Genome‐wide analysis of in vivo CcpA binding with and without its key co‐factor HPr in the major human pathogen group A Streptococcus

Catabolite control protein A (CcpA) is a master regulator of carbon source utilization and contributes to the virulence of numerous medically important Gram‐positive bacteria. Most functional assessments of CcpA, including interaction with its key co‐factor HPr, have been performed in nonpathogenic bacteria. In this study we aimed to identify the in vivo DNA binding profile of CcpA and assess the extent to which HPr is required for CcpA‐mediated regulation and DNA binding in the major human pathogen group A Streptococcus (GAS). Using a combination RNAseq/ChIP‐seq approach, we found that CcpA affects transcript levels of 514 of 1667 GAS genes (31%) whereas direct DNA binding was identified for 105 GAS genes. Three of the directly regulated genes encode the key GAS virulence factors Streptolysin S, PrtS (IL‐8 degrading proteinase), and SpeB (cysteine protease). Mutating CcpA Val301 to Ala (strain 2221‐CcpA‐V301A) abolished interaction between CcpA and HPr and impacted the transcript levels of 205 genes (40%) in the total CcpA regulon. By ChIP‐seq analysis, CcpAV301A bound to DNA from 74% of genes bound by wild‐type CcpA, but generally with lower affinity. These data delineate the direct CcpA regulon and clarify the HPr‐dependent and independent activities of CcpA in a key pathogenic bacterium.