checkpoints, that inhibit mitosis in the presence of DNA or spindle damage. The loss of these regulatory mechanisms, as occurs in certain types of cancer, leads to chromosome missegregation that in turn results in gross cellular defects and cell death. The molecular mechanism that enables mitotic arrest is still unknown. Recently, a gene encoding for a putative inhibitor of mitosis, called PDS1, has been identified in the yeast Saccharomyces cerevisiae. In the absence of PDS1, sister chromatids separate prematurely, cells fail to exhibit a DNA damage checkpoint arrest, and they have a spindle elongation defect. Pds1p appears to be a substrate of the anaphase promoting complex (APC), which is involved in the degradation of type-B cyclins. These results suggest that Pds1p in an inhibitor of mitosis and that its degradation allows mitosis to proceed. The role of Pds1p in the regulation of mitosis will be studied by examining the expression pattern and cellular localization of Pds1p during the cell cycle, by studying the effect of a non-degradable Pds1p derivative on the ability to execute mitosis, and by determining the role of APC and other degradation-associated factors in Pds1p degradation. Proteins that potentially interact with Pds1p will be identified by screening for extragenic suppressors of conditional pds1 alleles. The possibility that the involvement of Pds1p in spindle function and mitotic regulation stems from two different functions will be examined by attempting to generate pds1 alleles specifically defective in either function. The results of these experiment will shed light not only on Pds1p function but also on the molecular mechanisms of sister chromatid cohesion, DNA damage checkpoint and spindle elongation.
