Maltose fermentation in Saccharomyces is initiated by its active transport into the cell by maltose permease. The level of activity of maltose permease is under dual regulation. Firstly, transcription of the gene encoding maltose permease is induced by maltose and repressed by glucose. Secondly, maltose permease is glucose inactivated, that is, the ability to transport maltose is rapidly lost following the addition of glucose to a fermenting culture. We propose to determine the mechanism of this post-translational regulation of maltose permease. MA161 encodes maltose permease in Saccharomyces. We have inserted an epitope-tag at the N-terminal end of the Ma161 protein to enable us to follow the fate of the protein during glucose-induced inactivation by using techniques such as immunoprecipitation, immunofluorescence microscopy and Western analysis. The projects described in this proposal explore the possibility that maltose permease is modified by phosphorylation and/or proteolytically degraded during inactivation. We will use the tools of Saccharomyces genetics and the expanding number of mutant strains with alterations affecting the secretory, signal transduction and glucose-response pathways to determine the gene products utilized for the inactivation process. We will isolate glucose inactivation resistant mutations in MA161 and define new Saccharomyces genes encoding products involved in glucose-induced inactivation.
