Nucleic Acids Research, 1993, Vol. 21, No. 16 3821-3828
© 1993
ENZYMOLOGY |
Fission yeast with DNA polymerase 
temperature-sensitive alleles exhibits cell division cycle phenotype
Laboratory of Experimental Oncology, Department of Pathology and Department of Pharmacology, Stanford University School of Medicine Stanford, CA 94305-5324, USA
*To whom correspondence should be addressed
Received March 30, 1993. Revised June 29, 1993. Accepted June 29, 1993.
DNA polymerases 
and 
are essential enzymes believed to play critical roles in initiation and replication of chromosome DNA. In this study, we show that the genes for Schizosaccharomyces pombe (S.pombe) DNA polymerase and (pol+ and pol+) are essential for cell viability. Disruption of either the pol+ or pol+ gene results in distinct terminal phenotypes. The S.pombe pol+ gene is able to complement the thermosensitive cdc2-2 allele of Saccharomyces cerevisiae (S.cerevlslae) at the restrictive temperature. By random mutagenesis in vitro, we generated three pol conditional lethal alleles. We replaced the wild type chromosomal copy of pol+ gene with the mutagenized sequence and characterized the thermosensitlve alleles in vivo. All three thermosensltive mutants exhibit a typical cell division cycle (cdc) terminal phenotype similar to that of the disrupted pol+ gene. Flow cytometric analysis showed that at the nonpermissive temperature all three mutants were arrested in S phase of the cell cycle. The three S.pombe conditional pol alleles were recovered and sequenced. The mutations causing the thermosensltive phenotype are mlssense mutations. The altered amlno acid residues are uniquely conserved among the known polymerase 5 sequences.
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