Dividing cells require of a strict regulation of cell cycle in order to complete it correctly and obtain two healthy daughter cells genetically identical. Mitosis is a highly conserved and regulated process in eukaryotes. Mitosis is divided in four different phases: prophase, metaphase, anaphase and telophase. My research project is focused in mitotic exit regulation, concretely the mechanisms that ensure proper anaphase completion before exit from mitosis.
In Saccharomyces cerevisiae mitotic exit is regulated by two different pathways: FEAR (or Cdcfourteen Early Anaphase Release) and MEN (Mitotic Exit Network). Both cooperate in the Cdc14 phosphatase release from the nucleolus and activation. Cdc14 release is initiated by FEAR-dependent Net1 phosphorylation and allows the cells to exit from mitosis.
Net1 is phosphorylated by Clb2/Cdc28 complex (Cyclin/Cyclin-dependent-kinase complex) and Cdc5 kinase. Cdc5 is a polo-like kinase that mediates Cdc14 release regulating both FEAR and MEN.
Cdc5 direct role in phosphorylating Net1 in vivo has not been demonstrated so far. We have demonstrated how Cdc5 is activated by Cdc28, activation required for its kinase action. Then, we have mapped by mass spectrometry analysis and peptide array the presumably phosphorylated Net1 residues by Cdc5. We are currently studying the in vivo individual contribution of each Net1 residue phosphorylation by Cdc5.