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  • Title: Bromodeoxyuridine (BrdU) template and thymidine pool effects on high frequencies of sister-chromatid exchange (SCE) in Bloom syndrome cells and a mutant cell line (AsHa) originated from ataxia telangiectasia.
    Author: Shiraishi Y, Li MJ.
    Journal: Mutat Res; 1990 Jun; 230(2):177-86. PubMed ID: 2374555.
    Abstract:
    The effect of bromodeoxyuridine (BrdU)-substituted DNA template and thymidine (dT) pool on excess sister-chromatid exchanges (SCEs) was studied in Bloom syndrome (BS) cells and an ataxia telangiectasia (AT)-derived mutant cell line (AsHa). When BS endomitotic cells were labeled with low and high (or high and low) BrdU concentrations during S1 and S2, only the BrdU concentration during S1 phase affected the observed SCE. In BS cells about a 10-fold increase in SCEs occurs during or following replication on a BrdU-substituted template (high-high and high-low BrdU labeling) relative to the normal DNA template. SCEs decreased to about half in AsHa cells labeled with various BrdU doses (40, 60, 80 and 100 micrograms/ml) during only S1, compared with those labeled during S1 and S2. Co-cultivation of AsHa and BS cells resulted in a significant reduction in SCE level from 70 to 13-17 in BS cells, lowered the BrdU concentrations necessary for sister-chromatid differential (SCD) staining from 40 to 10 micrograms/ml with normal SCE level and resulted in decreased level of SCEs at high BrdU concentrations (80-100 micrograms/ml) (12-14 SCE) in AsHa cells, compared with the originally increased SCE level (36.65 SCE at 100 micrograms/ml) without co-culture. However, co-cultivation between AsHa and normal cells lowered the BrdU dose necessary for SCD staining from 40 to 30 micrograms/ml; the dT pool possibly balanced at this level, which is clearly higher than that at co-cultivation between AsHa and BS cells. The reason for the very high BrdU doses needed to achieve SCD would seem to be that AsHa cells have high levels of thymidylate (TMP) synthetase, which maintain a large endogenous thymidine pool. This has been confirmed by direct measurement. These findings strongly support that excess and decreased dT pools are closely related to the condition necessary for high SCE induction.
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