Intestinal Adherence of Vibrio cholerae Involves a Coordinated Interaction between Colonization Factor GbpA and Mucin

Abstract

The chitin-binding protein GbpA of Vibrio cholerae has been recently described as a common adherence factor for chitin and intestinal surface. Using an isogenic in-frame gbpA deletion mutant, we first show that V. cholerae O1 El Tor interacts with mouse intestinal mucus quickly, using GbpA in a specific manner. The gbpA mutant strain showed a significant decrease in intestinal adherence, leading to less colonization and fluid accumulation in a mouse in vivo model. Purified recombinant GbpA (rGbpA) specifically bound to N-acetyl-D-glucosamine residues of intestinal mucin in a dose-dependent, saturable manner with a dissociation constant of 11.2 M. Histopathology results from infected mouse intestine indicated that GbpA binding resulted in a time-dependent increase in mucus secretion. We found that rGbpA increased the production of intestinal secretory mucins (MUC2, MUC3, and MUC5AC) in HT-29 cells through upregulation of corresponding genes. The upregulation of MUC2 and MUC5AC genes was dependent on NF-B nuclear translocation. Interestingly, mucin could also increase GbpA expression in V. cholerae in a dose-dependent manner. Thus, we propose that there is a coordinated interaction between GbpA and mucin to upregulate each other in a cooperative manner, leading to increased levels of expression of both of these interactive factors and ultimately allowing successful intestinal colonization and pathogenesis by V. cholerae.Vibrio cholerae is the causative agent of the potentially lethal disease cholera. V. cholerae strains belonging to serogroups O1 and O139 are mainly responsible for cholera epidemics, while strains of other serogroups may cause sporadic outbreaks of the disease. Although the O139 strain has evolved recently, V. cholerae O1 biotype El Tor strains have still been responsible for most of the epidemics in recent years (20,26). In order to cause the disease, V. cholerae must adhere to the intestinal mucus barrier (52). The ability of V. cholerae to adhere to animal cells has been studied before (26,42), and various adherence factors have been proposed, including the virulence-associated toxin-coregulated pilus (5), outer membrane proteins (26, 42), and lipopolysaccharide (LPS) (11). Attachment of V. cholerae to abiotic surfaces has also been recently described (50). However, there is still no information about the factor(s) responsible for initial adherence of the bacteria to the intestine and whether the host plays any role in aiding the colonization of the intestine by the bacteria.Vibrios are marine organisms that adhere to chitin in the environment (12, 33) and utilize chitin as the sole source of nitrogen and carbon by using a family of glycosyl hydrolases, called chitinases (21). Genome analysis of V. cholerae O1 El Tor has revealed the presence of seven such chitinase genes (7), some of which have been characterized (27,37). One of these genes is the putative chitinase gene with locus number VCA0811, the product of which has been recently identified as a common adhesion molecule for both chitinou...

Vibrio cholerae GbpA elicits necrotic cell death in intestinal cells

“…Previously, it was demonstrated that the induction of mucin genes (Bhowmick et al, 2008) and secretion of pro-inflammatory cytokine IL-8 in intestinal cells by GbpA were regulated by NF-kB. Further influence of ROS on NF-kB activation induced by various environmental stimuli is reported.…”

“…The GbpA was found to increase the production of intestinal secretory mucins (MUC2, MUC3 and MUC5AC) in HT-29 cells through the activation of corresponding genes (Bhowmick et al, 2008). The activation of mucin genes (MUC2 and MUC5AC) was found to be dependent on nuclear translocation of NF-kB (Bhowmick et al, 2008). Therefore, GbpA can be considered as an effector molecule capable of eliciting host response.…”

“…In spite of being secretory in nature, GbpA has been found to be the factor bridging V. cholerae and mucin in the host (Bhowmick et al, 2008). The GbpA was found to increase the production of intestinal secretory mucins (MUC2, MUC3 and MUC5AC) in HT-29 cells through the activation of corresponding genes (Bhowmick et al, 2008). The activation of mucin genes (MUC2 and MUC5AC) was found to be dependent on nuclear translocation of NF-kB (Bhowmick et al, 2008).…”

“…It was identified as an adhesion factor for surfaces with N-acetylglucosamine (GlcNAc) and its oligomers (Meibom et al, 2004). In spite of being secretory in nature, GbpA has been found to be the factor bridging V. cholerae and mucin in the host (Bhowmick et al, 2008). The GbpA was found to increase the production of intestinal secretory mucins (MUC2, MUC3 and MUC5AC) in HT-29 cells through the activation of corresponding genes (Bhowmick et al, 2008).…”

“…This initial attachment is the key step in pathogenesis of V. cholerae. N-Acetylglucosamine-binding protein A (GbpA), the gene product of locus VCA0811, serves an important role as a common adhesin for chitinous surfaces as well as intestinal epithelium (Kirn et al, 2005;Bhowmick et al, 2008;Wong et al, 2012). It was identified as an adhesion factor for surfaces with N-acetylglucosamine (GlcNAc) and its oligomers (Meibom et al, 2004).…”