Bacillus subtilis 168, WT vs. ΔMWX, WT vs. Δ7ECF, Δ7ECF vs. ΔMWX.. Bacillus subtilis subsp. subtilis str. 168

Bacillus subtilis encodes seven extracytoplasmic function (ECF) sigma factors. Three (sigma M, sigma W and simga X) mediate responses to cell envelope active antibiotics. The functions of sigma Y, sigma Z, sigma V, and YlaC remain largely unknown, and strong inducers of these sigma factors and their regulons have yet to be defined. Here, we define transcriptomic and phenotypic differences under non-stress conditions between strains carrying deletions in all seven ECF sigma factor genes (Δ7ECF), a sigMWX triple mutant (∆MWX), and the parental 168 strain. Our results identify >80 genes as at least partially dependent on ECF sigma factors and, as expected, most of these are dependent on sigma M, sigma W or sigma X which are active at a significant basal level during growth. Several genes, including the eps operon encoding enzymes for exopolysaccharide (EPS) production, were decreased in expression in Δ7ECF but affected little if at all in ΔMWX. Consistent with this observation, Δ7ECF (but not ∆MWX) showed reduced biofilm formation. Extending previous observations, we also note that ∆MWX is sensitive to a variety of antibiotics and Δ7ECF is either as sensitive as, or slightly more sensitive than, the ΔMWX strain to these stressors. These findings emphasize the overlapping nature of the seven ECF s factor regulons in B. subtilis, confirm that three of these (sigma M, W or X) play the dominant role in conferring intrinsic resistance to antibiotics, and provide initial insights into the roles of the remaining ECF sigma factors. Overall design: Strains WT vs. ΔMWX, WT vs. Δ7ECF, Δ7ECF vs. ΔMWX. Each experiment was conducted three times using three independent total RNA preparations (biological triplicates). For each paried comparison, one sample was was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647. For each comparison, one replicate was performed with dyewap with the same RNA preparation.