Regulation and Characterization of a Newly Deduced Cell Wall Hydrolase Gene ( cwlJ ) Which Affects Germination of Bacillus subtilis Spores

Abstract

ABSTRACT The predicted amino acid sequence of Bacillus subtilis ycbQ (renamed cwlJ ) exhibits high similarity to those of the deduced C-terminal catalytic domain of SleBs, the specific cortex-hydrolyzing enzyme of B. cereus and the deduced one of B. subtilis . We constructed a cwlJ::lacZ fusion in the B. subtilis chromosome. The β-galactosidase activity and results of Northern hybridization and primer extension analyses of the cwlJ gene indicated that it is transcribed by Eς E RNA polymerase. cwlJ -deficient spores responded to both l -alanine and AGFK, the A 580 values of spore suspensions decreased more slowly than in the case of the wild-type strain, and the mutant spores released less dipicolinic acid than did those of the wild-type strain during germination. However, the mutant spores released only slightly less hexosamine than did the wild-type spores. In contrast, B. subtilis sleB spores did not release hexosamine at a significant level. While cwlJ and sleB spores were able to germinate, CJSB ( cwlJ sleB ) spores could not germinate but exhibited initial germination reactions, e.g., partial decrease in A 580 and slow release of dipicolinic acid. CJSB spores became slightly gray after 6 h in the germinant, but their refractility was much greater than that of sleB mutant spores. The roles of the sleB and cwlJ mutations in germination and spore maturation are also discussed.