scholarly journals Intracellular Ca2+Release Triggers Translocation of Membrane Marker FM1–43 from the Extracellular Leaflet of Plasma Membrane into Endoplasmic Reticulum in T Lymphocytes

2005 ◽  
Vol 280 (16) ◽  
pp. 16377-16382 ◽  
Author(s):  
Sepehr Dadsetan ◽  
Vyacheslav Shishkin ◽  
Alla F. Fomina
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
T Cell ◽  
T Cell Receptor ◽  
Membrane Trafficking ◽  
Cell Receptor ◽  
Multivesicular Bodies ◽  
Close Proximity ◽  
Intracellular Ca ◽  
Stimulation Of

Stimulation of T cell receptor in lymphocytes enhances Ca2+signaling and accelerates membrane trafficking. The relationships between these processes are not well understood. We employed membrane-impermeable lipid marker FM1–43 to explore membrane trafficking upon mobilization of intracellular Ca2+in Jurkat T cells. We established that liberation of intracellular Ca2+with T cell receptor agonist phytohemagglutinin P or with Ca2+-mobilizing agents ionomycin or thapsigargin induced accumulation of FM1–43 within the lumen of the endoplasmic reticulum (ER), nuclear envelope (NE), and Golgi. FM1–43 loading into ER-NE and Golgi was not mediated via the cytosol because other organelles such as mitochondria and multivesicular bodies located in close proximity to the FM1–43-containing ER were free of dye. Intralumenal FM1–43 accumulation was observed even when Ca2+signaling in the cytosol was abolished by the removal of extracellular Ca2+. Our findings strongly suggest that release of intracellular Ca2+may create continuity between the extracellular leaflet of the plasma membrane and the lumenal membrane leaflet of the ER by a mechanism that does not require global cytosolic Ca2+elevation.

scholarly journals The gamma and epsilon subunits of the CD3 complex inhibit pre-Golgi degradation of newly synthesized T cell antigen receptors.

1990 ◽  
Vol 110 (4) ◽  
pp. 973-986 ◽  
Author(s):  
T Wileman ◽  
G R Carson ◽  
M Concino ◽  
A Ahmed ◽  
C Terhorst
Keyword(s):  
T Cells ◽  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
T Cell ◽  
T Cell Receptor ◽  
Intracellular Transport ◽  
Cell Receptor ◽  
Dna Transfection ◽  
Antigen Receptors ◽  
Intracellular Degradation

The T cell receptor for antigen (TCR) is composed of six different transmembrane proteins. T cells carefully control the intracellular transport of the receptor and allow only complete receptors to reach the plasma membrane. In an attempt to understand how T cells regulate this process, we used c-DNA transfection and subunit-specific antibodies to follow the intracellular transport of five subunits (alpha beta gamma delta epsilon) of the receptor. In particular, we assessed the intracellular stability of each chain. Our results showed that the chains were markedly different in their susceptibility to intracellular degradation. TCR alpha and beta and CD3 delta were degraded rapidly, whereas CD3 gamma and epsilon were stable. An analysis of the N-linked oligosaccharides of the glycoprotein subunits suggested that the chains were unable to reach the medial Golgi during the metabolic chase. This was supported by immunofluorescence micrographs that showed both the stable CD3 gamma and unstable CD3 delta chain localized in the endoplasmic reticulum. To study the effects of subunit associations on intracellular transport we used cotransfection to reconstitute precise combinations of subunits. Associations between stable and unstable subunits expressed in the same cell led to the formation of stable complexes. These complexes were retained in or close to the endoplasmic reticulum. The results suggested that the intracellular transport of the T cell receptor could be regulated by two mechanisms. The TCR alpha and beta and CD3 delta subunits were degraded rapidly and as a consequence failed to reach the plasma membrane. CD3 gamma or epsilon were stable but were retained inside the cell. The results also demonstrated that there was an interplay between the two pathways such that the CD3 gamma and epsilon subunits were able to protect labile chains from rapid intracellular degradation. In this way, they could seed subunit assembly in or close to the endoplasmic reticulum and allow a stable receptor to form before its transport to the plasma membrane.

Stimulation of a major subset of lymphocytes expressing T cell receptor γδ by an antigen derived from mycobacterium tuberculosis

Cell ◽  
1989 ◽  
Vol 57 (4) ◽  
pp. 667-674 ◽  
Author(s):  
Rebecca L. O'Brien ◽  
Mary Pat Happ ◽  
Angela Dallas ◽  
Ed Palmer ◽  
Ralph Kubo ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
T Cell Receptor ◽  
Cell Receptor ◽  
Major Subset ◽  
Stimulation Of

scholarly journals Stimulation of T cells through the CD3/T-cell receptor complex: role of cytoplasmic calcium, protein kinase C translocation, and phosphorylation of pp60c-src in the activation pathway.

1987 ◽  
Vol 7 (2) ◽  
pp. 650-656 ◽  
Author(s):  
J A Ledbetter ◽  
L E Gentry ◽  
C H June ◽  
P S Rabinovitch ◽  
A F Purchio
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Solid Phase ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Jurkat Cells ◽  
Receptor Complex ◽  
Kinase C ◽  
Stimulation Of

Stimulation of T cells or the Jurkat T-cell line with soluble antibodies to the CD3/T-cell receptor complex causes mobilization of cytoplasmic Ca2+, which is blocked by pertussis toxin but not by ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid, and translocation of protein kinase C activity from the cytoplasm to the membrane. Such stimulation also causes phosphorylation of pp60c-src at an amino-terminal serine residue. These activities are consistent with induction of phosphatidylinositol metabolism after antibody binding. Anti-CD3 stimulation with antibody in solution, however, does not cause Jurkat cells to release interleukin 2 and blocks rather than induces proliferation of T cells. Induction of interleukin 2 production by Jurkat cells and proliferation by normal T cells requires anti-CD3 stimulation with antibody on a solid support, such as Sepharose beads or a plastic dish. Thus, we examined phosphorylation of pp60c-src after stimulation of Jurkat cells with anti-CD3 in solution or on solid phase. Both of these caused serine phosphorylation of pp60c-src that was indistinguishable even after 4 h of stimulation. These results indicate that the mode of anti-CD3 stimulation (in solution or on solid phase) controls a cellular function that modifies the consequences of signal transduction through phosphatidylinositol turnover.

scholarly journals Dual Fatty Acylation of p59Fyn Is Required for Association with the T Cell Receptor ζ Chain through Phosphotyrosine–Src Homology Domain-2 Interactions

1999 ◽  
Vol 145 (2) ◽  
pp. 377-389 ◽  
Author(s):  
Woutervan't Hof ◽  
Marilyn D. Resh
Keyword(s):  
Plasma Membrane ◽  
T Cell ◽  
T Cell Receptor ◽  
Cell Receptor ◽  
Insoluble Fraction ◽  
Sh2 Domain ◽  
Sh2 Domains ◽  
Fatty Acylation ◽  
Src Homology ◽  
Activation Motif

The first 10 residues within the Src homology domain (SH)–4 domain of the Src family kinase Fyn are required for binding to the immune receptor tyrosine-based activation motif (ITAM) of T cell receptor (TCR) subunits. Recently, mutation of glycine 2, cysteine 3, and lysines 7 and 9 was shown to block binding of Fyn to TCR ζ chain ITAMs, prompting the designation of these residues as an ITAM recognition motif (Gauen, L.K.T., M.E. Linder, and A.S. Shaw. 1996. J. Cell Biol. 133:1007–1015). Here we show that these residues do not mediate direct interactions with TCR ITAMs, but rather are required for efficient myristoylation and palmitoylation of Fyn. Specifically, coexpression of a K7,9A-Fyn mutant with N-myristoyltransferase restored myristoylation, membrane binding, and association with the cytoplasmic tail of TCR ζ fused to CD8. Conversely, treatment of cells with 2-hydroxymyristate, a myristoylation inhibitor, blocked association of wild-type Fyn with ζ. The Fyn NH2 terminus was necessary but not sufficient for interaction with ζ and both Fyn kinase and SH2 domains were required, directing phosphorylation of ζ ITAM tyrosines and binding to ζ ITAM phosphotyrosines. Fyn/ζ interaction was sensitive to octylglucoside and filipin, agents that disrupt membrane rafts. Moreover, a plasma membrane bound, farnesylated Fyn construct, G2A,C3S-FynKRas, was not enriched in the detergent insoluble fraction and did not associate with ζ. We conclude that the Fyn SH4 domain provides the signals for fatty acylation and specific plasma membrane localization, stabilizing the interactions between the Fyn SH2 domain and phosphotyrosines in TCR ζ chain ITAMs.

scholarly journals The T Cell Receptor Resides in Ordered Plasma Membrane Nanodomains that Aggregate Upon T Cell Activation

2015 ◽  
Vol 108 (2) ◽  
pp. 98a
Author(s):  
Ingela Parmryd ◽  
Astrid Riehl ◽  
Jelena Dinic ◽  
Jeremy Adler
Keyword(s):  
Plasma Membrane ◽  
T Cell ◽  
T Cell Receptor ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cell Receptor

scholarly journals ZAP-70 Association with T Cell Receptor ζ (TCRζ): Fluorescence Imaging of Dynamic Changes upon Cellular Stimulation

1998 ◽  
Vol 143 (3) ◽  
pp. 613-624 ◽  
Author(s):  
Joanne Sloan-Lancaster ◽  
John Presley ◽  
Jan Ellenberg ◽  
Tetsuo Yamazaki ◽  
Jennifer Lippincott-Schwartz ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
T Cell ◽  
Tyrosine Kinase ◽  
T Cell Receptor ◽  
Protein Tyrosine Kinase ◽  
Specific Interaction ◽  
Active Form ◽  
Lymphocyte Activation ◽  
Cell Receptor ◽  
Protein Tyrosine

The nonreceptor protein tyrosine kinase ZAP-70 is a critical enzyme required for successful T lymphocyte activation. After antigenic stimulation, ZAP-70 rapidly associates with T cell receptor (TCR) subunits. The kinetics of its translocation to the cell surface, the properties of its specific interaction with the TCRζ chain expressed as a chimeric protein (TTζ and Tζζ), and its mobility in different intracellular compartments were studied in individual live HeLa cells, using ZAP-70 and Tζζ fused to green fluorescent protein (ZAP-70 GFP and Tζζ–GFP, respectively). Time-lapse imaging using confocal microscopy indicated that the activation-induced redistribution of ZAP-70 to the plasma membrane, after a delayed onset, is of long duration. The presence of the TCRζ chain is critical for the redistribution, which is enhanced when an active form of the protein tyrosine kinase Lck is coexpressed. Binding specificity to TTζ was indicated using mutant ZAP-70 GFPs and a truncated ζ chimera. Photobleaching techniques revealed that ZAP-70 GFP has decreased mobility at the plasma membrane, in contrast to its rapid mobility in the cytosol and nucleus. Tζζ– GFP is relatively immobile, while peripherally located ZAP-70 in stimulated cells is less mobile than cytosolic ZAP-70 in unstimulated cells, a phenotype confirmed by determining the respective diffusion constants. Examination of the specific molecular association of signaling proteins using these approaches has provided new insights into the TCRζ–ZAP-70 interaction and will be a powerful tool for continuing studies of lymphocyte activation.

Establishment of T-cell Lines from Infiltrated NOD Islets by Stimulation of Specific T-Cell Receptor Vβ Segments

1993 ◽  
Vol 6 (4) ◽  
pp. 423-436 ◽  
Author(s):  
Didier Maugendre ◽  
Béatrice Legrand ◽  
Martine Olivi ◽  
Pierre Bedossa ◽  
Jean-François Bach ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Receptor ◽  
Cell Lines ◽  
Cell Receptor ◽  
T Cell Lines ◽  
T Cell Receptor Vβ ◽  
Stimulation Of

scholarly journals Thy-1 stimulation of mouse T cells induces a delayed T cell receptor-like signal that results in Ca2+-independent cytotoxicity

2017 ◽  
Vol 16 (4) ◽  
pp. 5683-5692 ◽  
Author(s):  
Suzanne Furlong ◽  
Melanie R. Power Coombs ◽  
David W. Hoskin
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Cell Receptor ◽  
Stimulation Of

scholarly journals Spatiotemporal Regulation of Signaling: Focus on T Cell Activation and the Immunological Synapse

2020 ◽  
Vol 21 (9) ◽  
pp. 3283
Author(s):  
Esther Garcia ◽  
Shehab Ismail
Keyword(s):  
Signal Transduction ◽  
Plasma Membrane ◽  
T Cell ◽  
T Cell Receptor ◽  
T Cell Activation ◽  
Cell Activation ◽  
Immunological Synapse ◽  
Cell Receptor ◽  
Signaling Network ◽  
Spatiotemporal Regulation

In a signaling network, not only the functions of molecules are important but when (temporal) and where (spatial) those functions are exerted and orchestrated is what defines the signaling output. To temporally and spatially modulate signaling events, cells generate specialized functional domains with variable lifetime and size that concentrate signaling molecules, enhancing their transduction potential. The plasma membrane is a key in this regulation, as it constitutes a primary signaling hub that integrates signals within and across the membrane. Here, we examine some of the mechanisms that cells exhibit to spatiotemporally regulate signal transduction, focusing on the early events of T cell activation from triggering of T cell receptor to formation and maturation of the immunological synapse.

scholarly journals T Cell Receptor (TCR) Interacting Molecule (TRIM), A Novel Disulfide-linked Dimer Associated with the TCR–CD3–ζ Complex, Recruits Intracellular Signaling Proteins to the Plasma Membrane

1998 ◽  
Vol 188 (3) ◽  
pp. 561-575 ◽  
Author(s):  
Eddy Bruyns ◽  
Anne Marie-Cardine ◽  
Henning Kirchgessner ◽  
Karin Sagolla ◽  
Andrej Shevchenko ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Amino Acid ◽  
T Cell ◽  
T Cell Receptor ◽  
Intracellular Signaling ◽  
Adaptor Protein ◽  
Cell Receptor ◽  
Sh2 Domain ◽  
Pi3 Kinase ◽  
Signaling Proteins

The molecular mechanisms regulating recruitment of intracellular signaling proteins like growth factor receptor–bound protein 2 (Grb2), phospholipase Cγ1, or phosphatidylinositol 3-kinase (PI3-kinase) to the plasma membrane after stimulation of the T cell receptor (TCR)– CD3–ζ complex are not very well understood. We describe here purification, tandem mass spectrometry sequencing, molecular cloning, and biochemical characterization of a novel transmembrane adaptor protein which associates and comodulates with the TCR–CD3–ζ complex in human T lymphocytes and T cell lines. This protein was termed T cell receptor interacting molecule (TRIM). TRIM is a disulfide-linked homodimer which is comprised of a short extracellular domain of 8 amino acids, a 19–amino acid transmembrane region, and a 159–amino acid cytoplasmic tail. In its intracellular domain, TRIM contains several tyrosine-based signaling motifs that could be involved in SH2 domain–mediated protein–protein interactions. Indeed, after T cell activation, TRIM becomes rapidly phosphorylated on tyrosine residues and then associates with the 85-kD regulatory subunit of PI3-kinase via an YxxM motif. Thus, TRIM represents a TCR-associated transmembrane adaptor protein which is likely involved in targeting of intracellular signaling proteins to the plasma membrane after triggering of the TCR.

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