A complex of YlbF, YmcA and YaaT regulates sporulation, competence and biofilm formation by accelerating the phosphorylation of Spo0A.

Bacillus subtilis has adopted a bet-hedging strategy to ensure survival in changing environments. From a clonal population, numerous sub-populations can emerge, expressing different sets of genes that govern the developmental processes of sporulation, competence and biofilm formation. The master transcriptional regulator Spo0A controls the entry into all three fates and the production of the phosphorylated active form of Spo0A is precisely regulated via a phosphorelay, involving at least four proteins. Two proteins, YmcA and YlbF were previously shown to play an unidentified role in the regulation of biofilm formation, and in addition, YlbF was shown to regulate competence and sporulation. Using an unbiased proteomics screen, we demonstrate that YmcA and YlbF interact with a third protein, YaaT to form a tripartite complex. We show that all three proteins are required for proper establishment of the three above-mentioned developmental states. We show that the complex regulates the activity of Spo0A in vivo and, using in vitro reconstitution experiments, determine that they stimulate the phosphorelay, probably by interacting with Spo0F and Spo0B. We propose that the YmcA-YlbF-YaaT ternary complex is required to increase Spo0A~P levels above the thresholds needed to induce development.

MecA dampens transitions to spore, biofilm exopolysaccharide and competence expression by two different mechanisms.

The adapter protein MecA targets the transcription factor ComK for degradation by the ClpC/ClpP proteolytic complex, thereby negatively regulating competence in Bacillus subtilis. Here we show that MecA also decreases the frequency of transitions to the sporulation pathway as well as the expression of eps, which encodes synthesis of the biofilm matrix exopolysaccharide. We present genetic and biophysical evidence that MecA downregulates eps expression and spore formation by directly interacting with Spo0A. MecA does not target Spo0A for degradation, and apparently does not prevent the phosphorylation of Spo0A. We propose that it inhibits the transcriptional activity of Spo0A∼P by direct binding. Thus, in its interaction with Spo0A, MecA differs from its role in the regulation of competence where it targets ComK for degradation. MecA acts as a general buffering protein for development, acting by two distinct mechanisms to regulate inappropriate transitions to energy-intensive pathways.