Bacillus subtilisfunctional genomics: global characterization of the stringent response by proteome and transcriptome analysis

Abstract

The stringent response in Bacillus subtilis was characterized by using proteome and transcriptome approaches. Comparison of protein synthesis patterns of wild-type and relA mutant cells cultivated under conditions which provoke the stringent response revealed significant differences. According to their altered synthesis patterns in response to DL-norvaline, proteins were assigned to four distinct classes: (i) negative stringent control, i.e., strongly decreased protein synthesis in the wild type but not in the relA mutant (e.g., r-proteins); (ii) positive stringent control, i.e., induction of protein synthesis in the wild type only (e.g., YvyD and LeuD); (iii) proteins that were induced independently of RelA (e.g., YjcI); and (iv) proteins downregulated independently of RelA (e.g., glycolytic enzymes). Transcriptome studies based on DNA macroarray techniques were used to complement the proteome data, resulting in comparable induction and repression patterns of almost all corresponding genes. However, a comparison of both approaches revealed that only a subset of RelA-dependent genes or proteins was detectable by proteomics, demonstrating that the transcriptome approach allows a more comprehensive global gene expression profile analysis. The present study presents the first comprehensive description of the stringent response of a bacterial species and an almost complete map of protein-encoding genes affected by (p)ppGpp. The negative stringent control concerns reactions typical of growth and reproduction (ribosome synthesis, DNA synthesis, cell wall synthesis, etc.). Negatively controlled unknown y-genes may also code for proteins with a specific function during growth and reproduction (e.g., YlaG). On the other hand, many genes are induced in a RelA-dependent manner, including genes coding for already-known and as-yet-unknown proteins. A passive model is preferred to explain this positive control relying on the redistribution of the RNA polymerase under the influence of (p)ppGpp.

Transcriptome Analysis of Sorbic Acid-Stressed Bacillus subtilis Reveals a Nutrient Limitation Response and Indicates Plasma Membrane Remodeling

“…The induction of transition-state and early-stationary-phase genes suggests that sorbic acidstressed cells respond as if they experience nutrient limitation. In amino acid-, glucose-, or oxygen-limited cells, the (RelAmediated) stringent response helps to prevent the waste of scarce nutrients (9,26). Hierarchical clustering revealed that only 8 out of 55 (P ϭ 0.017) RelA-dependent positive stringent control genes were significantly induced and that 4 out of 86 (P ϭ 0.79) RelA-dependent negative stringent control genes were significantly repressed.…”

“…The regulation of this operon is complex, as it is regulated by PucR, TnrA, CodY, and GlnR, and expression is dependent on SigA and SigH (10,72). In addition, the induction of ureA is also RelA dependent (26). Thus, the induction of the ureABC operon may be explained by the derepression of CodY-regulated genes and the induction of the stringent response ( Fig.…”

“…T-profiler was adapted for the use of B. subtilis transcriptome data by implementing predefined gene groups from the following sources: the Database of Transcriptional Regulation in Bacillus subtilis (DBTBS) (release May 2006) (47), the Kyoto Encyclopedia of Genes and Genomes database (KEGG) (release May 2006) (39), the SubtiList database (54), and the stringently controlled genes (26). Gene groups regulated positively or negatively by a specific transcription factor were named accordingly.…”

Post-transcriptional regulation of the Bacillus subtilis pst operon encoding a phosphate-specific ABC transporter

“…Total RNA was extracted from B. subtilis strains 168, 168-PR and ML6 (Table 1) as described previously (Eymann et al, 2002). Northern blot analysis was performed according to the manufacturer's instructions (DIG Northern Starter kit, Roche Diagnostics), using 5 mg total RNA per lane.…”

Integrated metatranscriptomic and metagenomic analyses of stratified microbial assemblages in the open ocean

“…Transcript abundance will not always correlate directly with cognate protein levels, and the kinetics that couple gene expression to phenotypic manifestation varies among different transcript classes (Steunou et al, 2008). Nonetheless, reasonably good correlation between transcriptomes and proteomes, especially for transcripts and peptides in higher abundance, has been observed in several model organisms (Eymann et al, 2002;Corbin et al, 2003;Scherl et al, 2005). In the Pelagibacter genome-centric analyses reported in this study (Figure 5), we observed considerable overlap between highly abundant transcripts and the most represented peptides previously reported in a SAR11-centric metaproteomic study (Sowell et al, 2008).…”