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 T cell receptor (TCR)– CD3–ζ complex are not very well understood. We describe here purification, tandem mass spectrometry sequencing, cloning, and biochemical characterization a novel transmembrane adaptor which associates comodulates with TCR–CD3–ζ in...

Abstract In vitro protein kinase assays of CD45 immunprecipitates prepared from digitonin lysates resting human T lymphocytes resulted in exclusive tyrosine phosphorylation a 32‐kDa (pp32). Reprecipitation the phosphorylated proteins and Western blot analysis whole immunoprecipitates employing antisera specifically directed at different kinases demonstrated that p56 lck was responsible for pp32. Since also pp32, present data strongly suggest functional complex is formed between CD45, Such...

In human T-lymphocytes the Src family protein tyrosine kinase p59(fyn) associates with three phosphoproteins of 43, 55, and 85 kDa (pp43, pp55, pp85). Employing a GST-Fyn-Src homology 2 (SH2) domain fusion pp55 was purified from lysates Jurkat T-cells. Molecular cloning cDNA reveals that gene codes for so far nondescribed polypeptide 359 amino acids comprises pleckstrin domain, C-terminal SH3 as well several potential phosphorylation sites, among which one fulfills criteria to bind Src-like...

Oxidative stress leads to T-cell hyporesponsiveness or death. The actin-binding protein cofilin is oxidized during oxidative stress, which provokes a stiff actin cytoskeleton and hyporesponsiveness. Here, we show that long-term translocation of into the mitochondria necrotic-like programmed cell death (PCD) in human T cells. Notably, mutants functionally mimic oxidation by single mutation at oxidation-sensitive cysteins (Cys-39 Cys-80) predominately localize within mitochondria. expression...

Formation of immune synapses (IS) between T cells and APC requires multiple rearrangements in the actin cytoskeleton selective receptor accumulation supramolecular activation clusters (SMAC). The inner cluster (central SMAC) contains TCR/CD3 complex. outer (peripheral integrin LFA-1 Talin. Molecular mechanisms selectively stabilizing receptors IS remained largely unknown. Here, we demonstrate that sustained clustering is a consequence combined activities actin-bundling protein L-plastin...

Abstract Background Tumor cell migration and metastasis require dynamic rearrangements of the actin cytoskeleton. Interestingly, F-actin cross-linking stabilizing protein L-plastin, originally described as a leukocyte specific protein, is aberrantly expressed in several non-hematopoietic malignant tumors. Therefore, it has been discussed tumor marker. However, systematic vivo analyses functional relevance L-plastin for were so far lacking. Methods We investigated expression phosphorylation...

T lymphocytes express several low molecular weight transmembrane adaptor proteins that recruit src homology (SH)2 domain–containing intracellular molecules to the cell membrane via tyrosine-based signaling motifs. We describe here a novel molecule of this group termed SIT (SHP2 interacting protein). is disulfide-linked homodimeric glycoprotein expressed in lymphocytes. After tyrosine phosphorylation by and possibly syk protein kinases recruits SH2 phosphatase SHP2 an immunoreceptor...

CD45, a leukocyte-specific protein tyrosine phosphatase involved in signal transduction, has previously been shown to associate with 32-kDa phosphoprotein human T-lymphocytes and T-lymphoma cell lines. The was purified its coding cDNA cloned. Since expression of the found be restricted B- it termed LPAP (lymphocyte phosphatase-associated phosphoprotein). exists two differentially phosphorylated forms resting c, both which undergo alterations during T-lymphocyte activation. Analysis mRNA...

T cell receptor (TCR)-interacting molecule (TRIM) is a recently identified transmembrane adaptor protein, which exclusively expressed in cells. Here we demonstrate that mature cells, TRIM preferentially interacts with the TCR via TCR-ζ chains and to lesser extent CD3-ε/γ heterodimer. Transient or stable overexpression of Jurkat cells results enhancement expression on surface elevated induction Ca2+ mobilization after activation. TRIM-mediated upregulation from inhibition spontaneous...

Activation of naïve T cells requires costimulation via TCR/CD3 plus accessory receptors, which enables the dynamic rearrangement actin cytoskeleton and immune synapse maturation. Signaling events induced following may thus be valuable targets for therapeutic immunosuppression. Phosphorylation actin-bundling protein L-plastin represents such a costimulatory signal in primary human cells. Phosphorylated has higher affinity toward F-actin. However, importance phosphorylation regulation upon...

Abstract Several antitumor therapies work by increasing reactive oxygen species (ROS) within the tumor micromilieu. Here, we reveal that L-plastin (LPL), an established marker, is reversibly regulated ROS-induced thiol oxidation on Cys101, which forms a disulfide bridge with Cys42. LPL reduction mediated Thioredoxin1 (TRX1) system, as shown TRX1 trapping, knockdown and blockade of reductase (TRXR1) auranofin. diminishes its actin-bundling capacity. Ratiometric imaging using LPL-roGFP-Orp1...

Chronic inflammation is a hallmark of chronic wounds and inflammatory skin diseases. Due to hyperactive prolonged triggered by proinflammatory immune cells, transitioning the repair healing phase halted. T cells may exacerbate milieu secreting cytokines.

Abstract A glutathione‐S‐transferase‐src‐homology domain 2 (GST‐SH2) fusion protein was employed to identify molecules interacting with the tyrosine kinase p59 fyn . Among several proteins which bound SH2 in lysates of human Jurkat T lymphocytes, α‐ and β‐tubulin were identified by N‐terminal sequencing. Further analysis established that α‐tubulin exists as a tyrosine‐phosphorylated cells, where it interacts , but not p56 lck By contrast, untransformed resting lymphocytes is detectable...

Abstract In human T lymphocytes a functional complex is formed between the protein tyrosine phosphatase CD45, kinase p56 lck and phospho‐protein, pp32, possible common substrate. Here we demonstrate that previously described pp32 composed of two distinct molecules (pp29 pp32) in both resting continuously proliferating lymphoma lines. Importantly, lymphocyte activation employing CD2 monoclonal antibodies (mAb), CD3 mAb or phorbol 12, 13 dibutyrate results loss pp29 from CD45/p56 molecular...

ABSTRACT Concentrations of chlorophyll a /freshweight (Chl FW) and photosynthetic pigments/chlorophyll were studied during one growing season in the current year's (CYN) last needles (LYN) from Norway spruce ( Picea abies (L.) Karst.) grown under natural or close‐to‐natural climate. Climate regimes differed active radiation (PAR), temperature T ) UV‐B radiation. Pigments not affected by but most differences between climate regimes, also seasonal variations within could be related to PAR ....

Cytotoxic T-cells (CTLs) play an important role in many immune-mediated inflammatory diseases. Targeting cytotoxicity of CTLs would allow to interfere with tissue destruction. Here we demonstrate that WF-10, a pro-oxidative compound, inhibits CTL-mediated cytotoxicity. WF-10 did not influence early steps target-cell killing, but impaired the ability detach from initial target cell and move second cell. This reduced serial killing was accompanied by stronger enrichment adhesion molecule LFA-1...

CD2-associated molecules were identified by means of in vitro kinase assays CD2 immunoprecipitates obtained from nontransformed human T lymphocytes lysed the detergent brij 58. Under these conditions was found to be associated with protein tyrosine kinases p56lck and p59fyn as well two low molecular weight phosphoproteins. The latter zeta epsilon chains, major signaling components CD/7 TcR complex. Importantly, induction cell unresponsiveness towards CD2-mediated stimuli CD3/T receptor (TcR)...

Oxidative stress can lead to T cell hyporesponsiveness. A reducing micromilieu (e.g. provided by dendritic cells) rescue cells from such oxidant-induced dysfunction. However, the effects on proteins leading restored activation remained unknown. One key molecule of is actin-remodeling protein cofilin, which dephosphorylated serine 3 upon costimulation and has an essential role in formation mature immune synapses between antigen-presenting cells. Cofilin spatiotemporally regulated; at plasma...

While several protein serine/threonine kinases control cytokine production by T cells, the roles of phosphatases are largely unexplored. Here, we analyzed involvement phosphatase 1α (PP1α) in synthesis following costimulation primary human cells. Small interfering RNA (siRNA)-mediated knockdown PP1α (PP1KD) or expression a dominant negative (D95N-PP1) drastically diminished interleukin-10 (IL-10) production. Focusing on key transcriptional activator IL-10, demonstrate that nuclear...

Use of chimeric antigen receptor (CAR) T cells to treat B cell lymphoma and leukemia has been remarkably successful. Unfortunately, the therapeutic efficacy CAR against solid tumors is very limited, with immunosuppression by pro-oxidative tumor microenvironment (TME) a major contributing factor. High levels reactive oxygen species are well-tolerated due their elevated expression antioxidant proteins; however, this not case for cells, which consequently become hypo-responsive. The aim study...