LC–MS/MS proteomic analysis of starved Bacillus subtilis cells overexpressing ribonucleotide reductase (nrdEF): implications in stress-associated mutagenesis

Abstract

The non-appropriate conditions faced by nutritionally stressed bacteria propitiate error-prone repair events underlying stationary-phase- or stress-associated mutagenesis (SPM). The genetic and molecular mechanisms involved in SPM have been deeply studied but the biochemical aspects of this process have so far been less explored. Previous evidence showed that under conditions of nutritional stress, non-dividing cells of strain B. subtilis YB955 overexpressing ribonucleotide reductase (RNR) exhibited a strong propensity to generate true reversions in the hisC952 (amber), metB5 (ochre) and leuC425 (missense) mutant alleles. To further advance our knowledge on the metabolic conditions underlying this hypermutagenic phenotype, a high-throughput LC-MS/MS proteomic analysis was performed in non-dividing cells of an amino acid-starved strain, deficient for NrdR, the RNR repressor. Compared with the parental strain, the level of 57 proteins was found to increase and of 80 decreases in the NrdR-deficient strain. The proteomic analysis revealed an altered content in proteins associated with the stringent response, nucleotide metabolism, DNA repair, and cell signaling in amino acid-starved cells of the ∆nrdR strain. Overall, our results revealed that amino acid-starved cells of strain B. subtilis ∆nrdR that escape from growth-limiting conditions exhibit a complex proteomic pattern reminiscent of a disturbed metabolism. Future experiments aimed to understand the consequences of disrupting the cell signaling pathways unveiled in this study, will advance our knowledge on the genetic adaptations deployed by bacteria to escape from growth-limiting environments.

Transcriptome profiling of gene expression in phosphate-solubilizing bacterium Acinetobacter sp. strain m01 interacting with melon (Cucumis melo L.) seedling

“…This phenomenon may be attributed to restrained cell growth due to limited nutrients. Under nutritional stress, bacteria exhibit low specific growth rates, for instance, Bacillus subtilis keeps long periods of non-dividing situation (Cummings and Macfarlane 1997;Castro-Cerritos et al 2018) that may lead to reduced gene expression. Moreover, many genes were upregulated in melon plant under PSB M01 inoculation (Figure 2), indicating that M01 can stimulate the gene expression to improve plant growth.…”

Mfd affects global transcription and the physiology of stressed Bacillus subtilis cells

“…The changes in gene expression of IMP biosynthetic genes are likely to affect nucleotide pools and induce a more sensitive state to trimethoprim; such sensitivity was reduced when mfd was overexpressed. Interestingly, activity of the ribonuclotide reductase, encoded by nrdEF , influences Mfd - dependent mutagenesis (Castro-Cerritos et al, 2017;Castro-Cerritos et al, 2018). The response in daptomycin correlated with low gene expression of the liaRS operon, which codes for factors that sense cell-envelope stress.…”