Evidence for substrate control of Cdk phosphorylation timing during the budding yeast cell cycle1757

The main intention of this project is to investigate how Cdk substrate phosphorylation is ordered during the cell cycle. We want to understand what makes a ‘good’ Cdk substrate that is phosphorylated early during the cell cycle, e. g. to promote S phase, as compared to a ‘bad’ Cdk substrate that is phosphorylated much later, e. g. when cells enter mitosis. This could be because a late substrate is indeed a poor Cdk substrate, or it could be that the late substrate is a good substrate for Cdk counteracting phosphatases. The two possibilities are not mutually exclusive. To compare phosphorylation and dephosphorylation dynamics we arrest cells in mitosis when most Cdk substrates are fully phosphorylated. Then, we acutely, and chemically inhibit Cdk. The rate of Cdk substrate dephosphorylation will be measured, which will be an indication of the efficiency of Cdk counteracting phosphatases on each substrate.