DNA polymerase α in the fission yeast Schizosaccharomyces pombe: Identification and tracing of the catalytic subunit during the cell cycle

Abstract

A recombinant protein was obtained in Escherichia coli by subcloning part of the Schizosaccharomyces pombe POL1 gene at the 3′-end of lacZ. Antibodies raised against this protein were used to identify the POL1 gene product in extracts of exponentially growing S. pombe cells. A major 170-kDa protein, whose structure and properties were typical of the catalytic subunit of eukaryotic DNA polymerases α (pol α), was detected. The same antibodies were used to trace pol α and to quantify its level during the S. pombe cell cycle. We found that pol α was present at all stages of the cycle and that its cellular pool was subject to limited (threefold) increase in G 1 and S phases, with a decline to the initial level soon after. In addition, we found that a second form of pol α with slightly lower molecular weight (165 kDa) existed only during late G 1 and S phases. Moreover, absence of initiation or perturbations in the course of DNA replication induced overproduction of the 165-kDa form.