Molecular mechanisms by which the large GTPase hGBP1 inhibits actin-based bacterial dissemination
Immunology
Final Report Abstract
The human-adapted bacterial pathogen Shigella flexneri hijacks the host cell actin cytoskeleton and uses the force of polymerizing actin to propel itself through the cytosol of infected cells. Shigella exploits actin-based motility also to spread directly from cell to cell allowing the enteric bacteria to disseminate rapidly through the intestinal epithelium and to efficiently avoid extracellular immune defenses. Host defense against Shigella therefore critically depends on cell-autonomous immunity, which is the ability of a single cell regardless of its tissue of origin to contain or control intracellular pathogens. Key players of cell-autonomous immunity are guanylatebinding proteins (GBPs). Two independent studies reported that human GBP1 (hGBP1) associates with intracellular Shigella and blocks bacterial actin tail formation thereby immobilizing Shigella in the cytosol of infected cells and preventing cell-to-cell spread. How hGBP1 associates with cytosolic bacteria and inhibits actin-based motility to block Shigella dissemination represented a critical gap in knowledge. By using a combination of biochemical cell-free approaches, cellular immunology, and microbiology, I discovered that hGBP1 is a sensor for the bacterial surface molecule lipopolysaccharide (LPS), which is anchored in the bacterial outer membrane via its lipid A moiety, consists further of inner and outer core oligosaccharides and an outward facing O-antigen polysaccharide. Polymeric hGBP1 associates with the bacterial surface via direct interaction with LPS before transforming into an intercalated protein mesh surrounding the bacterial cell. I found that dynamic polymerization is strictly required for encapsulating bacteria thereby revealing the biological function for the previously biochemical characterized hGBP1 polymer. Reorganization of the surface attached hGBP1 polymers to a stable protein coat depended on GBP1’s C-terminal triple arginine motif and the presence of O-antigen on bacteria. The O-antigen polysaccharides form a chemo-physical barrier which protect gram-negative bacteria from antimicrobial peptides and immune sensors targeting the lipid A moiety. I discovered that once integrated into the bacterial outer membrane, hGBP1 acts as an antibacterial molecular soap and emulsifies outer membrane-anchored LPS to break down the O-antigen barrier and unmask lipid A for recognition by antimicrobials. My studies revealed that detergent-like hGBP1 on its own was not bactericidal but facilitates bacterial killing synergistically with lipid A sensing last resort antibiotic polymyxin B rendering hGBP1 as potential target for host directed therapies against multi-resistant bacteria. Identifying hGBP1 as novel cytosolic surfactant further explained how hGBP1 restricts Shigella actin-based motility. By disrupting the outer membrane integrity, the hGBP1 microcapsule interferes with the localization and function of Shigella motility factor IcsA thereby inhibiting actin polymerization at the bacterial pole and preventing direct cell-to-cell spread. This finding suggests that the hGBP1 microcapsule likely affects other outer membrane virulence factors constituting a more broadly host defense strategy.
Publications
- (2020) A Rapidly Evolving Polybasic Motif Modulates Bacterial Detection by Guanylate Binding Proteins. mBio 11:e00340-20
Kohler KM, Kutsch M, Piro AS, Wallace GD, Coers J# & Barber MF
(See online at https://doi.org/10.1128/mbio.00340-20) - (2020) Direct binding of polymeric GBP1 to LPS disrupts bacterial cell envelope functions. EMBO J 39:e104926
Kutsch M, Sistemich L, Lesser CF, Goldberg MB, Herrmann C & Coers J
(See online at https://doi.org/10.15252/embj.2020104926) - (2021) Human guanylate binding proteins: nanomachines orchestrating host defense. FEBS J 288:5826-5849
Kutsch M & Coers J
(See online at https://doi.org/10.1111/febs.15662) - (2021) The GBP1 microcapsule interferes with IcsA-dependent septin cage assembly around Shigella flexneri. FEMS Pathogens and Disease 79:ftab023
Kutsch M, Gonzáles-Prieto C, Lesser CF & Coers J
(See online at https://doi.org/10.1093/femspd/ftab023)
