Helicobacter pylori cholesteryl glucosides interfere with host membrane phase and affect type IV secretion system function during infection in AGS cells

Abstract

SummaryHelicobacter pylori infection is an aetiological cause of gastric disorders worldwide. H. pylori has been shown to assimilate and convert host cholesterol into cholesteryl glucosides (CGs) by cholesterol-aglucosyltransferase encoded by capJ. Here, we show that CapJ-deficient (DcapJ) H. pylori resulted in greatly reduced type IV secretion system (TFSS)-associated activities, including the hummingbird phenotype of AGS cells, IL-8 production, CagA translocation/phosphorylation and CagA-mediated signalling events. Complementation of the DcapJ mutation with wild type cagJ or by adding CGs-containing lysates or exogenous fluorophore-tagged CGs reversed the mutant phenotypes. We also show that the wild-type but not DcapJ H. pylori recruited raft-associated components to sites of bacterial attachment. Fluorescence recovery after photobleaching (FRAP) analysis of AGS cells treated with fluorescence-tagged cholesterol/CGs revealed that there was a higher proportion of CGs associated with immobile fractions. CGs-associated membranes were also more resistant to a cold detergent extraction. Thus, we propose that CGs synthesized by H. pylori around host-pathogen contact sites partition in detergent-resistant membranes (DRMs), alters lateralphase segregation in membrane and reorganizes membrane architecture. These processes together promote the formation of a functional TFSS and H. pylori infection.

Hijacking and Use of Host Lipids by Intracellular Pathogens

“…Cholesteryl glucosides promote the formation of functional T4SS, and mutants of components of the pathogenicity island have impaired activities of their T4SS that can be restored by complementation. Glucosyl cholesterol synthesized by H. pylori collects around host-pathogen contact sites and promotes the formation of a functional T4SS and, therefore, H. pylori infection (162). …”

Colonize, evade, flourish

“…80 In the absence of Hp0421, H. pylori is more susceptible to phagocytosis and therefore triggers a strong T cell-mediated immune response. 84,85 Conversely, wild-type bacteria are phagocytosed to a limited extent and generate a weak immune response. Further, high concentrations of cholesterol encourage phagocytosis, suggesting that cholesterol is steadily converted to cholesteryl-α-glucoside and this is required to evade phagocytosis.…”

“…hp0421 mutant, suggesting that α-glucosylation of cholesterol is also required for T4SS functionality and lipid raft clustering. 85 As part of this process, the host receptor for the bacterial T4SS has been identified as α 5 β 1 integrin, which binds protein CagL at the periphery of the apparatus. 87 The integrin is raft-associated and therefore also recruited to sites of H. pylori-host cell contact, most likely by association with cholesterol.…”

Chemoenzymatic synthesis of cholesteryl-6-O-tetradecanoyl-α-d-glucopyranoside: a product of host cholesterol efflux promoted by Helicobacter pylori

“…8,9 Recent studies suggest that through a still unknown mechanism, H. pylori is able to sense cholesterol rich regions in host epithelial tissue and extract cholesterol from the host membrane for incorporation into α-CG and two other CGs modified at the C-6 hydroxyl of glucose. 12 One analog is acetylated with tetradecanoic acid and is referred to as α-CAG ( 1 ) (Figure 1), the other is phosphorylated and abbreviated as α-CPG. Together these three glycolipids make up approximately 25% of the lipid content of H. pylori and are a distinguishing feature of these bacteria.…”

“…19,20 While this enzyme is imbedded in the cell wall, presumably it loads UDP-Glc from the cytoplasm of H. pylori , which is then combined with extracellular cholesterol depleted from host membrane. 12 CG appears to be the substrate for an enzyme that transfers a tetradecanoic acid donor to the C-6 hydroxyl to form α-CAG but that enzyme and the exact acyl donor are yet to be identified. 20 Curiously, lyso-phosphoglycerolipid biosynthesis is correlated with increased α-CAG lipid content suggesting that at least some of the fatty acid donor may derive from phosphoglycerolipids.…”