Bacillus subtilis degR, a positive regulator of the production of degradative enzymes, is negatively regulated by the competence transcription factor ComK which is overproduced in mecA null mutants. We used transposon Tn10 to search for a mutation that reduced the repression level of degR caused by a mecA mutation. A new gene exerting positive regulation on comK was obtained and designated med (suppressor of mecA effect on degR). Sequence determination, Northern analysis, and primer extension analyses revealed that the med gene contained an open reading frame (ORF) composed of 317 codons and was transcribed into an approximately 1,250-nucleotide mRNA together with its short downstream gene. The expression of comK is positively regulated by factors such as ComK itself, ComS (SrfA)-MecA, DegU, SinR, and AbrB. Quantitative analyses using comK -lacZ, srfA-lacZ, degU -lacZ, and sinR -lacZ fusions showed that disruption of med caused a significant decrease in comK expression in both mecA ؉ and mecA strains, while expression of srfA, sinR, and degU was not affected by the mutation. An epistatic analysis revealed that overproduction of ComK resulted in alteration of med expression, suggesting a regulatory loop between comK and med. Several possible mechanisms for positive regulation of comK by Med are discussed.

DegR is a positive regulator for degradative enzyme synthesis in Bacillus subtilis. The degR gene is transcribed by RNA polymerase containing c c D , and the level of its expression is low in a mecA-deficient mutant. In a search for suppressors of the mecA effect through mini-Tn10 transposon mutagenesis, a lacR mutation designated lacR288 was discovered. The B. subtilis lacR gene encodes the repressor for lacA which specifies L L-galactosidase, and therefore, inactivation of the lacR gene results in overproduction of the enzyme. In the lacR288 mutant, however, the expression of lacA was at a negligible level, indicating that the repressor activity was not destroyed by the mutation. The putative gene product of the lacR288-containing gene is a 288-amino acid protein lacking the C-terminal 42 amino acids of intact LacR and carries no extra amino acids derived from the transposon sequence. The suppression by lacR288 of the decreased degR expression in the mecA background was found to be caused by an increase in the c c D level as shown by Western blot analysis. Furthermore, the increase was due to post-transcriptional regulation of sigD, the gene encoding c c D , as revealed by using both transcriptional and translational sigD-lacZ fusions. The lacR288 mutation had no effect on the stability of the c c D protein. Based on these results we conclude that the lacR288 mutation stimulates sigD expression at the translational level.z 1999 Federation of European Biochemical Societies.