Bacillus Subtilis Extracytoplasmic Function Sigma Factors Control Antibiotic Production And Resistance

Bacillus subtilis, the model organism for Gram-positive bacteria, expresses seven extracytoplasmic function (ECF) [sigma] factors (called [sigma]M, [sigma]W, [sigma]X, [sigma]Y, [sigma]V, [sigma]Z and [sigma]YlaC). [sigma]M, [sigma]W, [sigma]X, and [sigma]V are well-characterized and mediate cell envelope stress responses. The functions of [sigma]Y, [sigma]Z and [sigma]YlaC remain largely unknown. One challenge in understanding the regulatory roles of ECF [sigma] factor is that they display significant redundancy in their regulons. In this study, we investigate the regulation of B. subtilis antibiotic production and resistance by ECF [sigma] factors using both global analytical techniques (cDNA microarrays, phenotypic arrays, and transposon mutagenesis) and classical genetic and physiological tests. This work began with the observation that a triple sigMWX mutant (strain [DELTA]MWX) lost its ability to inhibit the growth of other bacilli strains. We present evidence that deletions of [sigma]X and [sigma]M underlie this phenotype and that in wild type cells these ECF [sigma] factors activate the expression of a transcription factor Abh. Abh stimulates the production of a peptide antibiotic called sublancin, which is able to inhibit the growth of other bacilli. We also compared the transcriptomic profiles and phenotypic traits of strains lacking these three [sigma] factors ([DELTA]MWX) and all 7 ECF [sigma] factors ([DELTA]7ECF) with a wild type strain. Deletion of all 7 ECF [sigma] factors affects the i transcriptions of over 80 genes, most of which are regulated by [sigma]M, [sigma]W, or [sigma]X. The [DELTA]MWX and [DELTA]7ECF strains are more sensitive to several cell envelope disrupting compounds when compared to wild type including two [beta]-lactam antibiotics (aztreonam and cefuroxime). Finally, we investigated the ECF [sigma] factor-dependent genetic and biochemical mechanisms that mediate resistance to cefuroxime. [sigma]M is the major determinant in cefuroxime resistance with [sigma]X playing a smaller role. These [sigma] factors regulate at least three pathways involving the regulatory proteins Abh and Spx, and a cyclic-di-AMP synthase DisA. Collectively, the data in this work suggests that ECF [sigma] factors regulate an intricate regulatory network that contributes to both antibiotic production and resistance in B. subtilis. i