Kindlin-3 is required for the stabilization of TCR-stimulated LFA-1:ICAM-1 bonds critical for lymphocyte arrest and spreading on dendritic cells

Blood ◽  
2011 ◽  
Vol 117 (26) ◽  
pp. 7042-7052 ◽  
Author(s):  
Sara W. Feigelson ◽  
Valentin Grabovsky ◽  
Eugenia Manevich-Mendelson ◽  
Ronit Pasvolsky ◽  
Ziv Shulman ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Leukocyte Adhesion ◽  
Lymphocyte Function ◽  
Leukocyte Adhesion Deficiency ◽  
Tcr Signaling ◽  
Normal Expression ◽  
Lymphocyte Motility ◽  
Tcr Activation

Kindlin-3 is a key lymphocyte function–associated antigen-1 (LFA-1) coactivator deleted in leukocyte adhesion deficiency-III (LAD-III). In the present study, we investigated the involvement of this adaptor in lymphocyte motility and TCR-triggered arrest on ICAM-1 or on dendritic cells (DCs). Kindlin-3–null primary T cells from a LAD-III patient migrated normally on the major lymph node chemokine CCL21 and engaged in normal TCR signaling. However, TCR activation of Kindlin-3–null T lymphocytes failed to trigger the robust LFA-1–mediated T-cell spreading on ICAM-1 and ICAM-1–expressing DCs that is observed in normal lymphocytes. Kindlin-3 was also essential for cytoskeletal anchorage of the LFA-1 heterodimer and for microclustering of LFA-1 within ventral focal dots of TCR-stimulated lymphocytes spread on ICAM-1. Surprisingly, LFA-1 on Kindlin-3–null lymphocytes migrating over CCL21 acquired normal expression of an epitope associated with the conformational activation of the key headpiece domain, β I. This activated LFA-1 was highly responsive to TCR-triggered ICAM-1–driven stop signals in normal T cells locomoting on CCL21, but not in their Kindlin-3–null T-cell counterparts. We suggest that Kindlin-3 selectively contributes to a final TCR-triggered outside-in stabilization of bonds generated between chemokine-primed LFA-1 molecules and cell-surface ICAM-1.

scholarly journals Monoclonal antibodies to LFA-1 and to CD4 inhibit the mixed leukocyte reaction after the antigen-dependent clustering of dendritic cells and T lymphocytes.

1987 ◽  
Vol 165 (5) ◽  
pp. 1403-1417 ◽  
Author(s):  
K Inaba ◽  
R M Steinman
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Cell Mediated Immunity ◽  
Lymphocyte Function ◽  
Mixed Leukocyte Reaction ◽  
Binding Assays ◽  
Lymphocyte Blastogenesis ◽  
And Function

T cell proliferation in response to many stimuli is known to occur in discrete clusters of dendritic cells (DC) and CD4+ helper lymphocytes. The role of lymphocyte function-associated antigen (LFA-1) and CD4 in the formation and function of these clusters has been evaluated in the mixed leukocyte reaction (MLR). By day 1 of the control MLR, most of the DC and responsive T cells are associated in discrete aggregates. Addition of anti-LFA-1 and CD4 reagents does not block DC-T aggregation but reduces the subsequent proliferative response by 80-90%. Anti-LFA-1 disassembles newly formed DC-T cell aggregates, whereas anti-CD4 inhibits blastogenesis without disrupting the cluster. Binding of DC to sensitized, antigen-specific CD4+ cells has been studied using lymphoblasts isolated at day 4 of the MLR. It has been shown previously that greater than 80% blasts rebind to DC in an antigen-specific fashion in rapid (10 min) binding assays. Antigen-dependent DC-T binding is blocked by anti-Ia but not by mAb to LFA-1 or CD4. However, the bound anti-CD4-coated lymphocytes are unable to release IL-2. Anti-LFA-1-coated T cells release IL-2 but are easily disaggregated after binding to DC. These findings lead to two conclusions. LFA-1 and CD4 are not involved in the initial steps whereby DC bind to T cells but exert an independent and subsequent role. LFA-1 acts to stabilize the DC-T cluster, while CD4 contributes to lymphocyte blastogenesis and IL-2 release. Because DC but not other presenting cells cluster unprimed lymphocytes, it seems likely that an antigen-independent mechanism distinct from LFA-1 and CD4 mediates aggregate formation at the onset of cell-mediated immunity.

The actin cloud induced by LFA-1–mediated outside-in signals lowers the threshold for T-cell activation

Blood ◽  
2006 ◽  
Vol 109 (1) ◽  
pp. 168-175 ◽  
Author(s):  
Jun-ichiro Suzuki ◽  
Sho Yamasaki ◽  
Jennifer Wu ◽  
Gary A. Koretzky ◽  
Takashi Saito
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Immunological Synapse ◽  
Adaptor Protein ◽  
Cell Receptor ◽  
Cloud Formation ◽  
Lymphocyte Function ◽  
Tcr Signaling

Abstract The dynamic rearrangement of the actin cytoskeleton plays critical roles in T-cell receptor (TCR) signaling and immunological synapse (IS) formation in T cells. Following actin rearrangement in T cells upon TCR stimulation, we found a unique ring-shaped reorganization of actin called the “actin cloud,” which was specifically induced by outside-in signals through lymphocyte function–associated antigen-1 (LFA-1) engagement. In T-cell–antigen-presenting cell (APC) interactions, the actin cloud is generated in the absence of antigen and localized at the center of the T-cell–APC interface, where it accumulates LFA-1 and tyrosine-phosphorylated proteins. The LFA-1–induced actin cloud formation involves ADAP (adhesion- and degranulation-promoting adaptor protein) phosphorylation, LFA-1/ADAP assembly, and c-Jun N-terminal kinase (JNK) activation, and occurs independent of TCR and its proximal signaling. The formation of the actin cloud lowers the threshold for subsequent T-cell activation. Thus, the actin cloud induced by LFA-1 engagement may serve as a possible platform for LFA-1–mediated costimulatory function for T-cell activation.

Somatic revertant mosaicism in a patient with leukocyte adhesion deficiency type 1

Blood ◽  
2006 ◽  
Vol 109 (3) ◽  
pp. 1182-1184 ◽  
Author(s):  
Yumi Tone ◽  
Taizo Wada ◽  
Fumie Shibata ◽  
Tomoko Toma ◽  
Yoko Hashida ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Leukocyte Adhesion ◽  
Somatic Mosaicism ◽  
Autosomal Recessive Disorder ◽  
Cell Receptor ◽  
Leukocyte Adhesion Deficiency ◽  
Deficiency Type

Abstract Leukocyte adhesion deficiency type 1 (LAD-1) is an autosomal recessive disorder caused by mutations in the ITGB2 (CD18) gene and characterized by recurrent severe infections, impaired pus formation, and defective wound healing. We describe an unusual case of severe phenotypic LAD-1 presenting with somatic mosaicism. The patient is a compound heterozygote bearing 2 different frameshift mutations that abrogate protein expression. However, CD18 expression was detected in a small proportion of T cells but was undetectable in granulocytes, monocytes, B cells, and natural killer (NK) cells. The T cells were not of maternal origin, lacked the paternal mutation, and showed a selective advantage in vivo. Molecular analysis using sorted CD18+ cells revealed them to be derived from a single CD8+ T cell carrying T-cell receptor VB22. These findings suggest that spontaneous in vivo reversion was responsible for the somatic mosaicism in our patient.

scholarly journals 752 Novel, orally administered HPK1 inhibitors demonstrate anti-tumor efficacy and enhanced immune response

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A786-A786
Author(s):  
Stefan Chmielewski ◽  
Maciej Kujawa ◽  
Eliza Zimolag ◽  
Michal Galezowski ◽  
Andrzej Gondela ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Small Molecule ◽  
Cell Function ◽  
Adaptor Protein ◽  
Negative Regulator ◽  
Tcr Signaling ◽  
T Cell Function ◽  
Tcr Activation

BackgroundHematopoietic progenitor kinase 1 (HPK1, MAP4K1) is emerging as a well-renowned, druggable target for T cell-based immunotherapies. HPK1 is a member of the serine/threonine MAP4K family, predominantly expressed in hematopoietic cell lineages and shown to be a negative regulator of the T cell receptor (TCR) signaling pathway. Upon TCR activation, HPK1 is recruited to the proximity of the cell membrane and phosphorylates an adaptor protein SLP-76 at the Ser376 residue which, in turn, abrogates TCR signaling. Other studies point to a potential role of HPK1 in T cell exhaustion as well as in functional re-programming of regulatory T cells. Moreover, mounting evidence suggest that HPK1 kinase activity suppresses the immune functions of a wide range of other immune cell subsets like B cells and dendritic cells. Taken together, these observations support small-molecule HPK1 inhibitors as an attractive modality in cancer immunotherapy either as single agents or in combination with immune checkpoint inhibitors.MethodsActivity of compounds against HPK1 and selected off- and anti-targets was assessed in biochemical assays. Phosphorylation of SLP-76 was measured either by flow cytometry or TR-FRET. Jurkat and primary T cells were activated and cultured in the presence of tested compounds and immunosuppressive agents. Impact on TCR selectivity and T cell function was measured by AlphaLISA and flow cytometry. Target engagement was measured in splenocytes of mice administered orally with tested compounds followed by IP injection of aCD3 antibody. Anti-tumor efficacy of HPK1 inhibitors was assessed in a syngeneic tumor model.ResultsRyvu's proprietary small molecule HPK1 inhibitors exhibit sub-nanomolar activity against human and mouse HPK1 proteins and good selectivity against other TCR pathway kinases. Tested compounds efficiently block phosphorylation of SLP-76 upon TCR engagement. TCR selectivity of Ryvu's inhibitors, measured as a ratio between CD69 and pSer376 SLP-76 inhibition, is on par or superior to reference molecules. Tested compounds are not only able to overcome PGE-2 induced resistance following TCR activation in human PBMCs, inducing elevated IL-2 release but also affect T cell function in co-culture assay. Developed molecules have favorable PK profiles, allowing for sustained target coverage in proposed dosing regimens and demonstrate efficacy in a mammary carcinoma syngeneic model.ConclusionsRyvu has developed potent and selective HPK1 inhibitors with favorable PK and PD profiles, whose activity in vitro translates to in vivo efficacy. Further preclinical work is warranted to select a lead candidate for IND-enabling studies and subsequently clinical studies across a variety of solid tumors.

scholarly journals Inhibition of T Cell Receptor Activation by Semi-Synthetic Sesquiterpene Lactone Derivatives and Molecular Modeling of Their Interaction with Glutathione and Tyrosine Kinase ZAP-70

Molecules ◽  
2019 ◽  
Vol 24 (2) ◽  
pp. 350 ◽  
Author(s):  
Andrei Khlebnikov ◽  
Igor Schepetkin ◽  
Anarkul Kishkentaeva ◽  
Zhanar Shaimerdenova ◽  
Gayane Atazhanova ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tyrosine Kinase ◽  
T Cell Receptor ◽  
Sesquiterpene Lactones ◽  
Cell Receptor ◽  
Receptor Activation ◽  
Tcr Signaling ◽  
Jurkat T Cells ◽  
Tcr Activation

A variety of natural compounds have been shown to modulate T cell receptor (TCR) activation, including natural sesquiterpene lactones (SLs). In the present studies, we evaluated the biological activity of 11 novel semi-synthetic SLs to determine their ability to modulate TCR activation. Of these compounds, α -epoxyarglabin, cytisinyl epoxyarglabin, 1 β ,10 α -epoxyargolide, and chloroacetate grosheimin inhibited anti-CD3-induced Ca2+ mobilization and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation in Jurkat T cells. We also found that the active SLs depleted intracellular glutathione (GSH) in Jurkat T cells, supporting their reactivity towards thiol groups. Because the zeta-chain associated tyrosine kinase 70 kDa (ZAP-70) is essential for TCR signaling and contains a tandem SH2 region that is highly enriched with multiple cysteines, we performed molecular docking of natural SLs and their semi-synthetic derivatives into the ZAP-70 binding site. The docking showed that the distance between the carbon atom of the exocyclic methylene group and the sulfur atom in Cys39 of the ZAP-70 tandem SH2 module was 3.04–5.3 Å for active compounds. Furthermore, the natural SLs and their derivatives could be differentiated by their ability to react with the Cys39 SH-group. We suggest that natural and/or semi-synthetic SLs with an α -methylene- γ -lactone moiety can specifically target GSH and the kinase site of ZAP-70 and inhibit the initial phases of TCR activation.

scholarly journals The GSK3β-β-catenin-TCF1 pathway improves naive T cell activation in old adults by upregulating miR-181a

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Zhongde Ye ◽  
Timothy M. Gould ◽  
Huimin Zhang ◽  
Jun Jin ◽  
Cornelia M. Weyand ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
The Elderly ◽  
Cell Receptor ◽  
Tcr Signaling ◽  
Activation Markers ◽  
Old Adults ◽  
Tcr Activation

AbstractMicroRNAs play an important role in the regulation of T cell development, activation, and differentiation. One of the most abundant microRNAs in lymphocytes is miR-181a, which controls T cell receptor (TCR) activation thresholds in thymic selection as well as in peripheral T cell responses. We previously found that miR-181a levels decline in T cells in the elderly. In this study, we identified TCF1 as a transcriptional regulator of pri-miR-181a. A decline in TCF1 levels in old individuals accounted for the reduced miR-181a expression impairing TCR signaling. Inhibition of GSK3ß restored expression of miR-181a by inducing TCF1 in T cells from old adults. GSK3ß inhibition enhanced TCR signaling to increase downstream expression of activation markers and production of IL-2. The effect involved the upregulation of miR-181a and the inhibition of DUSP6 expression. Thus, inhibition of GSK3ß can restore responses of old T cells by inducing miR-181a expression through TCF1.

scholarly journals THEMIS is a priming substrate of non-receptor tyrosine phosphatase PTPN6/SHP1 and plays dual roles during T cell development

2021 ◽  
Author(s):  
Jiali Zhang ◽  
Erwei Zuo ◽  
Minfang Song ◽  
Li Chen ◽  
Zhenzhou Jiang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Tyrosine Phosphatase ◽  
Cytokine Signaling ◽  
Tcr Signaling ◽  
Dual Roles ◽  
Thymic Development ◽  
Tcr Activation ◽  
Receptor Tyrosine Phosphatase

THEMIS plays an indispensable role in T cells, but its mechanism of action is highly controversial. Using the systematic proximity labeling methodology PEPSI, we identified THEMIS as an uncharacterized substrate for the phosphatase SHP1. Saturated mutagenesis analysis revealed that THEMIS phosphorylation at the evolutionally conserved Tyr34 residue was oppositely regulated by SHP1 and the kinase LCK. Like THEMIS-/- mice, THEMIS Y34F/Y34F knock-in mice showed a significant decrease in CD4 thymocytes and mature CD4 T cells, but a normal thymic development and peripheral homeostasis of CD8 T cells. Mechanistically, phosphorylated THEMIS induced by TCR activation acts as a "priming substrate" to bind SHP1 and convert its phosphatase activity from basal level to nearly fully activated level, ensuring an appropriate negative regulation of TCR signaling. However, cytokine signaling in CD8 T cells failed to elicit THEMIS Y34 phosphorylation, revealing both phosphorylation-dependent and -independent roles of THEMIS in controlling T cell maturation and expansion.

scholarly journals The interaction of dendritic cells and γδ T cells promotes the activation of γδ T cells in experimental autoimmune uveitis

2017 ◽  
Vol 10 (02) ◽  
pp. 1650042 ◽  
Author(s):  
Beibei Wang ◽  
Wei Lin ◽  
Jike Song ◽  
Xiaofeng Xie ◽  
Hongsheng Bi
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Γδ T Cells ◽  
Lymphocyte Function ◽  
Experimental Autoimmune Uveitis ◽  
Autoimmune Uveitis ◽  
Experimental Autoimmune

Uveitis is a severe inflammatory disease that can cause visual impairment. Recently, activated [Formula: see text] T cells were proved to play a central role in the development of experimental autoimmune uveitis (EAU). However, the mechanism underlying [Formula: see text] T-cell activation in EAU is incompletely known. In this study, we determined the percentage changes in and the phenotypes of [Formula: see text] T cells and dendritic cells (DCs) obtained from the spleens of immunized C57BL/6 (B6) mice, an animal model of EAU. We found that the number of [Formula: see text] T cells and DCs obviously increased during the inflammation phase of EAU (days 16–20 of our experiment), and that during this time, [Formula: see text] T cells expressed high levels of CD69 and the integrin lymphocyte function–associated antigen-1 (LFA-1) and secreted high levels of interleukin (IL)-17A. Moreover, DCs obtained during this phase expressed high levels of CD80, CD83, CD86, and intracellular cell adhesion molecule-1 (ICAM-1). Furthermore, we studied the interaction between DCs and [Formula: see text] T cells by using flow cytometry and confocal microscopy in order to determine whether DCs affected [Formula: see text] T-cell activation in vitro. Co-cultures of the two types of cells showed that DCs induced high levels of CD69, LFA-1, and IL-17A in [Formula: see text] T cells. Imaging studies revealed contact between the DCs and [Formula: see text] T cells. This interaction was mediated by the accumulation of ICAM-1 and LFA-1 at the interface of DCs-[Formula: see text] T cells. Thus, the activation of [Formula: see text] T cells in EAU was promoted by DCs interacting with [Formula: see text] T cells.

The mTOR Pathway Inhibits Expression of the Tumor Suppressors ELF4 and KLF4 Downstream of TCR Signaling to Induce T Cell Proliferation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3688-3688
Author(s):  
Takeshi Yamada ◽  
Kirsten Gierach ◽  
H. Daniel Lacorazza
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Cell Activation ◽  
Conflicts Of Interest ◽  
T Cell Proliferation ◽  
Tcr Signaling ◽  
Tcr Activation

Abstract Abstract 3688 Poster Board III-624 Quiescence of circulating naïve T cells is maintained by the transcription factors ELF4 and KLF4 downstream of T-cell receptor (TCR) signaling. Hence, loss of ELF4 leads to increased proliferation of CD8+ T cells in response to homeostatic and antigen driven stimuli (Yamada et al, Nature Immunology, 2009). The identification of signals that suppress this restraint of proliferation will aid to enhance immunological memory during vaccination and to better understand development of T-cell acute lymphoblastic leukemias. Consistent with lower threshold of activation by ELF4 deletion in unstimulated naïve T cells, we identified a significant downregulation of the dual-specificity phosphatases DUSP1 and DUSP5 in a global gene expression study, which was confirmed at a protein level. Consequently, Elf4−/− CD8 T cells showed sustained phosphorylation of Erk1/2 upon TCR activation. In addition, we found that the PD98059 and LY294002 inhibitors, but not Cyclosporin A, blocked inhibition of ELF4 transcription upon TCR activation independently of CD28 co-stimulation and signals emanating from IL-2R. Furthermore, rapamycin also prevented downregulation of ELF4 transcripts following T cell activation, suggesting that mTORC1 inhibits ELF4 transcription downstream of MAPK and PI3K/Akt pathways. We conclude that the transcription factor ELF4 sets a proliferation threshold in naïve T cells by activating DUSPs and that ELF4 suppression upon TCR activation is mediated by mTORC1 downstream of MAPK and PI3K/Akt pathways. Our findings provide important targets of this novel control of T cell proliferation to enhance immune response to vaccination and to prevent expansion of pre-leukemic clones in pediatric patients that fail to respond to current therapies. Disclosures: No relevant conflicts of interest to declare.

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