Membrane Cholesterol Is Crucial for Clostridium difficile Surface Layer Protein Binding and Triggering Inflammasome Activation

Abstract

Clostridium difficile, an obligate anaerobic gram-positive bacillus, generates spores and is commonly found colonizing the human gut. Patients with C. difficile infection (CDI) often exhibit clinical manifestations of pseudomembranous colitis or antibiotic-associated diarrhea. Surface layer proteins (SLPs) are the most abundant proteins in the C. difficile cell wall, suggesting that they might involve in immune recognition. Our previous results demonstrated that C. difficile triggers inflammasome activation. Here, we found SLPs as well as C. difficile induced inflammasome activation, and in a dose-dependent manner. In addition, the cholesterol-rich microdomains on the cell membrane (also referred to as lipid rafts) are thought to be crucial for bacterial adhesion and signal transduction. We demonstrated that lipid rafts participated in C. difficile SLPs binding to the cell membrane. Fluorescence microscopy showed that membrane cholesterol depletion by methyl-β-cyclodextrin (MβCD) reduced the association of SLPs with the cell surface. The coalescence of SLPs in the cholesterol-rich microdomains was confirmed in C. difficile-infected cells. Furthermore, the inflammasome activations induced by SLPs or C. difficile were abrogated by MβCD. Our results demonstrate that SLPs recruit the lipid rafts, which may be a key step for C. difficile colonization and inducing inflammasome activation.

Helicobacter pylori cholesterol-α-glucosyltransferase manipulates cholesterol for bacterial adherence to gastric epithelial cells

“…Bacterial virulence factors employing membrane cholesterol microdomains to gain access to host cells have been reported previously [34][35][36]. Cholesterol usurping/depleting agents have been used to alleviate infectious diseases by preventing microbial entry into host cells [37].…”

“…For instance, cholesterol-lowering agents (i.e., statins) potentially attenuate pathogen infectivity [38][39][40][41]. Another cholesterol-depleting agent, MβCD, is commonly employed to reduce microbial adherence to the host cell membrane [36,42,43]. Our previous studies demonstrated that statin use significantly reduced the incidence of H. pylori-related diseases [41,44,45].…”

Campylobacter jejuni Cytolethal Distending Toxin C Exploits Lipid Rafts to Mitigate Helicobacter pylori-Induced Pathogenesis

“…Pathogens and their virulence factors exploiting lipid rafts to gain entry into host cells have been reported elsewhere ( Wang and Hajishengallis, 2008 ; Frisan, 2016 ; Chen Y. et al, 2020 ). Previous studies have used cholesterol-depleting agents to reduce pathogen infections by inhibiting their entry into target cells.…”

“…Previous studies have used cholesterol-depleting agents to reduce pathogen infections by inhibiting their entry into target cells. For example, depletion of membrane cholesterol by methyl-β-cyclodextrin impaired pathogen attachment to the cell surface ( Guo et al, 2017 ; Owczarek et al, 2018 ; Chen Y. et al, 2020 ). Statins, inhibitors of HMG-CoA reductase, are cholesterol-lowering agents that have been employed to reduce microbial infectivity ( Boyd et al, 2012 ; Motzkus-Feagans et al, 2012 ; Skerry et al, 2014 ).…”

Antagonistic Activities of Lactobacillus rhamnosus JB3 Against Helicobacter pylori Infection Through Lipid Raft Formation

“…However, it has been indicated that the Le x antigen and TLR 4 are clustered in lipid rafts to facilitate the adherence of H. pylori to the gastric epithelial cells (9)(10)(11)(12), and disrupting lipid rafts may prevent H. pylori-associated gastric cancer (13). Lipid rafts consist of phospholipids, sphingolipids, and cholesterol and act as signaling platforms and gateways for microorganisms to adhere to host cells (10,(14)(15)(16)(17). Furthermore, H. pylori expresses cholesteryl a-D-glucopyranoside acyltransferase (CGAT), which is encoded by the hp0499 gene, to enhance lipid raft clustering on host cell membranes (9).…”