Molecular Dynamics of Protein Phosphatase 2A in Yeast

Syracuse University, Syracuse NY

Investigators

Abstract

ABSTRACT The project is to study the dynamics of the changes in cellular location of the major serine/threonine protein phosphatase, PP2A, in Saccharomyces cerevisiae. From studies mainly in higher organisms, it appears that this heterotrimeric enzyme, shown to be involved in the regulation of DNA replication, cell cycle control, RNA transcription, as well as signal transduction pathways, achieves specificity through the action of its so-called "regulatory" subunits. One proposed activity of these regulatory subunits is to facilitate the targeting of different forms of the PP2A trimer to specific cellular locations in order to achieve enzymatic specificity. As higher organisms express a large number of different regulatory subunits, S. cerevisisae is particularly well suited to testing the notion of selective targeting by regulatory subunits because its array of regulatory subunits is far smaller. In addition, all the genes encoding PP2A subunits have been cloned and genes expressing either GFP, YFP, or CFP-tagged forms of each protein subunit have been created. Using fluorescence microscopy, the cellular locations of one or more PP2A subunits throughout the cell cycle or growth phase will be followed in real time. Preliminary data generated in the laboratory has identified certain cellular locations, such as the spindle pole body, the emerging bud tip, the bud neck, and the periphery of the vacuole, as sites at which certain PP2A subunits transiently accumulate. Using genetic and microscopic tests, other putative proteins that co-localize with PP2A will be identified. Furthermore, using strains in which one or more PP2A subunit genes are either mutated or inactivated completely, the effects of such mutations on the abilities of the other PP2A subunits to be correctly localized will be assessed. These studies should directly test the notion of whether and how PP2A regulatory subunits control the specific localization of functioning PP2A enzymes.