A Multi-Serotype Approach Clarifies the Catabolite Control Protein A Regulon in the Major Human Pathogen Group A Streptococcus

Abstract

Catabolite control protein A (CcpA) is a highly conserved, master regulator of carbon source utilization in gram-positive bacteria, but the CcpA regulon remains ill-defined. In this study we aimed to clarify the CcpA regulon by determining the impact of CcpA-inactivation on the virulence and transcriptome of three distinct serotypes of the major human pathogen Group A Streptococcus (GAS). CcpA-inactivation significantly decreased GAS virulence in a broad array of animal challenge models consistent with the idea that CcpA is critical to gram-positive bacterial pathogenesis. Via comparative transcriptomics, we established that the GAS CcpA core regulon is enriched for highly conserved CcpA binding motifs (i.e. cre sites). Conversely, strain-specific differences in the CcpA transcriptome seems to consist primarily of affected secondary networks. Refinement of cre site composition via analysis of the core regulon facilitated development of a modified cre consensus that shows promise for improved prediction of CcpA targets in other medically relevant gram-positive pathogens.

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

“…were CcpA repressed) compared to genes (153) whose transcript levels were lower in 2221∆ccpA (i.e. were CcpA activated), which is in accordance with previous observations (Carvalho et al, 2011, DebRoy et al, 2016, Kietzman & Caparon, 2011, Shelburne et al, 2008. The transcript levels of twelve virulence factor encoding genes were impacted by CcpA inactivation including speB, speA2, sic, spd, ska, grab, the sag operon, the nga-slo operon, and prtS (Table 2).…”

“…Relative to our previously described in silico derived cre ( Figure 6E, (DebRoy et al, 2016)), the major difference for our ChIPseq derived cre was a lack of predominance of T at positions 1 and 2 ( Figure 6B). We compared the cre sites that we found using the ChIP method in this study to those that we had previously predicted in MGAS2221 using in silico analysis of RNAseq data (DebRoy et al, 2016). Both studies were conducted on mid-exponential cells grown in THY.…”

“…The LacI-GalR family transcriptional regulator catabolite control protein (CcpA) is a key mediator of CCR in many Gram-positive bacteria (Henkin et al, 1991, Titgemeyer & Hillen, 2002, Warner & Lolkema, 2003. CcpA inactivation impacts ~15-20% of the transcriptome of a broad array of Gram-positive bacteria with the majority of impacted genes encoding proteins involved in carbohydrate and nitrogen utilization (Antunes et al, 2012, DebRoy et al, 2016, Seidl et al, 2009, Zeng et al, 2013. Importantly, CcpA also affects the transcript levels of genes encoding known and putative virulence factors in human Gram-positive pathogens ranging from streptococci to Clostridia (Iyer et al, 2005, Mertins et al, 2007, Seidl et al, 2008a, Varga et al, 2004.…”

“…Although the pseudo-palindromic cre sequence seems to be the predominate site of CcpA-DNA interaction in Bacillus species, there have been suggestions that CcpA may bind other DNA sites. Using in silico approaches, cre sites have been predicted in only a small percentage of genes whose transcript levels are significantly altered by CcpA inactivation (Carvalho et al, 2011, DebRoy et al, 2016.…”

Integration of Biological Networks for Acidithiobacillus thiooxidans Describes a Modular Gene Regulatory Organization of Bioleaching Pathways

“…The generation of the Affinity transcriptional regulatory network was based on previously reported protocols DebRoy et al, 2016). Briefly, candidate transcription factors present in the genome of A. thiooxidans Licanantay were identified using the following protocol: First, using the results of genome annotation, each candidate must have at least a Helix-Turn-Helix domain, previously identified using HMMER software with Pfam database.…”