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CD8 kinetically promotes ligand binding to the T-cell antigen receptor
Authors:Gakamsky Dmitry M  Luescher Immanuel F  Pramanik Aladdin  Kopito Ronen B  Lemonnier François  Vogel Horst  Rigler Rudolf  Pecht Israel
Institution:Department of Immunology, and Department of Materials and Interfaces, Weizmann Institute of Science, 76100 Rehovot, Israel. lidima@wisemail.weizmann.ac.il
Abstract:The mechanism of CD8 cooperation with the TCR in antigen recognition was studied on live T cells. Fluorescence correlation measurements yielded evidence of the presence of two TCR and CD8 subpopulations with different lateral diffusion rate constants. Independently, evidence for two subpopulations was derived from the experimentally observed two distinct association phases of cognate peptide bound to class I MHC (pMHC) tetramers and the T cells. The fast phase rate constant ((1.7 +/- 0.2) x 10(5) M(-1) s(-1)) was independent of examined cell type or MHC-bound peptides' structure. Its value was much faster than that of the association of soluble pMHC and TCR ((7.0 +/- 0.3) x 10(3) M(-1) s(-1)), and close to that of the association of soluble pMHC with CD8 ((1-2) x 10(5) M(-1) s(-1)). The fast binding phase disappeared when CD8-pMHC interaction was blocked by a CD8-specific mAb. The latter rate constant was slowed down approximately 10-fold after cells treatment with methyl-beta-cyclodextrin. These results suggest that the most efficient pMHC-cell association route corresponds to a fast tetramer binding to a colocalized CD8-TCR subpopulation, which apparently resides within membrane rafts: the reaction starts by pMHC association with the CD8. This markedly faster step significantly increases the probability of pMHC-TCR encounters and thereby promotes pMHC association with CD8-proximal TCR. The slow binding phase is assigned to pMHC association with a noncolocalized CD8-TCR subpopulation. Taken together with results of cytotoxicity assays, our data suggest that the colocalized, raft-associated CD8-TCR subpopulation is the one capable of inducing T-cell activation.
Keywords:TCR  T-cell receptor  APC  antigen presenting cells  MHC  major histocompatibility complex  pMHC  peptide-MHC complex  PbCS(ABA)  an azido-benzoic acid (ABA) derivative of the peptide SYIPSAEK(ABA)I  pCw3  peptide derived from HLA-Cw3  FCS  fluorescence correlation spectroscopy  MβCD  methyl-β-cyclodextrin  PS days  poststimulation days  MFI  mean fluorescence intensity  FITC  fluorescein isothiocyanate  FITC-CTB  FITC-labeled cholera toxin B subunit
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