Transcription and translation of the sigG gene is tuned for proper execution of the switch from early to late gene expression in the developing Bacillus subtilis spore

Abstract

A cascade of alternative sigma factors directs developmental gene expression during spore formation by the bacterium Bacillus subtilis. As the spore develops, a tightly regulated switch occurs in which the early-acting sigma factor σF is replaced by the late-acting sigma factor σG. The gene encoding σG (sigG) is transcribed by σF and by σG itself in an autoregulatory loop; yet σG activity is not detected until σF-dependent gene expression is complete. This separation in σF and σG activities has been suggested to be due at least in part to a poorly understood intercellular checkpoint pathway that delays sigG expression by σF. Here we report the results of a careful examination of sigG expression during sporulation. Unexpectedly, our findings argue against the existence of a regulatory mechanism to delay sigG transcription by σF and instead support a model in which sigG is transcribed by σF with normal timing, but at levels that are very low. This low-level expression of sigG is the consequence of several intrinsic features of the sigG regulatory and coding sequence—promoter spacing, secondary structure potential of the mRNA, and start codon identity—that dampen its transcription and translation. Especially notable is the presence of a conserved hairpin in the 5’ leader sequence of the sigG mRNA that occludes the ribosome-binding site, reducing translation by up to 4-fold. Finally, we demonstrate that misexpression of sigG from regulatory and coding sequences lacking these features triggers premature σG activity in the forespore during sporulation, as well as inappropriate σG activity during vegetative growth. Altogether, these data indicate that transcription and translation of the sigG gene is tuned to prevent vegetative expression of σG and to ensure the precise timing of the switch from σF to σG in the developing spore.

From Root to Tips: Sporulation Evolution and Specialization inBacillus subtilisand the Intestinal PathogenClostridioides difficile

“…2016). Because of the autoregulatory nature of σ G , several mechanisms concur with inhibition by CsfB to prevent its premature activity (Mearls et al. 2018).…”

A genetic selection reveals functional metastable structures embedded in a toxin-encoding mRNA

“…Regulation of translation initiation via SD-sequestration is an old theme that initially started with the study of bacteriophage genes (de Smit and van Duin, 1990) and ribosomal proteins (for a review see Duval et al, 2015). More recently, its impact on other bacterial genes such as sigma factors (Mearls et al, 2018) and translational riboswitches (Rinaldi et al, 2016) has been reported.…”

Channels modestly impact compartment‐specific ATP levels during Bacillus subtilis sporulation and a rise in the mother cell ATP level is not necessary for Pro‐σK cleavage

“…Why then does σ G fail to become fully active in spoIIQ and spoIIIA mutants? The transmembrane segment of SpoIIQ regulates expression of the gene for the anti-σ G factor CsfB (Flanagan et al, 2016) and several features of the sigG promoter, mRNA leader sequence, and start codon dampen expression (Mearls et al, 2018), but these findings do not explain a more general requirement of SpoIIQ and…”

“…Activity of σ F initiates a signaling pathway that leads to proteolytic activation of Pro‐σ E in the MC (Konovalova et al ., 2014). During the ensuing engulfment process, σ F activity produces SpoIIQ, which inserts in the inner FS membrane, and σ E activity produces SpoIIIA proteins, which insert in the engulfing MC membrane (Figure 1, left side), forming channels between the two compartments that are required for full activity of σ G in the FS and maintenance of FS integrity (Blaylock et al ., 2004; Doan et al ., 2005; 2009; Camp and Losick, 2008; 2009; Meisner et al ., 2008; Mearls et al ., 2018). The activation of σ G coincides with completion of engulfment and initiates a signaling pathway that leads to proteolytic cleavage of Pro‐σ K to active σ K in the MC (Kroos and Akiyama, 2013; Konovalova et al ., 2014).…”

“…in the engulfing MC membrane (Figure 1, left side), forming channels between the two compartments that are required for full activity of σ G in the FS and maintenance of FS integrity (Blaylock et al, 2004;Doan et al, 2005;Camp and Losick, 2008;Meisner et al, 2008;Mearls et al, 2018). The activation of σ G coincides with completion of engulfment and initiates a signaling pathway that leads to proteolytic cleavage of Pro-σ K to active σ K in the MC (Kroos and Akiyama, 2013;Konovalova et al, 2014).…”