The membrane-bound and soluble forms of HLA-G bind identical sets of endogenous peptides but differ with respect to TAP association

Abstract

The class Ib antigen HLA-G is expressed as a membrane-bound protein like classical class Ia molecules (M.HLA-G) but, unlike typical class I, is also expressed as a soluble protein (S.HLA-G) with a unique C terminus. Our results show that, similar to classical class I proteins, the membrane-bound form of HLA-G associated with TAP, as evidenced by the ability to immunoprecipitate HLA-G class I heavy chain with TAP antisera. In contrast, the soluble G protein did not appear to associate with TAP in the same manner, since similar immunoprecipitation experiments failed to detect soluble G complex. A detailed analysis of peptides bound to the soluble and membrane HLA-G proteins expressed in the B lymphoblastoid cell line 721.221 showed that, like class Ia complexes, both HLA-G proteins consist of heavy and light chains complexed with nonameric peptides in a 1:1:1 ratio. The two proteins bind essentially the same set of peptides, which are derived from a variety of intracellular proteins and define a peptide motif for HLA-G. The peptides contain Leu at the C terminus and Pro or small hydrophobic amino acids in position 3 followed by Pro or Gly in position 4. The complexity of the bound peptides is lower than that found for some class Ia complexes, but is more similar to class Ia than to the limited repertoire of some murine class Ib molecules.

Alloreactive CD4+ and CD8+ T cells express the immunotolerant HLA‐G molecule in mixed lymphocyte reactions: in vivo implications in transplanted patients

“…cDNA were prepared from 1-5 ? g of total RNA using Ready To Go TM kit and oligo (dT) [12][13][14][15][16][17][18] , according to the manufacturer's recommendations. For HLA-G1 and HLA-G5 transcript detection, PCR amplification and hybridization of the PCR products were performed as described [34].…”

“…The 39-kDa full-length HLA-G1 heavy chain is constituted by three extracellular domains ( § 1, § 2 and § 3) associated with the g 2-microglobulin ( g 2m) and a nonapeptide. The 37-kDa soluble HLA-G5 heavy chain is encoded by an alternative transcript which retains part of intron 4 and lacks both transmembrane and cytoplasmic domains [13]. HLA-G5 also associates to g 2m and peptide [13].…”

“…The 37-kDa soluble HLA-G5 heavy chain is encoded by an alternative transcript which retains part of intron 4 and lacks both transmembrane and cytoplasmic domains [13]. HLA-G5 also associates to g 2m and peptide [13]. In terms of function, both HLA-G1 and HLA-G5 isoforms display immunotolerant properties [12].…”

Induction of HLA-G-restricted human cytomegalovirus pp65 (UL83)-specific cytotoxic T lymphocytes in HLA-G transgenic mice

“…The TAP1 2 /TAP2 2 -deficient T2 cell line (Salter & Cresswell, 1986) Peptide-binding assay. Six pp65-derived peptides were designed based on the HLA-G-binding motif (Diehl et al, 1996;Lee et al, 1995). We also used peptides eluted from HLA-G molecules (Lee et al, 1995) and an HLA-B27-binding peptide (Solache et al, 1999) as positive and negative controls, respectively.…”

“…HLA-G molecules have a peptide-binding groove that is homologous to those of classical MHC class I molecules. In addition, HLA-G was shown to present nonameric peptides derived from a variety of intracellular proteins (Diehl et al, 1996;Lee et al, 1995). It was proposed that HLA-G on trophoblast cells interacts with several inhibitory or triggering NK cell receptors to protect placental cells against the cytotoxic activity of NK cells (Lanier, 1999).…”