Proteins that negatively regulate the transition from stationary phase to the cell division cycle in the yeast Saccharomyces cerevisiae

Our lab has discovered a novel mutation in the GCS1 gene of yeast that prevents the developmental transition from stationary phase to the cell division cycle. The Gcs1 protein is a highly conserved GAP ( GTPase-Activating Protein) for both Arf and Arl1 proteins involved in vesicular transport. The deletion of the GCS1 gene ( gcs1Delta) prevents mutant cells from completing the developmental transition from stationary phase to the cell division cycle at the restrictive temperature of 15°C (termed 'reentry defect'). To further characterize the roles of Gcs1 in the process of vesicular transport and in the regulation of cell proliferation from a stationary-phase state, I have screened for genes encoding negative regulators of the gcs1Delta cold-sensitive reentry defect using a robotic colony arrayer and have identified 32 novel negative regulators. Interestingly, five of the identified negative regulators are known to affect a single 'Arl pathway' implicated in a transport-vesicle tethering process which assists in the targeting and binding of transport vesicles to specific membranes within the cell. In this pathway, Imh1 (a vesicle-tethering factor) is recruited to the cytosolic membrane of the trans-Golgi by the activated (GTP-bound) form of Arl1 in a GRIP domain-dependent manner. Imh1 is a known dosage suppressor (or positive regulator) of the gcs1Delta resumption phenotype, and I show that Imh1 protein deleted for its GRIP domain alleviates the gcs1Delta reentry defect in high doses, while the GRIP domain alone does not, suggesting that Imh1 positive regulation occurs independent of GRIP-domain interactions. Thus I hypothesize that Imh1 plays a key role in mediating the transition from stationary phase to the cell division cycle. I also show that members of the 'Ypt6 pathway' involved in vesicle tethering, and the Osh1-Scs2 complex, negatively regulate the transition from stationary phase to the cell division cycle.