title: An underlying mechanism for the increased mutagenesis of lagging-strand genes in Bacillus subtilis.
abstract: We previously reported that lagging-strand genes accumulate mutations faster than those encoded on the leading strand in Bacillus subtilis. Although we proposed that orientation-specific encounters between replication and transcription underlie this phenomenon, the mechanism leading to the increased mutagenesis of lagging-strand genes remained unknown. Here, we report that the transcription-dependent and orientation-specific differences in mutation rates of genes require the B. subtilis Y-family polymerase, PolY1 (yqjH). We find that without PolY1, association of the replicative helicase, DnaC, and the recombination protein, RecA, with lagging-strand genes increases in a transcription-dependent manner. These data suggest that PolY1 promotes efficient replisome progression through lagging-strand genes, thereby reducing potentially detrimental breaks and single-stranded DNA at these loci. Y-family polymerases can alleviate potential obstacles to replisome progression by facilitating DNA lesion bypass, extension of D-loops, or excision repair. We find that the nucleotide excision repair (NER) proteins UvrA, UvrB, and UvrC, but not RecA, are required for transcription-dependent asymmetry in mutation rates of genes in the two orientations. Furthermore, we find that the transcription-coupling repair factor Mfd functions in the same pathway as PolY1 and is also required for increased mutagenesis of lagging-strand genes. Experimental and SNP analyses of B. subtilis genomes show mutational footprints consistent with these findings. We propose that the interplay between replication and transcription increases lesion susceptibility of, specifically, lagging-strand genes, activating an Mfd-dependent error-prone NER mechanism. We propose that this process, at least partially, underlies the accelerated evolution of lagging-strand genes.
authors.affiliation: Departments of Microbiology and.
authors.forename: Samuel
authors.initials: S
authors.lastname: Million-Weaver
authors.affiliation: Departments of Microbiology and.
authors.forename: Ariana N
authors.initials: AN
authors.lastname: Samadpour
authors.affiliation: Departments of Microbiology and.
authors.forename: Daniela A
authors.initials: DA
authors.lastname: Moreno-Habel
authors.affiliation: Departments of Microbiology and.
authors.forename: Patrick
authors.initials: P
authors.lastname: Nugent
authors.affiliation: Departments of Microbiology and.
authors.forename: Mitchell J
authors.initials: MJ
authors.lastname: Brittnacher
authors.affiliation: Departments of Microbiology and.
authors.forename: Eli
authors.initials: E
authors.lastname: Weiss
authors.affiliation: Departments of Microbiology and.
authors.forename: Hillary S
authors.initials: HS
authors.lastname: Hayden
authors.affiliation: Departments of Microbiology and.
authors.forename: Samuel I
authors.initials: SI
authors.lastname: Miller
authors.affiliation: Genome Sciences, University of Washington, Seattle, WA 98195.
authors.forename: Ivan
authors.initials: I
authors.lastname: Liachko
authors.affiliation: Departments of Microbiology and merrikh@uw.edu.
authors.forename: Houra
authors.initials: H
authors.lastname: Merrikh
mesh_terms: D001412:Bacillus subtilis / Q000235:genetics*; D004249:DNA Damage; D004261:DNA Replication; D005798:Genes, Bacterial; D016296:Mutagenesis; D014158:Transcription, Genetic
keywords: TCR; Y-family polymerases; excision repair; replication conflicts; transcription orientation