α-Glucuronosyl and α-Glucosyl Diacylglycerides, Natural Killer T Cell-Activating Lipids from Bacteria and Fungi

Abstract

Natural killer T cells express T cell receptors (TCRs) that recognize glycolipid antigens in association with the antigen-presenting molecule CD1d. Here, we report the concise chemical synthesis of a range of saturated and unsaturated α-glucosyl and α-glucuronosyl diacylglycerides of bacterial and fungal origins from allyl α-glucoside with Jacobsen kinetic resolution as a key step. We show that these glycolipids could be recognized by a classical type I NKT TCR that uses an invariant Vα14-Jα18 TCR α-chain, but also by an atypical NKT TCR that uses a different TCR α-chain (Vα10-Jα50). In both cases, recognition was sensitive to the lipid fine structure, and included recognition of glycosyl diacylglycerides bearing branched (R- and S-tuberculostearic acid) and unsaturated (oleic and vaccenic) acids. The TCR footprints on CD1d-loaded with a mycobacterial α-glucuronosyl diacylglyceride was assessed using mutant CD1d molecules and, while similar to that for α-GalCer recognition by a type I NKT TCR, were more sensitive to mutations when α-glucuronosyl diacylglyceride was the antigen. In summary, we provide an efficient approach for synthesis of a broad class of bacterial and fungal α-glycosyl diacylglyceride antigens and demonstrate that they can be recognised by TCRs derived from type I and atypical NKT cells.

Benzofuran sulfonates and small self-lipid antigens activate type II NKT cells via CD1d

“…Overall, the CD1d docking footprints of the PPBF-reactive TCRs were distinct from the F′ docking of type I NKT TCRs (Fig. 4) (40,41), α-GlcADAG, or α-GalCer-reactive TRAV13-TRAJ50 + TCRs (37,42). These footprints were also distinct to those described for other type II and atypical NKT TCRs (34,38,39,43,44).…”

Benzofuran sulfonates and small self-lipid antigens activate type II NKT cells via CD1d

“…When PPBF was not included, similar results were observed for both the ABd and the VD1 TCR (data not shown). Overall, the CD1d docking footprint of the VD1G9 and ABd PPBF-reactive TCRs were distinct to the F′-oriented docking of type I NKT TCRs (Figure 4) 39, 40 , a-GlcADAG, a-GalCer reactive (type Ia ) TRAV13-TRAJ50 + TCRs 36,41 , and other type II NKT TCRs 38 or the A′-oriented docking of atypical 37 or sulfatide-reactive type II NKT 42,43 and TRDV1 + CD1d-restricted TCRs 33 . Thus, PPBF-reactive TCRs potentially dock over the top of the antigen-binding cleft of CD1d, which might allow productive analysis of molecules that are trapped between CD1d and TCR using TCR trap technology 23,24 .…”

Lipid antigens in adipose tissue

“…Precise determination of CD1d lipid antigens in general has proven to be challenging. Previous research based on a variety of different methods including extrapolation of data from cells deficient in lipid synthesis genes [87,88], application of antibodies that detect specific antigens loaded into CD1d [89,90], the use of specific catabolic enzymes to modify candidate lipid antigens in vitro prior to testing [89], fractionation of various lipid sources to identify candidates [60,[91][92][93][94][95], plate-bound CD1d tetramers to assess lipid antigen stability, efficiency and efficacy during iNKT cross talk assays [60,91,96,97], synthesis of αGalCer analogues and exogenous bacterial antigens to elicit a controlled cytokine response from iNKT's [98][99][100], and predictive modeling alongside molecular dynamics approaches [101]. Together all previously used methods have revealed a large spectrum of lipid antigen candidates that fit the criteria to bind CD1d in a context dependent manner corroborating the promiscuous nature of CD1d.…”