Novel rpoB mutations conferring rifampin resistance on Bacillus subtilis: global effects on growth, competence, sporulation, and germination

Abstract

Previously, spontaneous rifampin resistance mutations were isolated in cluster I of the rpoB gene, resulting in amino acid replacements (Q469R, H482R, H482Y, or S487L) in the Bacillus subtilis RNA polymerase beta subunit (W. L. Nicholson and H. Maughan, J. Bacteriol. 184:4936-4940, 2002). In this study, each amino acid change in the beta subunit was observed to result in its own unique spectrum of effects on growth and various developmental events, including sporulation, germination, and competence for transformation. The results thus establish the important role played by the RNA polymerase beta subunit, not only in the catalytic aspect of transcription, but also in the regulation of major developmental events in B. subtilis.

FIG. 1.

Growth rates of isogenic strains carrying the indicated rpoB alleles in LB medium (white bars) or LB medium plus Rif (hatched bars). Data are expressed as average doubling times in minutes ± standard deviations (n = 3). Data that were not significantly different (P > 0.01 by ANOVA) were placed into the same group (groups are indicated by lowercase letters). w.t., wild type.

FIG. 2.

Spore germination of Rif^r mutants and their wild-type (w.t.) parental strain. Germinated spore titers after 24 h at 48°C (hatched bars) or after a shift from 24 h at 48°C to an additional 24 h at 37°C (black bars) are depicted, normalized to the germinated spore titers of parallel plates of the same strain after 24 h of incubation at 37°C (white bars). Plus and minus signs indicate that spores were produced by growth in SMM in the presence or absence, respectively, of RIF (50 μg/ml). Strains are identified by the amino acid alteration in rpoB. Results are from two independent experiments performed in duplicate, from which the data differed by <10%.