Current understanding of UV-induced base pair substitution mutation in E. coli with particular reference to the DNA polymerase III complex.

Abstract:

UV mutagenesis in E. coli is believed to occur in two discrete steps. The second step involves continued DNA synthesis beyond a blocking lesion in the template strand. This bypass step requires induced levels of umuD and umuC gene products and activated recA protein. DNA polymerase III may be involved since a dnaE mutator strain (believed to have defective base selection) is associated with enhanced UV mutagenesis in conjunction with a genetic background permitting the bypass step. In non-UV-mutable umu and lexA strains, UV mutagenesis can be demonstrated if delayed photoreversal is given. This is interpreted as indicating that an earlier misincorporation step can occur in such strains but the resulting mutations do not survive because the bypass step is blocked. The misincorporation step does not require any induced SOS gene products and can occur either at the replication fork or during repair replication following excision of a DNA lesion. Neither a dnaE mutator gene (leading to a defective alpha subunit of DNA polymerase III holoenzyme) nor a mutD5 mutator gene (leading to a defective epsilon proofreading subunit) had any effect on the misincorporation step. Although this is consistent with DNA polymerase III holoenzyme not being involved in the misincorporation step, other interpretations involving the inhibition of epsilon proofreading activity by recA protein are possible. In vitro studies are reported in which sites of termination of synthesis by DNA polymerase III holoenzyme on UV-irradiated M13 mp8 DNA were examined in the presence of inhibitors of the 3'-5' proofreading exonuclease (including recA protein). No evidence was found for incorporation of bases opposite photoproducts suggesting that either inhibition is more complete in the cell and/or that other factors are involved in the misincorporation step.

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