Overexpression of the pleiotropic regulator CodY decreases sporulation, attachment and pellicle formation in Bacillus anthracis

CodY, a global transcriptional regulator, primarily functions as a nutrient and energy sensor. It is activated by metabolic effectors like BCAA and GTP. In low G + C Gram positive bacteria, it facilitates coupling of changes in the cellular metabolite pool with those required in the transcriptome of the cell. This pleiotropic regulator controls the expression of a vast number of genes as the cell transits from exponential to the stationary phase. Earlier studies have shown that CodY is required for the virulence of Bacillus anthracis. We sought to investigate the effect of its overexpression on the physiology of B. anthracis. In our study, we found that cellular CodY levels were unchanged during this phase-transition. Expression of endogenous CodY remained the same in different nutrient limiting conditions. Immunoblotting studies revealed CodY presence in the whole spore lysate of B. anthracis indicating it to be a component of the spore proteome. We could also detect CodY in the secretome of B. anthracis. Further, CodY was overexpressed in B. anthracis Sterne strain and this led to a 100-fold decrease in the sporulation titer and a 2.5-fold decrease in the in vitro attachment ability of the bacteria. We also observed a decrease in the pellicle formation by CodY overexpressed strain when compared to wildtype bacilli. The CodY overexpressed strain showed chaining phenotype during growth in liquid media and pellicle.