A single amino acid residue of CDR3β alters the antigenicity and autoreactivity of human invariant natural killer T cell receptors (IRM7P.707)

Abstract

Abstract Detection of human invariant natural killer T (iNKT) cells relies on staining for their T cell receptor (TCR), invariant Vα24 TCRα chain paired with semi-variant Vβ11 TCRβ chain, or α-GalCer-CD1d tetramer. It has been assumed that all Vα24+ iNKT cells are stained with similar intensity by anti-Vα24 mAbs, clones C15 and 6B11, regardless of the hypervariable Vβ11 CDR3β sequences. By analyzing a large panel of autoreactive human iNKT TCRs, we found that TCRs encoding CASR at the start of CDR3β were stained with significantly lower intensity by C15 than those encoding CASS. Substitution of S to R at the fourth CDR3β residue was sufficient to diminish staining intensity by anti-Vα24 mAbs. Interestingly, the autoreactivity was also reduced in the TCRs encoding CASR. Importantly, α-GalCer tetramer positive population of human primary T cells presented low and high intensity subsets when stained with anti-Vα24 mAbs. Vβ11 TCRβ chains were cloned from each sorted population, and the Vα24 low population contained a significantly higher frequency of Vβ11 TCRs encoding CASR compared to the Vα24 high population. Therefore, human iNKT TCRs encoding CASR may falsely be negative when stained with anti-Vα24 mAbs, which underestimate the frequency of bona fide Vα24+ iNKT cells. Our data demonstrate that the fourth CDR3β amino acid residue affects iNKT TCR autoreactivity and binding of anti-Vα24 mAbs, suggesting that the CDR3β sequence alters the structural conformation of the Vα24 TCRα chain.