CHARACTERIZATION OF SPOOA HOMOLOGS IN DIVERSE BACILLUS AND CLOSTRIDIUM SPECIES IDENTIFIES A PROBABLE DNA-BINDING DOMAIN

SpoOA is a phosphorylation-activated transcription factor of Bacillus subtilis. It is a member of the response regulator superfamily of bact erial signal transduction proteins and controls many of the changes in gene expression that occur during the transition into stationary phas e and during the initiation of sporulation. To identify the domains of SpoOA most critical for determining its structural and functional fea tures, presumptive homologues of the spoOA gene were characterized in a collection of eight Bacillus species and six Clostridium species rep resenting phylogenetically diverse members of these genera. An alignme nt of the partial or complete DNA sequences of these homologues reveal ed three regions of especially high conservation in the effector domai n. We speculate that the most highly conserved of these corresponds to the recognition helix of a putative helix-turn-helix motif, and, ther efore, represents the actual DNA-contacting surface of the protein. In the case of homologues identified in Bacillus anthracis and Clostridi um acetobutylicum and retrieved by polymerase chain reaction amplifica tion, we confirmed by gene-disruption analysis that the homologue actu ally is required for initiation of sporulation. Apparent homologues of the B. subtilis spoIVB gene were also discovered immediately upstream from the spoOA homologues in all Bacillus and Clostridium species exa mined. The discovery of homologues of B. subtilis sporulation genes in these diverse species implies that the gene products required for spe cifying pathways of sporulation-specific gene activation and for deter mining key morphogenetic changes may be highly conserved and suggests that an approach similar to that undertaken here might be used as a ge neral strategy to retrieve and compare their gene sequences. Exhaustiv e efforts to detect a spoOA-like gene in non-endospore formers, includ ing close relatives of Bacillus such as Listeria and Staphylococcus, w ere uniformly unsuccessful, suggesting that regulation of gene activit y during the transition into stationary phase mediated by SpoOA-like p roteins may be exclusive to the endospore-forming bacteria.