Spx (YjbD), a negative effector of competence in Bacillus subtilis, enhances ClpC-MecA-ComK interaction

ComK, a key transcriptional regulator in the development of competence in Bacillus subtilis, is required for its own transcription as well as that of the late competence genes encoding proteins involved in DNA uptake. ComK is sequestered in a complex with ClpC and MecA until a peptide, ComS, accumulates in cells. ComS releases ComK from the inhibitory complex, thus allowing ComK to carry out its function as a transcriptional activator. Spx (formerly YjbD), a negative effector of competence, accumulates in clpP mutants. High concentrations of Spx may be responsible for the inability of clpP mutants to become competent because spx mutations are able to restore competence in the clpP mutant. In this paper, we showed, based on in vitro experiments, that Spx forms a quaternary complex with ClpC, MecA and ComK and enhances ComK binding to ClpC-MecA. Two ComS alanine substitution mutants (I13A and W43A), previously shown to be defective in vivo, were less efficient in releasing ComK from ClpC-MecA. The I13A mutant with a weaker binding affinity to MecA was inefficient in releasing ComK regardless of whether Spx was present. In contrast, the defect of the W43A mutant in dissociating ComK was more readily observed in the presence of Spx. Spx is a highly conserved protein among Gram-positive bacteria, in which it may function closely with the protease adaptor protein, MecA.