The uracil-DNA glycosylase activity was compared in cell-free extracts of normal (NHSF6) and Bloom's syndrome (BS) skin fibroblasts (BS1KA and BS2KA from Japanese patients) at middle culture age. The enzyme activity in the extracts of exponentially growing NHSF6 and BS2KA cells increased linearly with the DNA synthetic activity, while such a relation was not obvious in BS1KA cells. The thermal stability and the inhibition by the end product uracil of the BS1KA enzyme did not differ from those of the NHSF6 enzyme. Synchronized-cell studies showed the following characteristics. (i) Uracil-DNA glycosylase activity was enhanced in a temporal sequence only during S phase and reached a peak level a few hours prior to that of DNA synthesis in NHSF6. (ii) BS2KA cells were normal in the temporal induction sequence but BS1KA cells revealed the delayed peak induction of the enzyme occurring simultaneously with peak DNA synthesis. (iii) With progress of culture passage, the uracil-DNA glycosylase activity became highly expressed at G0 and during G1 despite a little change in the S-phase activity in NHSF6 cells. (iv) Another abnormality of BS1KA cells was that such a culture-age-dependent dysregulation occurred earlier during middle passages. The above results (i) and (ii) suggest that the delayed enzyme induction in BS1KA cells may be related to the observation that BS1KA cells were more sensitive to 5-bromodeoxyuridine-induced cell killing and to sister chromatid exchange formation than BS2KA cells of the clinically milder subject.