In Escherichia coli, the dinB gene is required for the SOS-induced lambda untargeted mutagenesis pathway and confers a mutator phenotype to the cell when the gene product is overexpressed. Here, we report that the purified DinB protein is a DNA polymerase. This novel E. coli DNA polymerase (pol IV) is shown to be strictly distributive, devoid of proofreading activity, and prone to elongate bulged (misaligned) primer/template structures. Site-directed mutagenesis experiments of dinB also demonstrate that the polymerase activity of DinB is required for its in vivo mutagenicity. Along with the sequence homologies previously found within the UmuC-like protein family, these results indicate that the uncovered DNA polymerase activity may be a common feature of all these homologous proteins.

The hOGG1 gene encodes a DNA glycosylase that excises 8-hydroxyguanine (oh 8 Gua) from damaged DNA. Structural analyses of the hOGG1 gene and its transcripts were performed in normal and lung cancer cells. Due to a genetic polymorphism at codon 326, hOGG1-Ser 326 and hOGG1-Cys 326 proteins were produced in human cells. Activity in the repair of oh 8 Gua was greater in hOGG1-Ser 326 protein than in hOGG1-Cys 326 protein in the complementation assay of an E. coli mutant defective in the repair of oh 8 Gua. Two isoforms of hOGG1 transcripts produced by alternative splicing encoded distinct hOGG1 proteins: one with and the other without a putative nuclear localization signal. Loss of heterozygosity at the hOGG1 locus was frequently (15/ 23, 62.2%) detected in lung cancer cells, and a cell line NCI-H526 had a mutation leading to the formation of the transcripts encoding a truncated hOGG1 protein. However, the oh 8 Gua levels in nuclear DNA were similar among lung cancer cells and leukocytes irrespective of the type of hOGG1 proteins expressed. These results suggest that the oh 8 Gua levels are maintained at a steady level, even though multiple hOGG1 proteins are produced due to genetic polymorphisms, mutations and alternative splicing of the hOGG1 gene.

ABSTRACTdinP is an Escherichia coli gene recently identified at 5.5 min of the genetic map, whose product shows a similarity in amino acid sequence to the E. coli UmuC protein involved in DNA damage-induced mutagenesis. In this paper we show that the gene is identical to dinB, an SOS gene previously localized near the lac locus at 8 min, the function of which was shown to be required for mutagenesis of nonirradiated phage infecting UV-preirradiated bacterial cells (termed UTM for untargeted mutagenesis). A newly constructed dinP null mutant exhibited the same defect for UTM as observed previously with a dinB::Mu mutant, and the defect was complemented by plasmids carrying dinP as the only intact bacterial gene. Furthermore, merely increasing the dinP gene expression, without UV irradiation or any other DNAdamaging treatment, resulted in a strong enhancement of mutagenesis in Flac plasmids; at most, 800-fold increase in the G6-to-G5 change. The enhanced mutagenesis did not depend on recA, uvrA, or umuDC. Thus, our results establish that E. coli has at least two distinct pathways for SOS-induced mutagenesis: one dependent on umuDC and the other on dinB͞P.