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Molecular analysis of mutations in the hprt gene of V79 hamster fibroblasts: effects of imbalances in the dCTP, dGTP and dTTP pools.

Abstract：

:dCMP-deaminase-deficient V79/dC hamster cells have highly imbalanced deoxyribonucleoside triphosphate (dNTP) pools, i.e. a 17-fold larger dCTP pool, a slightly reduced dTTP and a very low dGTP pool, compared to dCMP-deaminase-proficient V79/p cells. Nevertheless, the two lines showed the same rates of spontaneous mutation at the hprt and ouabain-resistance loci. Analysis of spontaneous hprt mutations indicated an increase in misincorporation of C in V79/dC cells, although it was not statistically significant. When the dCTP pool was further increased fivefold by incubating V79/dC cells with cytidine, C misincorporation increased to 88%, but the mutation frequency remained unchanged. The dNTP pools of V79/dC cells were also altered by treatment with thymidine, or with thymidine plus deoxycytidine. After incubation with thymidine alone, the dCTP pool all but disappeared, whereas it maintained a normal level in the presence of deoxycytidine. In both cases dTTP rose to nmol amounts, and dGTP accumulated. Incubation with 10 mM thymidine was the only treatment that increased the mutation frequency; T misincorporation then accounted for 94% of the base substitutions. In the presence of deoxycytidine the cells had a dTTP/dCTP ratio of 0.04, but 86% of the base substitutions involved C misincorporation and most probably originated from G mis-incorporation caused by excess dGTP. Alterations of RNA splicing and hot spots for base substitutions varied with the imbalance, the latter showed "next-nucleotide effects". Our results suggest that the fidelity of DNA replication in V79 cells is only affected by large changes in the pool and is more sensitive to changes in dGTP than in dCTP or dTTP.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Darè E,Zhang LH,Jenssen D,Bianchi V

doi

10.1006/jmbi.1995.0516

subject

Has Abstract

pub_date

1995-10-06 00:00:00

pages

514-21

issue

5

eissn

0022-2836

issn

1089-8638

pii

S0022283685705165

journal_volume

252

pub_type

杂志文章

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