Decoding Gut microbial immunity: How Natural Killer T cells recognise Bacteroides fragilis glycosphingolipids 2256741

Abstract Introduction The human gut microbiota comprises more than 50% of Bacteroides species that produce small diffusible molecules like sphingolipids that play a key role in modulating the host’s immune responses. In particular, Bacteroides fragilis produces glycosphingolipids termed ‘BfaGCs’ that can activate type I Natural Killer T (NKT) cells. BfaGCs exhibit distinct structural characteristics, including truncated sphinganine chains, varied branching patterns, and specific functional groups, distinguishing them from the canonical type I NKT cell marker, α-galactosylceramide (KRN7000). Methods Using a combinatorial cellular immunology, mass spectrometry and X-ray crystallography approach, we provide the first insights into the molecular mechanism of recognition of four novel BfaGCs presented by the antigen-presenting molecule CD1d, in complex with the type I NKT TCR. Results The co-culture assay performed with bone marrow-derived dendritic cells and NKT cells in the presence of specific BfaGCs indicated that branching in their sphinganine chain is a critical determinant of NKT cell activation. The three-dimensional complex structures revealed that the TCR adopted a parallel docking topology atop the F’-pocket of CD1d in recognising the presented BfaGCs, reminiscent of other published type I NKT TCR lipid complexes. Nevertheless, the terminal sphinganine branching of BfaGCs facilitates unique interactions within the CD1d F’-pocket, thereby underpinning their distinctive agonistic properties. Also, the NKT TCR demonstrates high binding affinities for both stimulatory and non-stimulatory forms of CD1d-presented BfaGCs, affirming BfaGCs as bona fide CD1d ligands that modulate host immune defences via NKT cells. Conclusion Thus, BfaGCs were demonstrated to function as immunomodulatory mediators influencing the host’s defence in the context of NKT cells. Collectively, our findings enhance understanding of the symbiotic interplay between lipid-producing gut microbes and the host. Funding Source Australian Research Council Topic Categories Mucosal and Regional Immunology (MUC)