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Basic residues in the T-cell receptor ? cytoplasmic domain mediate membrane association and modulate signaling.

ABSTRACT: The T-cell receptor (TCR) consists of a TCR?? heterodimer, a TCR? homodimer, and CD3?? and CD3?? heterodimers. The precise mechanism of T-cell triggering following TCR ligand engagement remains elusive. Previous studies reported that the cytoplasmic tail of CD3? binds to the plasma membrane through a basic residue-rich stretch (BRS) and proposed that dissociation from the membrane is required for phosphorylation thereof. In this report we show that BRS motifs within the cytoplasmic tail of TCR? mediate association with the plasma membrane and that TCR engagement results in TCR? dissociation from the membrane. This dissociation requires phosphorylation of the TCR? immunoreceptor tyrosine-based activation motifs by lymphocyte cell-specificprotein tyrosine kinase (Lck) but not ?-chain-associated protein kinase 70 binding. Mutations of the TCR? BRS motifs that disrupt this membrane association attenuate proximal and distal responses induced by TCR engagement. These mutations appear to alter the localization of TCR? with respect to Lck as well as the mobility of the TCR complex. This study reveals that tyrosine phosphorylation of the TCR? cytoplasmic domain regulates its association with the plasma membrane and highlights the functional importance of TCR? BRS motifs.

SUBMITTER: Zhang H

PROVIDER: S-EPMC3228420 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Basic residues in the T-cell receptor ζ cytoplasmic domain mediate membrane association and modulate signaling.

Zhang Hao H Cordoba Shaun-Paul SP Dushek Omer O van der Merwe P Anton PA

Proceedings of the National Academy of Sciences of the United States of America 20111114 48

The T-cell receptor (TCR) consists of a TCRαβ heterodimer, a TCRζ homodimer, and CD3γε and CD3δε heterodimers. The precise mechanism of T-cell triggering following TCR ligand engagement remains elusive. Previous studies reported that the cytoplasmic tail of CD3ε binds to the plasma membrane through a basic residue-rich stretch (BRS) and proposed that dissociation from the membrane is required for phosphorylation thereof. In this report we show that BRS motifs within the cytoplasmic tail of TCRζ ...[more]

PMID: 22084078

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Project description:We previously reported that neuraminidase (NA) pretreatment of human PBMCs markedly increased their cytokine response to lipopolysaccharide (LPS). To study the mechanisms by which this occurs, we transfected HEK293T cells with plasmids encoding TLR4, CD14, and MD2 (three components of the LPS receptor complex), as well as a NFκB luciferase reporting system. Both TLR4 and MD2 encoded by the plasmids are α-2,6 sialylated. HEK293T cells transfected with TLR4/MD2/CD14 responded robustly to the addition of LPS; however, omission of the MD2 plasmid abrogated this response. Addition of culture supernatants from MD2 (sMD2)-transfected HEK293T cells, but not recombinant, non-glycosylated MD2 reconstituted this response. NA treatment of sMD2 enhanced the LPS response as did NA treatment of the TLR4/CD14-transfected cell supplemented with untreated sMD2, but optimal LPS-initiated responses were observed with NA-treated TLR4/CD14-transfected cells supplemented with NA-treated sMD2. We hypothesized that removal of negatively charged sialyl residues from glycans on the TLR4 complex would hasten the dimerization of TLR4 monomers required for signaling. Co-transfection of HEK293T cells with separate plasmids encoding either YFP- or FLAG-tagged TLR4, followed by treatment with NA and stimulation with LPS, led to an earlier and more robust time-dependent dimerization of TLR4 monomers on co-immunoprecipitation, compared to untreated cells. These findings were confirmed by fluorescence resonance energy transfer (FRET) analysis. Overexpression of human Neu1 increased LPS-initiated TLR4-mediated NFκB activation and a NA inhibitor suppressed its activation. We conclude that (1) sialyl residues on TLR4 modulate LPS responsiveness, perhaps by facilitating clustering of the homodimers, and that (2) sialic acid, and perhaps other glycosyl species, regulate MD2 activity required for LPS-mediated signaling. We speculate that endogenous sialidase activity mobilized during cell activation may play a role in this regulation.

Dataset Information

Basic residues in the T-cell receptor ? cytoplasmic domain mediate membrane association and modulate signaling.

Publications

Basic residues in the T-cell receptor ζ cytoplasmic domain mediate membrane association and modulate signaling.

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