Isolation, characterisation and molecular cloning of new mutant alleles of the fission yeast p34 cdc2+ protein kinase gene: identification of temperature-sensitive G2-arresting alleles

Abstract

The protein serine-threonine kinase p34cdc2+ plays a central role in the control of the mitotic cell cycle of the fission yeast Schizosaccharomyces pombe. p34cdc2+ function is required both for the initiation of DNA replication and for entry into mitosis, and is also required for the initiation of the second meiotic nuclear division. Recent extensive analysis of p34cdc2+ homologue proteins in higher eukaryotes has demonstrated that p34cdc2+ function is likely to be conserved in all eukaryotic cells. Here we report the isolation and characterisation of five new temperature-sensitive alleles of the cdc2+ gene. All five have been cloned and sequenced, together with the meiotically defective cdc2-N22 allele, bringing the total of p34cdc2+ mutants cloned in this and previous reports to seventeen. The five temperature-sensitive alleles define four separate mutations within the p34cdc2+ protein sequence, two of which give rise to cell cycle arrest in G2 only, when shifted to the restrictive temperature. The nature of the mutation in each protein is described and possible implications for the structure and function of p34cdc2+ discussed.