Using Schizosaccharomyces pombe to isolate repressors of Xenopus laevis Wee2 kinase activity

The cell cycle is evolutionary conserved among eukaryotes. Here we use this principle of conservation to identify repressors of the Xenopus Wee2 (XWee2) kinase activity using S.pombe as a model organism. Wee2 is a conserved inhibitor of mitotic entry. Active Wee2 blocks entry into mitosis by phosphorylating and inhibiting the mitotic cyclin dependent kinase, Cdk1. When Xenopus Wee2 is over expressed in S.pombe, it causes the yeast to arrest in G2. The expression of a Xenopus cDNA that represses the XWee2 kinase activity would be expected to rescue the G2 arrest in the yeast and allow cells to enter mitosis. A S.pombe strain engineered to conditionally over-express XWee2 was transformed with three different Xenopus oocyte cDNA libraries and selection for suppressors of the Wee2 kinase was carried out. Here we describe the challenges in performing this selection and report the isolation and analysis of one possible Xenopus cDNA suppressor candidate (pAXADH-SC96) that partially suppresses the G2 arrest phenotype of S.pombe over expressing Xenopus Wee2.