An amino-terminal threonine/serine motif is necessary for activity of the Crp/Fnr homolog, MrpC and for Myxococcus xanthus developmental robustness.

The Crp/Fnr family of transcriptional regulators play central roles in transcriptional control of diverse physiological responses, and are activated by a surprising diversity of mechanisms. MrpC is a Crp/Fnr homolog that controls the Myxococcus xanthus developmental program. A long-standing model proposed that MrpC activity is controlled by the Pkn8/Pkn14 serine/threonine kinase cascade, which phosphorylates MrpC on threonine residue(s) located in its extreme amino-terminus. In this study, we demonstrate that a stretch of consecutive threonine and serine residues, T21 T22 S23 S24, is necessary for MrpC activity by promoting efficient DNA binding. Mass spectrometry analysis indicated the TTSS motif is not directly phosphorylated by Pkn14 in vitro but is necessary for efficient Pkn14-dependent phosphorylation on several residues in the remainder of the protein. In an important correction to a long-standing model, we show Pkn8 and Pkn14 kinase activities do not play obvious roles in controlling MrpC activity in wild-type M. xanthus under laboratory conditions. Instead, we propose Pkn14 modulates MrpC DNA binding in response to unknown environmental conditions. Interestingly, substitutions in the TTSS motif caused developmental defects that varied between biological replicates, revealing that MrpC plays a role in promoting a robust developmental phenotype.

MrpC, a CRP/Fnr homolog, functions as a negative autoregulator during the Myxococcus xanthus multicellular developmental program.

MrpC, a member of the CRP/Fnr superfamily of transcriptional regulators, plays a key role in coordination of the multicellular developmental program in Myxococcus xanthus. Previous reports suggest MrpC is subject to complex regulation including activation by an unusual LonD-dependent proteolytic processing event that removes its unique N-terminal peptide, producing the isoform MrpC2. MrpC2 is proposed to positively autoregulate and regulate transcription of hundreds of genes necessary for both the aggregation and sporulation phases of the developmental program. We demonstrate here that mrpC expression bifurcates corresponding to different cell populations within the developmental program. During our analysis of regulatory events controlling this process, we demonstrate that MrpC2 is not an active isoform; rather, the N-terminal peptide is instead essential for MrpC function in vivo. We also demonstrate that MrpC is instead a negative autoregulator and represses its own expression by specifically competing with its enhancer binding protein, MrpB. These results provide an additional rare example of CRP/EBP coordinated regulation, and significantly revise the model for control of the central developmental transcriptional activator of the M. xanthus developmental program.