Comprehensive and Comparative Transcriptional Profiling of the Cell Wall Stress Response inBacillus subtilis

Abstract

The bacterial cell wall (CW) is an essential protective barrier and the frontline of cellular interactions with the environment and also a target for numerous antimicrobial agents. Accordingly, its integrity and homeostasis are closely monitored and rapid adaptive responses by transcriptional reprogramming induce appropriate counter-measures against perturbations. Here, we report a comprehensive and comparative transcriptional profiling of the primary cell envelope stress responses (CESR), based on combining RNAseq and high-resolution tiling array studies of the Gram-positive model bacteriumBacillus subtilisexposed to a range of antimicrobial compounds that interfere with cytoplasmic, membrane-coupled or extracellular steps of peptidoglycan (PG) biosynthesis. It revealed the complexity of the CESR ofB. subtilisand unraveled the contribution of extracytoplasmic function sigma factors (ECFs) and two-component signal transduction systems (TCSs) to protect the cell envelope. While membrane-anchored steps are tightly controlled, early cytoplasmic and late extracellular steps of PG biosynthesis are hardly monitored at all. The ECF σ factors σW and particularly σM provide a general CESR, while σV is almost exclusively induced by lysozyme, against which it provides specific resistance. Remarkably, σXwas slightly repressed by most antibiotics, pointing towards a role in envelope homeostasis rather than CESR. It shares this role with the WalRK TCS, which balances CW growth with controlled autolysis. In contrast, all remaining TCSs are envelope stress-inducible systems. LiaRS is induced by a wide range of PG synthesis inhibitors, while the three paralogous systems BceAB, PsdRS and ApeRS are more compound-specific detoxification modules. Induction of the CssRS TCS by all antibiotics interfering with membrane-anchored steps of PG biosynthesis points towards a physiological link between CESR and secretion stress. Based on the expression signatures, a suite of CESR-specificB. subtiliswhole cell biosensors were developed and carefully evaluated. This is the first comprehensive transcriptomic study focusing exclusively on the primary effects of envelope perturbances that shall provide a reference point for future studies on Gram-positive CESR.

TheBacillus subtiliscell envelope stress-inducibleytpABoperon modulates membrane properties and contributes to bacitracin resistance

“…Early genome-wide transcriptome studies have revealed that ytpAB is most strongly induced by inhibitors of the membrane-associated steps of peptidoglycan biosynthesis, and in particular by those compounds that interfere with the lipid II cycle such as bacitracin and vancomycin (18, 52). This pattern of response is consistent with a recent comprehensive profiling study using both RNA-seq and tiling array methodologies in which bacitracin and vancomycin were the strongest inducers, followed by tunicamycin, moenomycin, and lysozyme (53).…”

Implementation of Fluorescent-Protein-Based Quantification Analysis in L-Form Bacteria

“…It is worth noting that the cells cultured in NB/MSM showed a remarkable diversity of shapes, regardless of DCS addition. Surprisingly, there appears to be a correlation between the higher mKO2 fluorescence level and the presence of DCS, as enhanced fluorescence was observed (Figure S6A), presumably implying that DCS may affect promoters or other unknown intracellular targets [51].…”

Time-Course Transcriptome Analysis of Bacillus subtilis DB104 during Growth

“…For example, transcriptome or proteome analysis was conducted during specific growth stages, such as early to middle sporulation [ 12 ] and germination [ 13 ]. Zhang et al [ 14 ] revealed the transcriptional response to cell envelope stress in B . subtilis .…”

“…For example, transcriptome or proteome analysis was conducted during specific growth stages, such as early to middle sporulation [12] and germination [13]. Zhang et al [14] revealed the transcriptional response to cell envelope stress in B. subtilis. Nicolas et al [15] explored the regulatory architecture of B. subtilis using transcriptome analysis in a wide range of conditions.…”