H-040. Genetic Analysis of the Putative swrA Operon of Bacillus subtilis

J. E. Patrick, D. B. Kearns;
Indiana Univ., Bloomington, IN.

B. subtilis is a Gram-positive soil microbe capable of motility via peritrichous flagella. In liquid environments it swims and on solid surfaces it swarms in dynamic multicellular rafts. Swarming motility is controled by swrA which acts directly or indirectly to increase transcription from the flagellar and chemotaxis operon, fla/che. In laboratory strains, the swrA gene was proposed to be part of a discistronic operon with the downstream gene yvjD, and yvjD was proposed to be necessary for swarming motility at the wild type level. We performed genetic analysis of swrA and yvjD in an undomesticate strain to investigate these claims. In order to investigate the operon structure of swrA and yvjD, transcriptional lacZ fusions to PswrA and to the putative PyvjD were constructed. PyvjD-lacZ produced equivalent Beta-galactosidase activity to PswrA-lacZ fusions, suggesting that yvjD has a promoter separate from that of swrA. Next we sought to determing if the genes had similar functions. There was a recombination defect in strains lacking yvjD. ΔyvjD strains could be transduced with SPP1 phage and could maintain a plasmid extrachromosomally, but failed to incorporate the plasmid into the genome. This defect could be complemented using either PswrA-swrAyvjD or PyvjD-yvjD. Presence or absence of a functinal swrA allele had no effect on recombination. Introduction of PyvjD-yvjD restored the ability of ΔyvjD strains to recombine. Also, Campbell integrants into yvjD could not be excised unless an additional copy of the allele, under either PswrA or PyvjD control, was present. Taken together, these data suggest that yvjD has its own promoter separate from the promoter of swrA, and the promoter is sufficient to restore the recombination phenotype observed in ΔyvjD strains of B. subtilis. We conclude that yvjD is not part of an obligate operon with the upstream gene swrA. This far, all evidence suggests that the functions of swrA and yvjD are unrelated.