THE RAD21 GENE-PRODUCT OF SCHIZOSACCHAROMYCES-POMBE IS A NUCLEAR, CELL-CYCLE-REGULATED PHOSPHOPROTEIN

The rad21 gene of Schizosaccharomyces pombe is involved in the repair of double-strand breaks in DNA and is essential for mitotic growth (Birkenbihl, R. P., and Subramani, S. (1992) Nucleic Acids Res. 20, 6605-6611), We show that the Rad21 protein migrates with an aberrantly slow mobility, has a thrombin cleavage site, and is multiply phosphorylated mainly at serine residues. The expression of the rad21 mRNA and the Rad21 protein is cell cycle-regulated, with the peak of mRNA and protein expression occurring near the G(1) to S transition. Following translation of the protein, hypophosphorylated forms of the protein appear. However, the most phosphorylated form of Rad21 appears only later in the cell cycle (in S to G(2)). Analysis of the radiosensitive mutant rad21-45 revealed that the mutant protein is permanently hypophosphorylated. The Rad21 protein is nuclear during the cell cycle. The nuclear localization signal was identified in the C-terminal third of the protein. Upon repression of the Rad21 protein expressed from the repressible nmt1 promoter, the unphosphorylated and hypophosphorylated forms of Rad21 disappeared first. When the concentration of the most highly phosphorylated form of Rad21 sank under a critical level, the cells underwent aberrant mitoses, They exhibited loss of proper nuclear organization and abnormal septation.