Cell wall glycosphingolipids ofSphingomonas paucimobilisare CD1d-specific ligands for NKT cells

Abstract

The current consensus on characterization of NKT cells is based on their reactivity to the synthetic glycolipid, a-galactosylceramide (a-GalCer) in a CD1d-dependent manner. Because of the limited availability of a-GalCer, there is a constant search for CD1d-presented ligands that activate NKT cells. The a-anomericity of the carbohydrate is considered to be an important requisite for the CD1d-specific activation of NKT cells. The gram-negative, lipopolysaccharide-free bacterium Sphingomonas paucimobilis is known to contain glycosphingolipids (GSL) with a-anomeric sugars attached to the lipid chain. Here, we report that GSL extracted from this bacterium are able to stimulate NKT cells in a CD1d-specific manner. In addition, soluble CD1d-Ig dimers loaded with this lipid extract specifically bind to NKT cells (but not conventional T cells). Further studies on the S. paucimobilis GSL could potentially lead to other natural sources of CD1d-specific ligands useful for NKT cell analyses and aimed at identifying novel therapies for a variety of disease states. IntroductionLipid antigens, including phospholipids and glycosphingolipids (GSL), are presented by CD1d -a subset of the CD1 family of MHC class I-like molecules -to specialized immune effector cells called NKT cells [1]. Located on a different chromosome than the MHC, five different CD1 genes encode CD1 molecules -CD1a, CD1b, CD1c, CD1d and CD1e -in humans, whereas only two homologues of CD1d (CD1d1 and CD1d2) are present in mice [2]. On the basis of the crystal structure of mouse CD1d1 [3], it is predicted that the fatty-acyl chains are buried in the hydrophobic pocket of the molecule with the hydrophilic head-group of the lipid antigen available outside the molecule for its interaction with the NKT cell receptor. Even though the lipidbinding groove of CD1d is widely accommodative of many lipid groups, it is believed that only the sugar structures in a-anomeric orientation are able to stimulate NKT cells [4]. Because of the lack of physiological glycolipids with terminal a-anomeric sugars, it is hypothesized that both altered selfglycolipids during a pathological process and exogenous antigenic glycolipids could be potential ligands presented by CD1d [4]. The presence of GSL in the cell wall of some bacterial strains indicates the possibility of these bacterial GSL being presented by CD1d. Although human CD1a, b and c molecules are known to present mycobacterial antigens to human NKT cells, there is a lack of substantial evidence to show that CD1d has the ability to present these microbial lipids [5]. In a recent study, Fischer et al. [6] were able to show that mycobacterial phosphoinositolmannosides (PIM) could bind to soluble CD1d and activate human and mouse NKT cells. However, only a fraction of NKT cells detected by agalactosylceramide (a-GalCer) could be stained by a CD1d tetramer loaded with mycobacterial PIM [6]. A recent study [7] was able to show the binding of synthetic a-GalNAc containing GSL to cell surface CD1d, but no functional studies were done t...

Retracted: α‐GalCer ameliorates listeriosis by accelerating infiltration of Gr‐1+ cells into the liver

“…a-glucuronosylceramide from Sphingomonas sp. [26][27][28][29] and Ehrlichia sp. [28], a-galactosyldiacylglycerol from Borrelia sp.…”

“…a-glucuronosylceramide from Sphingomonas sp. [26][27][28][29] and Ehrlichia sp.[28], a-galactosyldiacylglycerol from Borrelia sp.[30], phosphatidylinositol mannoside from Mycobacterium sp.[31] and phosphoethanolamine [32] have recently been identified as ligands for iNKT cells, although all of these activate iNKT cells to a lesser degree than a-GalCer. Although lysosomal glycosphingolipid, isoglobotrihexosylceramide has been reported as a possible endogenous ligand for iNKT cells [28,33], a recent study argues against this possibility [34].…”

CD1d-restricted iNKT cells, the ‘Swiss-Army knife’ of the immune system

“…Whereas the ability of some previously described ligands of microbial origin -Plasmodium (and Trypanosoma) glycosylphosphatidylinositol (GPI) and Mycobacterium phosphatidylinositol mannoside (PIM) -to activate iNKT cells has been questioned, recent work has rigorously demonstrated that GSL-1 from Sphingomonas and BbGL-II from Borrelia undoubtedly stimulate NKT cells in a CD1d-dependent manner [49,[63][64][65]. It remains to be determined whether direct recognition of these glycolipids by iNKT cells is responsible for the iNKT cell-dependent anti-bacterial activity noted during infection with these bacteria [49,63].…”

“…iNKT cells have been shown to directly recognize α-linked glycosphingolipids and diacylglycerol antigens that are expressed by bacteria such as Sphingomonas, Ehrlichia and Borrelia burgdorferi in a CD1d-dependent manner [49,[63][64][65] (Figure 1C). The biological response to these glycolipid antigens includes the production of IFNγ and IL-4 by iNKT cells.…”

NKT cells: In the beginning…

“…I will not relate these developments here in any detail, since the focus of this "historical feature" has been to recount the circumstances that led to our current definition of iNKT cells. Suffice it to say that there is now overwhelming evidence that iNKT cells can recognize a variety of microbial glycolipids [42][43][44], although the self glycolipids responsible for thymic selection of iNKT cells remain controversial [45,46]. It is obvious, however, that our ultimate understanding of the biological relevance of iNKT cells will depend to a large extent on the future identification of relevant endogenous and exogenous physiological glycolipid ligands.…”