Roles of YqjH and YqjW, homologs of the Escherichia coli UmuC/DinB or Y superfamily of DNA polymerases, in stationary-phase mutagenesis and UV-induced mutagenesis of Bacillus subtilis

Abstract

YqjH and YqjW are Bacillus subtilis homologs of the UmuC/DinB or Y superfamily of DNA polymerases that are involved in SOS-induced mutagenesis in Escherichia coli. While the functions of YqjH and YqjW in B. subtilis are still unclear, the comparisons of protein structures demonstrate that YqjH has 36% identity to E. coli DNA polymerase IV (DinB protein), and YqjW has 26% identity to E. coli DNA polymerase V (UmuC protein). In this report, we demonstrate that both YqjH and the products of the yqjW operon are involved in UV-induced mutagenesis in this bacterium. Furthermore, resistance to UV-induced damage is significantly reduced in cells lacking a functional YqjH protein. Analysis of stationary-phase mutagenesis indicates that absences of YqjH, but not that of YqjW, decreases the ability of B. subtilis to generate revertants at the hisC952 allele via this system. These data suggest a role for YqjH in the generation of at least some types of stationary-phase-induced mutagenesis.

FIG. 1.

Survival curves of B. subtilis strains following UV irradiation. The results presented are averages of six different plates from each experiment for each strain following UV irradiation (see Material and Methods for details). Error bars represent 1 standard error. The results are representative, and the experiments were repeated at least four times.

FIG. 2.

UV mutagenesis of YB955 and isogenic derivatives thereof. The results presented are the average number of His⁺ revertants from six different plates from each experiment for each strain following UV irradiation (see Material and Methods for details). Error bars represent 1 standard error. The results are representative, and the experiments were repeated at least four times.

FIG. 3.

Stationary-phase-induced His⁺ reversion frequencies of YB955 and isogenic derivatives thereof. The results presented are the average number of revertants on each day from five different selection plates. Error bars represent 1 standard error, as described in Materials and Methods. These results are representative of experiments repeated at least three times.

FIG. 4.

Abilities of YB955 and isogenic derivatives thereof to survive under histidine starvation. Three plugs of bacteria containing agar were taken from selection plates each day for testing of the viability of bacteria on the selecting medium (see Material and Methods for details). The experiments were repeated at least twice.

FIG. 5.

Analysis of mutation rates. YB955 and isogenic derivatives of strain YB955 carrying the yqjH and/or yqjW alleles were tested for the ability to produce His⁺ revertants during exponential growth as described in Materials and Methods. The revertants were scored and recorded in 48 h after plating. The median (ŕ) is the mean of the 19th and 20th values of r (the observed number of mutants per culture in a 38-test-tube fluctuation test) when the r values are ranked. The number of mutations per culture (m) was calculated with the Lea-Coulson formula (ŕ/m − ln(m) = 1.24). Three parallel cultures were used to determine N_t by titration. The mutation rates were calculated with the formula m/2N_t (25, 35, 50). The results presented are the average mutation rates from three individual fluctuation tests. Error bars represent 1 standard error.