T cell specific adaptor protein (TSAd) promotes interaction of Nck with Lck and SLP-76 in T cells

Buday, László and Sundvold-Gjerstad, V. and Granum, S. and Koll, L. and Abrahamsen, G. (2015) T cell specific adaptor protein (TSAd) promotes interaction of Nck with Lck and SLP-76 in T cells. CELL COMMUNICATION AND SIGNALING, 13 (1). p. 31. ISSN 1478-811X


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Background: The Lck and Src binding adaptor protein TSAd (T cell specific adaptor) regulates actin polymerization in T cells and endothelial cells. The molecular details as to how TSAd regulates this process remain to be elucidated. Results: To identify novel interaction partners for TSAd, we used a scoring matrix-assisted ligand algorithm (SMALI), and found that the Src homology 2 (SH2) domain of the actin regulator Non-catalytic region of tyrosine kinase adaptor protein (Nck) potentially binds to TSAd phosphorylated on Tyr280 (pTyr280) and pTyr305. These predictions were confirmed by peptide array analysis, showing direct binding of recombinant Nck SH2 to both pTyr280 and pTyr305 on TSAd. In addition, the SH3 domains of Nck interacted with the proline rich region (PRR) of TSAd. Pull-down and immunoprecipitation experiments further confirmed the Nck-TSAd interactions through Nck SH2 and SH3 domains. In line with this Nck and TSAd co-localized in Jurkat cells as assessed by confocal microscopy and imaging flow cytometry. Co-immunoprecipitation experiments in Jurkat TAg cells lacking TSAd revealed that TSAd promotes interaction of Nck with Lck and SLP-76, but not Vav1. TSAd expressing Jurkat cells contained more polymerized actin, an effect dependent on TSAd exon 7, which includes interactions sites for both Nck and Lck. Conclusions: TSAd binds to and co-localizes with Nck. Expression of TSAd increases both Nck-Lck and Nck-SLP-76 interaction in T cells. Recruitment of Lck and SLP-76 to Nck by TSAd could be one mechanism by which TSAd promotes actin polymerization in activated T cells. © 2015 Hem et al.

Item Type: Article
Uncontrolled Keywords: T lymphocyte activation; T lymphocyte; protein protein interaction; protein processing; protein phosphorylation; protein localization; PROTEIN FUNCTION; protein expression; protein domain; Protein Binding; proline rich protein domain; priority journal; nonhuman; MOUSE; jurkat cell line; human cell; human; Flow Cytometry; embryo; controlled study; confocal microscopy; CHROMATIN IMMUNOPRECIPITATION; carboxy terminal sequence; binding affinity; ARTICLE; animal cell; amino terminal sequence; ACTIN POLYMERIZATION; actin filament; Vav1 protein; Vav protein; unclassified drug; SLP 76 protein; protein SH3; protein SH2; protein kinase Lck; Nck protein; TSAd; T cell specific adaptor protein; SLP-76; SH2D2A; SH2 domain; NCK; LCK; adaptor protein
Subjects: R Medicine / orvostudomány > R1 Medicine (General) / orvostudomány általában
Depositing User: MTMT SWORD
Date Deposited: 31 Aug 2015 20:23
Last Modified: 31 Aug 2015 20:23

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