scholarly journals T cell receptor ligation induces the formation of dynamically regulated signaling assemblies

2002 ◽  
Vol 158 (7) ◽  
pp. 1263-1275 ◽  
Author(s):  
Stephen C. Bunnell ◽  
David I. Hong ◽  
Julia R. Kardon ◽  
Tetsuo Yamazaki ◽  
C. Jane McGlade ◽  
...  
Keyword(s):  
T Cells ◽  
Lipid Raft ◽  
Synapse Formation ◽  
Fluorescent Protein ◽  
Yellow Fluorescent Protein ◽  
Cell Receptor ◽  
Rich Cluster ◽  
Enhanced Yellow Fluorescent Protein ◽  
Immune Synapse ◽  
Signaling Complex

Tcell antigen receptor (TCR) ligation initiates tyrosine kinase activation, signaling complex assembly, and immune synapse formation. Here, we studied the kinetics and mechanics of signaling complex formation in live Jurkat leukemic T cells using signaling proteins fluorescently tagged with variants of enhanced GFP (EGFP). Within seconds of contacting coverslips coated with stimulatory antibodies, T cells developed small, dynamically regulated clusters which were enriched in the TCR, phosphotyrosine, ZAP-70, LAT, Grb2, Gads, and SLP-76, excluded the lipid raft marker enhanced yellow fluorescent protein–GPI, and were competent to induce calcium elevations. LAT, Grb2, and Gads were transiently associated with the TCR. Although ZAP-70–containing clusters persisted for more than 20 min, photobleaching studies revealed that ZAP-70 continuously dissociated from and returned to these complexes. Strikingly, SLP-76 translocated to a perinuclear structure after clustering with the TCR. Our results emphasize the dynamically changing composition of signaling complexes and indicate that these complexes can form within seconds of TCR engagement, in the absence of either lipid raft aggregation or the formation of a central TCR-rich cluster.

scholarly journals Identification of antigen-presenting dendritic cells in mouse aorta and cardiac valves

2009 ◽  
Vol 206 (3) ◽  
pp. 497-505 ◽  
Author(s):  
Jae-Hoon Choi ◽  
Yoonkyung Do ◽  
Cheolho Cheong ◽  
Hyein Koh ◽  
Silvia B. Boscardin ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Fluorescent Protein ◽  
Yellow Fluorescent Protein ◽  
Human Aorta ◽  
Protein Antigens ◽  
Cardiac Valves ◽  
Aortic Sinus ◽  
Enhanced Yellow Fluorescent Protein ◽  
Antigen Presenting

Presumptive dendritic cells (DCs) bearing the CD11c integrin and other markers have previously been identified in normal mouse and human aorta. We used CD11c promoter–enhanced yellow fluorescent protein (EYFP) transgenic mice to visualize aortic DCs and study their antigen-presenting capacity. Stellate EYFP+ cells were readily identified in the aorta and could be double labeled with antibodies to CD11c and antigen-presenting major histocompatability complex (MHC) II products. The DCs proved to be particularly abundant in the cardiac valves and aortic sinus. In all aortic locations, the CD11c+ cells localized to the subintimal space with occasional processes probing the vascular lumen. Aortic DCs expressed little CD40 but expressed low levels of CD1d, CD80, and CD86. In studies of antigen presentation, DCs selected on the basis of EYFP expression or binding of anti-CD11c antibody were as effective as DCs similarly selected from the spleen. In particular, the aortic DCs could cross-present two different protein antigens on MHC class I to CD8+ TCR transgenic T cells. In addition, after intravenous injection, aortic DCs could capture anti-CD11c antibody and cross-present ovalbumin to T cells. These results indicate that bona fide DCs are a constituent of the normal aorta and cardiac valves.

pH dependence of the fluorescence lifetime of enhanced yellow fluorescent protein in solution and cells

2012 ◽  
Vol 235 ◽  
pp. 65-71 ◽  
Author(s):  
Takakazu Nakabayashi ◽  
Shugo Oshita ◽  
Ryoya Sumikawa ◽  
Fan Sun ◽  
Masataka Kinjo ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Fluorescent Protein ◽  
Ph Dependence ◽  
Yellow Fluorescent Protein ◽  
Enhanced Yellow Fluorescent Protein

The costimulatory activity of Tim-3 requires Akt and MAPK signaling and its recruitment to the immune synapse

2021 ◽  
Vol 14 (687) ◽  
pp. eaba0717
Author(s):  
Shunsuke Kataoka ◽  
Priyanka Manandhar ◽  
Judong Lee ◽  
Creg J. Workman ◽  
Hridesh Banerjee ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Transmembrane Domain ◽  
Transmembrane Protein ◽  
Cell Receptor ◽  
Mapk Signaling ◽  
Inhibitory Protein ◽  
Immune Synapse

Expression of the transmembrane protein Tim-3 is increased on dysregulated T cells undergoing chronic activation, including during chronic infection and in solid tumors. Thus, Tim-3 is generally thought of as an inhibitory protein. We and others previously reported that under some circumstances, Tim-3 exerts paradoxical costimulatory activity in T cells (and other cells), including enhancement of the phosphorylation of ribosomal S6 protein. Here, we examined the upstream signaling pathways that control Tim-3–mediated increases in phosphorylated S6 in T cells. We also defined the localization of Tim-3 relative to the T cell immune synapse and its effects on downstream signaling. Recruitment of Tim-3 to the immune synapse was mediated exclusively by the transmembrane domain, replacement of which impaired the ability of Tim-3 to costimulate T cell receptor (TCR)–dependent S6 phosphorylation. Furthermore, enforced localization of the Tim-3 cytoplasmic domain to the immune synapse in a chimeric antigen receptor still enabled T cell activation. Together, our findings are consistent with a model whereby Tim-3 enhances TCR-proximal signaling under acute conditions.

scholarly journals Genetic and transgenic reagents for Drosophila simulans, D. mauritiana, D. yakuba, D. santomea and D. virilis

2016 ◽  
Author(s):  
David L. Stern ◽  
Justin Crocker ◽  
Yun Ding ◽  
Nicolas Frankel ◽  
Gretchen Kappes ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Integration Site ◽  
Yellow Fluorescent Protein ◽  
Model Systems ◽  
Enhanced Yellow Fluorescent Protein ◽  
Functional Studies ◽  
Site Specific Integration ◽  
Genetic Strains ◽  
Fluorescent Molecules ◽  
Integration Efficiency

AbstractSpecies of the Drosophila melanogaster species subgroup, including the species D. simulans, D. mauritiana, D. yakuba, and D. santomea, have long served as model systems for studying evolution. Studies in these species have been limited, however, by a paucity of genetic and transgenic reagents. Here we describe a collection of transgenic and genetic strains generated to facilitate genetic studies within and between these species. We have generated many strains of each species containing mapped piggyBac transposons including an enhanced yellow fluorescent protein gene expressed in the eyes and a phiC31 attP site-specific integration site. We have tested a subset of these lines for integration efficiency and reporter gene expression levels. We have also generated a smaller collection of other lines expressing other genetically encoded fluorescent molecules in the eyes and a number of other transgenic reagents that will be useful for functional studies in these species. In addition, we have mapped the insertion locations of 58 transposable elements in D. virilis that will be useful for genetic mapping studies.

scholarly journals Mechanisms for segregating T cell receptor and adhesion molecules during immunological synapse formation in Jurkat T cells

2007 ◽  
Vol 104 (51) ◽  
pp. 20296-20301 ◽  
Author(s):  
Y. Kaizuka ◽  
A. D. Douglass ◽  
R. Varma ◽  
M. L. Dustin ◽  
R. D. Vale
Keyword(s):  
T Cells ◽  
T Cell ◽  
Adhesion Molecules ◽  
T Cell Receptor ◽  
Synapse Formation ◽  
Immunological Synapse ◽  
Cell Receptor ◽  
Jurkat T Cells

scholarly journals Agent-Based Modeling of T Cell Receptor Cooperativity

2020 ◽  
Vol 21 (18) ◽  
pp. 6473
Author(s):  
Anastasios Siokis ◽  
Philippe A. Robert ◽  
Michael Meyer-Hermann
Keyword(s):  
T Cell ◽  
Synapse Formation ◽  
Immunological Synapse ◽  
Cell Receptor ◽  
Agent Based Model ◽  
Tcr Signaling ◽  
Immune Synapse ◽  
Agent Based ◽  
Affinity Modulation ◽  
Antigen Peptide

Immunological synapse (IS) formation is a key event during antigen recognition by T cells. Recent experimental evidence suggests that the affinity between T cell receptors (TCRs) and antigen is actively modulated during the early steps of TCR signaling. In this work, we used an agent-based model to study possible mechanisms for affinity modulation during IS formation. We show that, without any specific active mechanism, the observed affinity between receptors and ligands evolves over time and depends on the density of ligands of the antigen peptide presented by major histocompatibility complexes (pMHC) and TCR molecules. A comparison between the presence or absence of TCR–pMHC centrally directed flow due to F-actin coupling suggests that centripetal transport is a potential mechanism for affinity modulation. The model further suggests that the time point of affinity measurement during immune synapse formation is critical. Finally, a mathematical model of F-actin foci formation incorporated in the agent-based model shows that TCR affinity can potentially be actively modulated by positive/negative feedback of the F-actin foci on the TCR-pMHC association rate kon.

scholarly journals Pancreatic islet function in a transgenic mouse expressing fluorescent protein

2005 ◽  
Vol 186 (2) ◽  
pp. 333-341 ◽  
Author(s):  
Daniel Nyqvist ◽  
Göran Mattsson ◽  
Martin Köhler ◽  
Varda Lev-Ram ◽  
Arne Andersson ◽  
...  
Keyword(s):  
Pancreatic Islet ◽  
Glucose Homeostasis ◽  
Fluorescent Protein ◽  
Regional Blood Flow ◽  
Animal Health ◽  
Yellow Fluorescent Protein ◽  
Islet Function ◽  
Enhanced Yellow Fluorescent Protein ◽  
Isolated Pancreatic Islets ◽  
Arterial Blood

Pancreatic islet function and glucose homeostasis have been characterized in the transgenic YC-3.0 mouse, which expresses the yellow chameleon 3.0 (YC-3.0) protein under the control of the β-actin and the cytomegalovirus promoters. Fluorescence from the enhanced yellow fluorescent protein (EYFP), one part of the yellow chameleon protein, was used as a reporter of transgene expression. EYFP was expressed in different quantities throughout most cell types, including islet endocrine and stromal cells. No adverse effects of the transgene on animal health, growth or fertility were observed. Likewise, in vivo glucose homeostasis, mean arterial blood pressure and regional blood flow values were normal. Furthermore, the transgenic YC-3.0 mouse had a normal β-cell volume and mass as well as glucose-stimulated insulin release in vitro, compared with the C57BL/6 control mouse. Isolated islets from YC-3.0 animals continuously expressed the transgene and reversed hyperglycemia when transplanted under the renal capsule of alloxan-diabetic nude mice. We conclude that isolated pancreatic islets from YC-3.0 animals implanted into recipients without any EYFP expression, constitute a novel and versatile model for studies of islet engraftment.

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