Hcp and VgrG1 are secreted components of theHelicobacter hepaticustype VI secretion system and VgrG1 increases the bacterial colitogenic potential

Abstract

SummaryThe enterohepatic Epsilonproteobacterium Helicobacter hepaticus persistently colonizes the intestine of mice and causes chronic inflammatory symptoms in susceptible mouse strains. The bacterial factors causing intestinal inflammation are poorly characterized. A large genomic pathogenicity island, HHGI1, which encodes components of a type VI secretion system (T6SS), was previously shown to contribute to the colitogenic potential of H. hepaticus. We have now characterized the T6SS components Hcp, VgrG1, VgrG2 and VgrG3, encoded on HHGI1, including the potential impact of the T6SS on intestinal inflammation in a mouse T-cell transfer model. The H. hepaticus T6SS components were expressed during the infection and secreted in a T6SS-dependent manner, when the bacteria were cultured either in the presence or in the absence of mouse intestinal epithelial cells. Mutants deficient in VgrG1 displayed a significantly lower colitogenic potential in Tcell-transferred C57BL/6 Rag2 -/-mice, despite an unaltered ability to colonize mice persistently. Intestinal microbiota analyses demonstrated only minor changes in mice infected with wild-type H. hepaticus as compared with mice infected with VgrG1-deficient isogenic bacteria. In addition, competitive assays between both wild-type and T6SS-deficient H. hepaticus, and between wildtype H. hepaticus and Campylobacter jejuni or Enterobacteriaceae species did not show an effect of the T6SS on interbacterial competitiveness. Therefore, we suggest that microbiota alterations did not play a major role in the changes of proinflammatory potential mediated by the T6SS. Cellular innate pro-inflammatory responses were increased by the secreted T6SS proteins VgrG1 and VgrG2. We therefore concluded that the type VI secretion component VgrG1 can modulate and specifically exacerbate the innate proinflammatory effect of the chronic H. hepaticus infection.

Intestinal Microbiota Composition of Interleukin-10 Deficient C57BL/6J Mice and Susceptibility to Helicobacter hepaticus-Induced Colitis

“…Furthermore, the strain 3B1 used at MHH caused robust inflammation comparable with results from other laboratories in different (e.g. T cell transfer) models of H. hepaticus -induced colitis [17]. We therefore hypothesized that differences of the intestinal microbiota composition between C57BL/6 Il10 −/− mice kept at the two different facilities might be responsible for these observed major differences of susceptibility to H. hepaticus -induced pathology.…”

“…The H. hepaticus genome [15] contains a gene cluster encoding a cytolethal distending toxin (CDT), a homolog of the Campylobacter jejuni CDT, which causes cell cycle arrest, chromatin fragmentation, and apoptosis [16]. It also includes a pathogenicity island HHGI1, which encodes a functional type VI secretion system [17], [18]. Strains carrying the island had a higher potential to induce hepatitis in A/JCr mice [19].…”

Role of Energy Sensor TlpD of Helicobacter pylori in Gerbil Colonization and Genome Analyses after Adaptation in the Gerbil

“…Densitometry of specific Western blot bands was performed using ImageJ software (http://rsbweb.nih.gov/ij/), and intensity values were normalized against the constitutive signal generated by anti-H. pylori antiserum. RNA from gerbil tissue was prepared for quantitative PCR (qPCR) as previously described for mouse tissue (35). qPCR was performed as described previously (35,36), using oligonucleotide primers directed against gerbil GAPDH (glyceraldehyde-3-phosphate dehydrogenase), gerbil IL-10, gerbil gamma interferon (IFN-␥) (37), gerbil IL-6 (38), and gerbil inducible nitric oxide synthase (iNOS) (39).…”

Screening for Inhibition of Vibrio cholerae VipA-VipB Interaction Identifies Small-Molecule Compounds Active against Type VI Secretion

“…The contribution of T6SS in this regard may be dependent on its central role in interbacterial competition, since this should be a very important virulence mechanism in complex bacterial communities, such as the gastrointestinal tract (41). In addition, there is evidence that T6SS targets the intestinal epithelium and thereby modulates the inflammatory response such that bacterial survival will be favored (33). Thus, inhibition of T6SS may adversely affect the virulence of gastrointestinal pathogens by impairing their interbacterial competitiveness as well as their ability to modulate the local immune response.…”

“…Thus, the coordinated functions of VipA, VipB, and ClpV are needed for active secretion. In most T6SSs, hemolysin-coregulated protein (Hcp) and valine-glycine repeat protein G (VgrG) are exported by the secretion machinery, but most likely numerous other proteins are secreted as well (30)(31)(32)(33)(34)(35)(36). The well-characterized Hcp protein is not only secreted but also suggested to be a chaperone that binds to cognate T6SS effector molecules, and it forms a hexameric pore that is required for secretion of diverse effectors encompassing several enzymatic classes (37)(38)(39).…”