Molecular Interaction between the Fyn-associated Protein SKAP55 and the SLP-76-associated Phosphoprotein SLAP-130

Marie‐Cardine, Anne; Hendricks-Taylor, L. Ranee; Boerth, Nancy J.; Zhao, Hang; Schraven, Burkhart; Koretzky, Gary A.

doi:10.1074/jbc.273.40.25789

Cited by 89 publications

(121 citation statements)

References 26 publications

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“…ADAP itself has been directly linked to integrin activation, as ADAP -/-T cells do not adhere to fibronectin, ICAM, or VCAM following TCR ligation [31,32]. One model for how a SLP-76/ ADAP association may lead to integrin activation is through Src kinase-associated phosphoprotein of 55 kDa, an adaptor that is constitutively associated with ADAP [52] and has been linked to TCR-induced integrin activation [53,54].…”

Section: Discussionmentioning

confidence: 99%

In vivo disruption of T cell development by expression of a dominant‐negative polypeptide designed to abolish the SLP‐76/Gads interaction

Jordan¹,

Kliche²,

Shabason³

et al. 2007

Eur J Immunol

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Multi-molecular complexes nucleated by adaptor proteins play a central role in signal transduction. In T cells, one central axis consists of the assembly of several signaling proteins linked together by the adaptors linker of activated T cells (LAT), Src homology 2 domain-containing leukocyte-specific phosphoprotein of 76 kDa (SLP-76), and Grb2-related adaptor downstream of Shc (Gads). Each of these adaptors has been shown to be important for normal T cell development, and their proper sub-cellular localization is critical for optimal function in cell lines. We previously demonstrated in Jurkat T cells and a rat basophilic leukemic cell line that expression of a 50-amino acid polypeptide identical to the site on SLP-76 that binds to Gads blocks proper localization of SLP-76 and SLP-76-dependent signaling events. Here we extend these studies to investigate the ability of this polypeptide to inhibit TCR-induced integrin activity in Jurkat cells and to inhibit in vivo thymocyte development and primary T cell function. These data provide evidence for the in vivo function of a dominant-negative peptide based upon the biology of SLP-76 action and suggest the possibility of therapeutic potential of targeting the SLP-76/Gads interaction. IntroductionStimulation of T cells through the T cell receptor (TCR) leads to recruitment and localization of several critical signaling proteins, which results in the generation of a macro-molecular signaling complex. Key components of this complex, or signalosome, include the adaptors Src homology 2 (SH2) domain-containing leukocyte-specific phosphoprotein of 76 kDa (SLP-76), and Grb2-related adaptor downstream of Shc (Gads), which serves to link SLP-76 and its associated molecules to the transmembrane adaptor linker of activated T cells (LAT) [1].As an adaptor protein, SLP-76 possesses no enzymatic activity but functions via interaction with multiple proteins by way of three structural domains [2][3][4]. The N terminus harbors three tyrosines that become phos- phorylated upon TCR engagement [5,6] and have been reported to serve as docking sites for the guanine nucleotide exchange factor Vav, the adaptor protein Nck, the Tec family kinase Itk, and the p85 subunit of PI3K [7][8][9][10][11][12][13][14][15][16]. A single SH2 domain resides in the C-terminal portion of SLP-76 and links it to the adhesion and degranulation-promoting adaptor protein (ADAP) and to the hematopoietic progenitor kinase 1 [17][18][19]. Hematopoietic progenitor kinase 1, in turn, has been shown to phosphorylate SLP-76, creating a binding site for 14-3-3, a negative regulator of TCR signaling [20,21]. The central proline-rich region binds phospholipase Cc- Targeted deletion of SLP-76 in mice results in an inability of thymocytes to progress past the CD4 -CD8 -double-negative (DN) stage of development and thus a complete absence of mature T cells [28,29]. Since T cells do not develop in the absence of SLP-76, much of our knowledge of the role of SLP-76 in mature TCR signaling has been derived from the study of J14 ...

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Section: Discussionmentioning

confidence: 99%

In vivo disruption of T cell development by expression of a dominant‐negative polypeptide designed to abolish the SLP‐76/Gads interaction

Jordan¹,

Kliche²,

Shabason³

et al. 2007

Eur J Immunol

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“…The SH3 domain of SKAP55 (Src-kinase associated phosphoprotein of 55 kDa) interacts with a proline-rich sequence (PRS) stretch in ADAP (Fig. 1A) (10,11), whereas another PRS (FPPPP) is responsible for the interaction with the actin regulator Ena/ VASP-like protein (EVL) (12). Membrane binding of the ADAP-SKAP55 complex is conferred by the PH domain of SKAP55 and to a lower extend by the C-terminal hSH3 domain (hSH3 C ) of ADAP (13)(14)(15)(16).…”

Section: Stimulation Of T Cells Leads To Distinct Changes Of Theirmentioning

confidence: 99%

“…4A, TCR stimulation as well as CXCL12 treatment of Jurkat T cells led to an inducible association of ADAP and ZAP70. The functional integrity of ADAP is indicated by coprecipitation of SKAP55, a constitutive binding partner of ADAP (10). To address the question of whether the interaction of ADAP and ZAP70 is mediated by ADAP-Y571, we made use of a suppression/re-expression vector system that allowed shRNA-mediated suppression of endogenous ADAP and simultaneous re-expression of Flag-tagged wildtype (WT) or ADAP-Y571F mutant (Fig.…”

Section: The Adap and Zap70 Interaction Is Inducible In A Cellularmentioning

confidence: 99%

Analysis of Phosphorylation-dependent Protein Interactions of Adhesion and Degranulation Promoting Adaptor Protein (ADAP) Reveals Novel Interaction Partners Required for Chemokine-directed T cell Migration

Kuropka

Witte

Sticht

et al. 2015

Molecular & Cellular Proteomics

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Stimulation of T cells leads to distinct changes of theirT cell migration and the establishment of productive T cell/APC interactions are regulated by the activity of integrins. In resting T cells, integrins are expressed in an inactive state that adopts a conformation with low affinity for their ligands. Members of the intercellular adhesion molecule family (ICAM 1-5) are the physiological ligands of lymphocyte functionassociated antigen 1 (LFA-1, ␣L␤2-integrin) whereas vascular cell adhesion molecule (VCAM) and fibronectin are the ligands for the ␤1-integrin very late antigen 4 (VLA-4) (1, 2). Triggering of the T cell receptor (TCR) by peptide-major histocompati-

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“…Via its own proline-rich motif, FYB associates with the adapter proteins SKAP1/SKAP55 (Src kinase-associated phosphoprotein 1/of 55 kDa) or SKAP2/ SKAP55R (Src kinase-associated phosphoprotein 2/of 55 kDa-related protein). 24,25 The formed protein complex allows for membrane-recruitment of the small Ras-like GTPase Rap1 which then triggers integrin activation required for lymphocyte extravasation as well as formation of ISs. 26 Accordingly, FYB reorients towards the target cell and like Nck is enriched in the proximity of the IS.…”

Section: Identification Of Nck Interaction Partnersmentioning

confidence: 99%

The adapter protein Nck: Role of individual SH3 and SH2 binding modules for protein interactions in T lymphocytes

et al. 2010

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Nck is a ubiquitously expressed, primarily cytosolic adapter protein consisting of one SH2 domain and three SH3 domains. It links receptor and nonreceptor tyrosine kinases to actin cytoskeleton reorganizing proteins. In T lymphocytes, Nck is a crucial component of signaling pathways for T cell activation and effector function. It recruits actin remodeling proteins to T cell receptor (TCR)-associated activation clusters and thereby initiates changes in cell polarity and morphology. Moreover, Nck is crucial for the TCR-induced mobilization of secretory vesicles to the cytotoxic immunological synapse. To identify the interactome of Nck in human T cells, we performed a systematic screen for interaction partners in untreated or pervanadate-treated cells. We used GST fusion proteins containing full length Nck, the combined SH3 domains or the individual SH3 and SH2 domains to precipitate putative Nck interactors from cellular lysates. Protein bands were excised from gels, processed by tryptic in-gel digestion and analyzed by mass spectrometry. Using this approach, we confirmed previously established interactions (e.g., with Slp76, CD3e, WASP, and WIPF1) and identified several novel putative Nck-binding proteins. We subsequently verified the SH2 domain binding to the actin-binding protein HIP55 and to FYB/ADAP, and the SH3-mediated binding to the nuclear proteins SFPQ/NONO. Using laser scanning microscopy, we provide new evidence for a nuclear localization of Nck in human T cells. Our data highlight the fundamental role of Nck in the TCR-to-cytoskeleton crosstalk and point to yet unknown nuclear functions of Nck also in T lymphocytes.

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Molecular Interaction between the Fyn-associated Protein SKAP55 and the SLP-76-associated Phosphoprotein SLAP-130

Cited by 89 publications

References 26 publications

In vivo disruption of T cell development by expression of a dominant‐negative polypeptide designed to abolish the SLP‐76/Gads interaction

In vivo disruption of T cell development by expression of a dominant‐negative polypeptide designed to abolish the SLP‐76/Gads interaction

Analysis of Phosphorylation-dependent Protein Interactions of Adhesion and Degranulation Promoting Adaptor Protein (ADAP) Reveals Novel Interaction Partners Required for Chemokine-directed T cell Migration

The adapter protein Nck: Role of individual SH3 and SH2 binding modules for protein interactions in T lymphocytes

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