Thymidine 5′-Monophosphate-Requiring Mutants of Saccharomyces cerevisiae Are Deficient in Thymidylate Synthetase
- 1 October 1977
- journal article
- research article
- Published by American Society for Microbiology in Journal of Bacteriology
- Vol. 132 (1), 44-50
- https://doi.org/10.1128/jb.132.1.44-50.1977
Abstract
Thymidylate synthetase activity was measured in crude extracts of the yeast S. cerevisiae by a sensitive radiochemical assay. Spontaneous non-conditional mutants auxotrophic for TMP (tmp1) lacked detectable thymidylate synthetase activity in cell-free extracts. In contrast, the parent strains (tup1, -2 or -4), which were permeable to TMP, contained levels of activity similar to those found in wild-type cells. Specific activity of thymidylate synthetase in crude extracts of normal cells or of cells carrying tup mutations was essentially unaffected by the ploidy or mating type of the cells, by the medium used for growth by the respiratory capacity of the cells, by concentrations of exogenous TMP as high as 50 .mu.g/ml or by subsequent removal of TMP from the medium. Extracts of a strain bearing the temperature-sensitive cell division cycle mutation cdc21 lacked detectable thymidylate synthetase activity under all conditions tested. Its parent and another mutant (cdc8), which arrests with the same terminal phenotype under restrictive conditions, had normal levels of the enzyme. Cells of a temperature-sensitive TMP auxotroph was arrested with a morphology identical to the cdc21 strain at the nonpermissive temperature and contained demonstrably thermolabile thymidylate synthetase activity. Tetrad analysis and the properties of revertants showed that the thymidylate synthetase defects were a consequence of the same mutation causing, in the auxotrophs, a requirement for TMP and, in the conditional mutants, temperature sensitivity. Complementation tests indicated that tmp1 and cdc21 are the same locus. These results identify tmp1 as the structural gene for yeast thymidylate synthetase.This publication has 23 references indexed in Scilit:
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