A class IV adenylate cyclase CyaB is required for capsule polysaccharide production and biofilm formation inVibrio parahaemolyticus

Abstract

CRP (cyclic AMP receptor protein), encoded by crp, is a global transcription factor that is activated by cyclic AMP (cAMP), a second messenger synthesized from ATP by adenylate cyclase, cyaA in Escherichia coli. In this species, growth on the preferred carbon source glucose, results in low levels of cAMP, and inactive CRP. When glucose is absent, cAMP levels are high which activates CRP to regulate 100s of genes. To investigate the role of CRP in Vibrio parahaemolyticus, we constructed non-polar deletions of the crp and cyaA homologs. The Dcrp mutant did not grow in minimal media supplemented with non-glucose carbon sources, however the DcyaA mutant grew similar to wild type suggesting the presence of an additional adenylate cyclase. Bioinformatics analysis identified a homolog of a class IV adenylate cyclase, CyaB present in V. parahaemolyticus. Both V. parahaemolyticus cyaA and cyaB functionally complemented an E. coli DcyaA mutant indicating adenylate cyclase activity. Growth pattern analysis showed that cyaA is controlled by CRP, but cyaB is not. cAMP-CRP was required for motility with the Dcrp and DcyaB/DcyaA mutants having a defect in swimming and swarming motility. The DcyaA mutant produces capsule polysaccharide (CPS) and biofilm similar to wild type. However, the DcyaB mutant was defective in CPS production and biofilm formation, similar to the Dcrp and DcyaB/DcyaA mutants, indicating that CyaB, but not CyaA, is required for these phenotypes. Phylogenetic analysis of cyaB showed broad distribution within the Vibrionaceae family and was acquired by horizontal gene transfer in several species.