A Negative Feedback Loop That Limits the Ectopic Activation of a Cell Type–Specific Sporulation Sigma Factor of Bacillus subtilis

Two highly similar RNA polymerase sigma subunits, σF and σG, govern the early and late phases of forespore-specific gene expression during spore differentiation in Bacillus subtilis. σF drives synthesis of σG but the latter only becomes active once engulfment of the forespore by the mother cell is completed, its levels rising quickly due to a positive feedback loop. The mechanisms that prevent premature or ectopic activation of σG while discriminating between σF and σG in the forespore are not fully comprehended. Here, we report that the substitution of an asparagine by a glutamic acid at position 45 of σG (N45E) strongly reduced binding by a previously characterized anti-sigma factor, CsfB (also known as Gin), in vitro, and increased the activity of σG in vivo. The N45E mutation caused the appearance of a sub-population of pre-divisional cells with strong activity of σG. CsfB is normally produced in the forespore, under σF control, but sigGN45E mutant cells also expressed csfB and did so in a σG-dependent manner, autonomously from σF. Thus, a negative feedback loop involving CsfB counteracts the positive feedback loop resulting from ectopic σG activity. N45 is invariant in the homologous position of σG orthologues, whereas its functional equivalent in σF proteins, E39, is highly conserved. While CsfB does not bind to wild-type σF, a E39N substitution in σF resulted in efficient binding of CsfB to σF. Moreover, under certain conditions, the E39N alteration strongly restrains the activity of σF in vivo, in a csfB-dependent manner, and the efficiency of sporulation. Therefore, a single amino residue, N45/E39, is sufficient for the ability of CsfB to discriminate between the two forespore-specific sigma factors in B. subtilis

Structural basis of mitochondrial transcription.

The mitochondrial genome is transcribed by a single-subunit DNA-dependent RNA polymerase (mtRNAP) and its auxiliary factors. Structural studies have elucidated how mtRNAP cooperates with its dedicated transcription factors to direct RNA synthesis: initiation factors TFAM and TFB2M assist in promoter-DNA binding and opening by mtRNAP while the elongation factor TEFM increases polymerase processivity to the levels required for synthesis of long polycistronic mtRNA transcripts. Here, we review the emerging body of structural and functional studies of human mitochondrial transcription, provide a molecular movie that can be used for teaching purposes and discuss the open questions to guide future directions of investigation