Isolation and properties of yeast mutants with highly efficient thymidylate utilization.
A screening procedure is presented which allows the isolation of yeast mutants (typ tir) with highly efficient utilization of exogenous deoxythymidine-5'-monophosphate (5'-dTMP) (greater than 50%). Data are given concerning the phenomenon of 5'-dTMP utilization in general: (i) The ability of S. cerevisiae to incorporate exogenous 5'-dTMP was found to already to be a wild type feature of this yeast, i.e. apparently not to be due to any mutation such as typ, tup, tmp, per or tum. Consequently these mutations are interpreted as amplifiers of a pre-given wild type potency. So far eight stages of 5'-dTMP utilization were detected as classified by the optimal 5'-dTMP requirement, with 5'-dTMP biosynthesis blocked, of the corresponding mutant strains isolated. All of them fit well into a mathematical series of the type "2n x 1.5" (n = 0, 1, 2, ..., 11), where the product term for n = 11 represents the 5'-dTMP requirement (mug/ml) of the best 5'-dTMP utilizing wild type strain found. (ii) Amplification of the 5'-dTMP utilizing potency obviously is due to any genetically determined alteration of the yeast 5'-dTMP uptaking principle itself or of physiological processes accompanying the monophosphate's uptake. (iii) The functioning of 5'-dTMP uptake requires acidic (less than or equal to pH 6) conditions in the yeast cell's outer environment. (iv) Some yeast typ and typ tlr mutants were found to exhibit a more or less pronounced sensitivity towards exogenously offered 5'-dTMP. The response of a sensitive strain towards inhibitory concentrations of the nucleotide apparently is co-conditioned by the presence or absence of thymidylate biosynthesis. With 5'-dTMP biosynthesis blocked the 5'-dTMP mediated inhibition is a permanent one and finally leads to the death of a cell. With a functioning thymidylate biosynthesis, in contrast, the inhibition is only temporary. (v) Yeast typ or typ tlr strains were observed to dephosphorylate exogenous 5'-dTMP to thymidine due to a phosphatase activity which cannot be eliminated at pH 7 + 70 mM inorganic phosphate conditions in the growth medium. This 5'-dTMP cleavage obviously occurs outside the cell and does not seem to be correlated both to the monophosphate's uptake and to the phenomenon of 5'-dTMP sensitivity. The destruction of 5'-dTMP does not disturb (5'-dTMP) DNA-specific labelling.
Isolation and properties of yeast mutants with highly efficient thymidylate utilization. A screening procedure is presented which allows the isolation of yeast mutants (typ tir) with highly efficient utilization of exogenous deoxythymidine-5'-monophosphate (5'-dTMP) (greater than 50%). Data are given concerning the phenomenon of 5'-dTMP utilization in general: (i) The ability of S. cerevisiae to incorporate exogenous 5'-dTMP was found to already to be a wild type feature of this yeast, i.e. apparently not to be due to any mutation such as typ, tup, tmp, per or tum. Consequently these mutations are interpreted as amplifiers of a pre-given wild type potency. So far eight stages of 5'-dTMP utilization were detected as classified by the optimal 5'-dTMP requirement, with 5'-dTMP biosynthesis blocked, of the corresponding mutant strains isolated. All of them fit well into a mathematical series of the type "2n x 1.5" (n = 0, 1, 2, ..., 11), where the product term for n = 11 represents the 5'-dTMP requirement (mug/ml) of the best 5'-dTMP utilizing wild type strain found. (ii) Amplification of the 5'-dTMP utilizing potency obviously is due to any genetically determined alteration of the yeast 5'-dTMP uptaking principle itself or of physiological processes accompanying the monophosphate's uptake. (iii) The functioning of 5'-dTMP uptake requires acidic (less than or equal to pH 6) conditions in the yeast cell's outer environment. (iv) Some yeast typ and typ tlr mutants were found to exhibit a more or less pronounced sensitivity towards exogenously offered 5'-dTMP. The response of a sensitive strain towards inhibitory concentrations of the nucleotide apparently is co-conditioned by the presence or absence of thymidylate biosynthesis. With 5'-dTMP biosynthesis blocked the 5'-dTMP mediated inhibition is a permanent one and finally leads to the death of a cell. With a functioning thymidylate biosynthesis, in contrast, the inhibition is only temporary. (v) Yeast typ or typ tlr strains were observed to dephosphorylate exogenous 5'-dTMP to thymidine due to a phosphatase activity which cannot be eliminated at pH 7 + 70 mM inorganic phosphate conditions in the growth medium. This 5'-dTMP cleavage obviously occurs outside the cell and does not seem to be correlated both to the monophosphate's uptake and to the phenomenon of 5'-dTMP sensitivity. The destruction of 5'-dTMP does not disturb (5'-dTMP) DNA-specific labelling.