A tyrosine phosphoregulatory system controls exopolysaccharide biosynthesis and biofilm formation in Vibrio cholerae

Production of an extracellular matrix is essential for biofilm formation, as this matrix both secures and protects the cells it encases. Mechanisms underlying production and assembly of matrices are poorly understood.Vibrio cholerae, relies heavily on biofilm formation for survival, infectivity, and transmission. Biofilm formation requiresVibriopolysaccharide (VPS), which is produced byvpsgene-products, yet the function of these products remains unknown. Here, we demonstrate that thevpsgene-productsvpsOandvpsUencode respectively for a tyrosine kinase and a cognate tyrosine phosphatase. Collectively, VpsO and VpsU act as a tyrosine phosphoregulatory system to modulate VPS production. We present structures of VpsU and the kinase domain of VpsO, and we report observed autocatalytic tyrosine phosphorylation of the VpsO C-terminal tail. The position and amount of tyrosine phosphorylation in the VpsO C-terminal tail represses VPS production and biofilm formation through a mechanism involving the modulation of VpsO oligomerization. We found that tyrosine phosphorylation enhances stability of VpsO. Regulation of VpsO phosphorylation by the phosphatase VpsU is vital for maintaining native VPS levels. This study provides new insights into the mechanism and regulation of VPS production and establishes general principles of biofilm matrix production and its inhibition. Author summary The biofilm life style protects microbes from a plethora of harm, to increase their survival and pathogenicity. Exopolysaccharides are the essential glue of the microbial biofilm matrix, and loss of this glue negates biofilm formation and renders cells more sensitive to antimicrobial agents. Here, we show that a tyrosine phosphoregulatory system controls the biosynthesis and abundance ofVibrioexopolysaccharide (VPS), an essential biofilm component of the pathogenVibrio cholerae. The phosphorylation state of the tyrosine autokinase VpsO, mediated by the tyrosine phosphatase VpsU, directly modulates VPS production and also affects the kinase's own degradation, to regulate VPS production. This study provides new insights into the mechanisms ofV.choleraebiofilm formation and consequently ways to combat pathogens more broadly, due conservation of tyrosine phosphoregulatory systems among exopolysaccharide producing bacteria.