The Escherichia coli polB gene, which encodes DNA polymerase II, is regulated by the SOS system

Abstract

The dinA (damage inducible) gene was previously identified as one of the SOS genes with no known function; it was mapped near the leuB gene, where the poiB gene encoding DNA polymerase H was also mapped. We cloned the chromosomal fragment carrying the dinA region from the ordered Escherichia coli genomic library and mapped the dinA promoter precisely on the physical map of the chromosome. The cells that harbored multicopy plasmids with the dimA region expressed very high levels of DNA polymerase activity, which was sensitive to N-ethylmaleimide, an inhibitor of DNA polymerase II. Expression of the polymerase activity encoded by the dinA locus was regulated by SOS system, and the dinA promoter was the promoter of the gene encoding the DNA polymerase. From these data we conclude that the polB gene is identical to the dinA gene and is regulated by the SOS system. The product of the polB (dinA) gene was identified as an 80-kDa protein by the maxicell method.In Escherichia coli, three DNA polymerases have been identified (15). DNA polymerase III has been shown to be essential for chromosomal DNA replication and to consist of multiple species of subunits. The nucleotide polymerizing activity resides in the a subunit encoded by the dnaE (poIC) gene. DNA polymerase I, encoded by polA, has been shown to be involved in DNA repair and to be required for DNA replication when the cell is growing fast on rich medium (11). However, no biological role has been assigned to DNA polymerase II, encoded by polB. The polB mutants that have been isolated exhibit no defect in DNA replication and DNA repair (5, 9).We suspected that the existing polB mutants retained substantial levels of DNA polymerase II activity in vivo and that this might have prevented us from revealing the specific phenotype of the mutants so far. To elucidate the role of DNA polymerase II in DNA replication, repair, recombination, and mutagenesis, we thought it necessary to construct apolB deletion mutant by using a cloned gene (25). Biochemical studies on DNA polymerase II have been hampered by the relative scarcity of the enzyme in the cell. The overproduction of the enzyme with the cloned gene would facilitate these studies. Therefore, we have been trying to clone the polB gene. Bonnar et al. (3) presented evidence that synthesis of DNA polymerase X, presumably DNA polymerase II, is regulated by LexA repressor. The dinA gene (13) was identified as an SOS-regulated gene with no known function; it was mapped near leuB, where polB was also mapped (4, 9). It occurred to us that dinA might be identical to polB. In this work, we examined this possibility and found that it is indeed so. The polB gene was recently cloned by Chen et al. (6).MATERIALS AND METHODS Bacterial strains, plasmids, phages, and media. The E. coli strains used in this study are listed in Table 1. Strain HRS5200 was constructed by P1 transduction of leu+ and polB1OO of HMS83 (5) cysteine, and 14C-labeled proteins for molecular weight standards were purchased from Amersham Japan (Tokyo, Japan). M...

Gene duplications in evolution of archaeal family B DNA polymerases

“…Scenario (i), which postulates a loss of a cellular encoded family B DNA polymerase, has been observed in two eubacteria. Neither Haemophilus influenzae Rd nor Mycoplasma genitalium possesses a family B homolog (18,22), yet this DNA polymerase is present in E. coli, where it functions as a repair polymerase (4,8,26). This DNA polymerase must have been lost independently in the lineages leading to H. influenzae and Mycoplasma genitalium.…”

Roles of DNA polymerases V and II in SOS-induced error-prone and error-free repair in Escherichia coli

“…A tentative link between the two was established when it was determined that pol II was induced as part of the LexA-regulon (2). pol II was subsequently shown to be encoded by the DNA damage-inducible polB (dinA) gene (3)(4)(5). A ⌬polB strain shows no measurable UV sensitivity, and SOS-induced mutagenesis occurs at normal levels (6,7).…”

The SOS response regulates adaptive mutation

“…A central part of the SOS response is the de-repression of more than 20 genes under the direct and indirect transcriptional control of the LexA repressor. The LexA regulon includes recombination and repair genes recA, recN, and ruvAB, nucleotide excision repair genes uvrAB and uvrD, the error-prone DNA polymerase (pol) genes dinB (encoding pol IV) (2) and umuDC (encoding pol V) (3), and DNA polymerase II (4,5) in addition to many functions not yet understood. In the absence of a functional SOS response, cells are sensitive to DNA damaging agents.…”