The Complementarity Determining Region 2 of BV8S2 (Vβ8.2) Contributes to Antigen Recognition by Rat Invariant NKT Cell TCR

Pyż, Elwira; Naidenko, Olga V.; Miyake, Sachiko; Yamamura, Takashi; Berberich, Ingolf; Cardell, Susanna; Kronenberg, Mitchell; Herrmann, Thomas

doi:10.4049/jimmunol.176.12.7447

Cited by 32 publications

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“…While the rAV14S6 93A containing iNKT TCR revealed substantial binding to rCD1d, there was nearly no binding to mCD1d or huCD1d detectable (Fig. 1C), which is in accordance with previous reports describing the lack of species cross-reactivity of rat iNKT cells [15,22]. Intriguingly, the rAV14S2 93G-containing iNKT TCR exhibited a lower rCD1d but an increased mCD1d and huCD1d binding when compared to rAV14S6 93A (Fig.…”

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confidence: 80%

“…Our analysis of iNKT TCR transductants revealed substantial differences in responses to αGC, depending on the type of rat iNKT α chain as well as reduced cross-reactivity with mouse CD1d, reflected by poor or missing binding of αGC-loaded mouse CD1d tetramers and dimers [15,22]. Finally, our site-directed mutagenesis analysis demonstrated for the first time the crucial role of the CDR2β for the response to αGC [15].The representatives of the rat multimember AV14 gene family differ at several positions and have been divided into type 1 and type 2 sequences by the similarity of their CDR2α [20,23]. In addition, variability within the TCRα chain of primary iNKT-cell populations has been observed at position 93 (V-J transition), i.e.…”

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confidence: 99%

“…Two α chains with AV14/AJ18 rearrangements containing different AV14 and amino acids at position 93 (Fig. 1A) were cloned from F344 rats and expressed together with a characterized rBV8S4-like β chain (CDR2+4) in a TCR negative T cell hybridoma [15] (Fig. 1B).…”

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The hypervariable region 4 (HV4) and position 93 of the α chain modulate CD1d‐glycolipid binding of iNKT TCRs

et al. 2015

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TCRs of invariant NKT (iNKT) cells bind α-galactosylceramide (αGC) loaded CD1d in a highly conserved fashion and show a characteristic TCR gene usage: An "invariant" α chain with a canonical AV14/AJ18 rearrangement in mice (AV24/AJ18 in humans) is paired with β chains containing characteristic Vβ segments. In the rat, a multimember AV14 gene family increases the variability within this system. This study characterizes CD1d binding of rat AV14 gene segments in TCR transductants as well as CD1d binding and iNKT TCR expression of expanded polyclonal F344 rat iNKT populations. It defines an important role of position 93 at the V-J transition for TCR avidity and species cross-reactivity of the rat iNKT TCR. Furthermore, for the first time we identified variability within the fourth hypervariable loop (HV4) of the α chain as a modulator of CD1d:αGC binding in rat and mouse. Additionally, we confirmed the importance of the CDR2β for CD1d:αGC binding, but also show that the CDR3β may even have opposite effects on binding depending on the pairing α chain. Altogether, we characterized naturally occurring sources of variability for the iNKT TCR and speculate that they rather level than increase the largely germline encoded differences of iNKT TCR ligand avidity.Keywords: CD1d r iNKT cells r Molecular modeling r Rat r T-cell receptors Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionType 1 NKT cells (invariant NKT (iNKT) cells) are a subset of αβ T cells characterized by the expression of a semi-invariant TCR. In contrast to conventional αβ T cells, iNKT cells are not MHCrestricted, but recognize CD1d, a nonpolymorphic MHC class Ilike molecule presenting lipid antigens [1,2]. The lineage-defining iNKT TCR contains an invariant α chain with a uniform AV14/AJ18 rearrangement in mice (AV24/AJ18 in human) and a limited number of β chain V segments (mostly BV8S2, BV7, and BV2 in mice Correspondence: Prof. Thomas Herrmann e-mail: herrmann-t@vim.uni-wuerzburg.de and the BV8S2 homologue BV11 in humans) [3][4][5][6][7]. This decreased variability within the TCR leads to a very restricted CDR repertoire with CDR3β as the only exception.The first characterized and one of the strongest antigens for iNKT cells is KRN7000 (α-galactosylceramide, αGC), the synthetic derivative of a compound isolated from the marine sponge Agelas mauritanus [3]. The rather fixed mode of interaction between the iNKT TCR and the CD1d:αGC complex does not only allow for direct identification of iNKT cells by CD1d oligomers loaded with αGC or its derivative PBS57, but also defines this lineage of αβ T cells. Crystallographic studies have shown that, for mouse and human, direct contact is mostly mediated by CDR1α, CDR3α, and CDR2β of the iNKT TCR, displaying the restricted usage of CDRs within this system [8,9]. Thereby, an alanine C 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu Eur. J. Immunol. 2015. 45: 2122-2133 Molecular immunology 2123 scanning mutagenesis study...

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The hypervariable region 4 (HV4) and position 93 of the α chain modulate CD1d‐glycolipid binding of iNKT TCRs

et al. 2015

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“…The mouse iNKTa chain uses a glycine at position 93, with rare variants containing alanine, valine or isoleucine [11] but the impact of these variants on iNKT-cell binding or function has not been assessed, and this specific site has not been included in any alanine scanning studies of the mouse iNKT-cell TCR:CD1d interaction. Interestingly, natural iNKTa variants at p93 also occur in the rat, where they have been shown to impact on iNKTcell function [12]. Furthermore, subtle variations at the TCR Va7.2-Ja33 junction that are regularly found in mucosa-associated invariant T (MAIT) cells, which are another ''invariant'' T lymphocyte subset in humans restricted by the non-polymorphic MHC class I-related MR1 molecule [13].…”

Section: Discussionmentioning

confidence: 99%

Natural variations at position 93 of the invariant Vα24‐Jα18 α chain of human iNKT‐cell TCRs strongly impact on CD1d binding

et al. 2011

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Human invariant natural killer T (NKT) cell TCRs bind to CD1d via an ''invariant'' Va24-Ja18 chain (iNKTa) paired to semi-invariant Vb11 chains (iNKTb). Single-amino acid variations at position 93 (p93) of iNKTa, immediately upstream of the ''invariant'' CDR3a region, have been reported in a substantial proportion of human iNKT-cell clones (4-30%). Although p93, a serine in most human iNKT-cell TCRs, makes no contact with CD1d, it could affect CD1d binding by altering the conformation of the crucial CDR3a loop. By generating recombinant refolded iNKT-cell TCRs, we show that natural single-nucleotide variations in iNKTa, translating to serine, threonine, asparagine or isoleucine at p93, exert a powerful effect on CD1d binding, with up to 28-fold differences in affinity between these variants. This effect was observed with CD1d loaded with either the artificial a-galactosylceramide antigens KRN7000 or OCH, or the endogenous glycolipid b-galactosylceramide, and its importance for autoreactive recognition of endogenous lipids was demonstrated by the binding of variant iNKT-cell TCR tetramers to cell surface expressed CD1d. The serine-containing variant showed the strongest CD1d binding, offering an explanation for its predominance in vivo. Complementary molecular dynamics modeling studies were consistent with an impact of p93 on the conformation of the CDR3a loop.Key words: CD1d . iNKT cells . Molecular modeling . Surface plasmon resonance . TCR IntroductionInvariant natural killer T (iNKT) lymphocytes are a versatile and potent subset of T cells which play important roles in both host defense and tolerance. In contrast to conventional T cells, they recognize the non-polymorphic lipid-antigen-presenting molecule CD1d via highly conserved TCRs. In fact, all iNKT-cell TCRs use an ''invariant'' TCRa chain (iNKTa), Va24-Ja18 in humans, Va14-Ja18 in mice, paired to ''semi-invariant'' TCRb chains (iNKTb), which in humans solely utilize Vb11 rearranged with diverse TCR Ã These authors contributed equally to this work. 248Jb segments. For all iNKT-cell TCRs, binding to CD1d is primarily mediated by the Va-Ja rearranged ''invariant'' CDR3a loop as well as the two germline-encoded CDR1a and CDR2b loops [1,2]. In addition, we have recently shown that the only variable region of human iNKT-cell TCRs, i.e. the CDR3b loop, strongly modulates the affinity to CD1d [3].Human iNKT-cell clones with single-amino acid variations in the ''invariant'' Va24-Ja18 iNKTa chain have been previously identified [4,5], and a recent study found that 4-30% of human iNKT-cell clones carried such variations [4]. However, the effect of these variations on iNKT-cell TCR binding to CD1d has not been examined. All of the described variations to date are located immediately upstream of the CDR3a loop region and most of these involve residue 93 (p93). The most commonly used amino acid at this position is serine (Ser93), which is encoded for by the germline sequence of human Va24S1 (AE00521.1), but other identified natural variations at this position inclu...

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“…All Chinese hamster ovary (CHO) cell derivatives were retrovirally transduced with human CD80 as described in [16]. For transduction of BTN3A1 the same type of retroviral vector was used but containing a fulllength BTN3A1 coding sequence obtained by RT-PCR of RAJI cells.…”

Section: Cell Linesmentioning

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Vγ9Vδ2 TCR‐activation by phosphorylated antigens requires butyrophilin 3 A1 (BTN3A1) and additional genes on human chromosome 6

et al. 2014

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The Complementarity Determining Region 2 of BV8S2 (Vβ8.2) Contributes to Antigen Recognition by Rat Invariant NKT Cell TCR

Cited by 32 publications

References 45 publications

The hypervariable region 4 (HV4) and position 93 of the α chain modulate CD1d‐glycolipid binding of iNKT TCRs

The hypervariable region 4 (HV4) and position 93 of the α chain modulate CD1d‐glycolipid binding of iNKT TCRs

Natural variations at position 93 of the invariant Vα24‐Jα18 α chain of human iNKT‐cell TCRs strongly impact on CD1d binding

Vγ9Vδ2 TCR‐activation by phosphorylated antigens requires butyrophilin 3 A1 (BTN3A1) and additional genes on human chromosome 6

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