Quantitative phosphoproteomics reveals SLP-76 dependent regulation of PAG and Src family kinases in T cells

• Published in: PloS one 2012-10, Vol.7 (10), p.e46725
• Main Authors: Cao, Lulu, Ding, Yiyuan, Hung, Norris, Yu, Kebing, Ritz, Anna, Raphael, Benjamin J, Salomon, Arthur R
• Format: Article
• Language: English
• Subjects: 1-Phosphatidylinositol 3-kinase; Adapter proteins; Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - metabolism; Amino Acid Motifs; Biochemistry; Biology; Cell adhesion & migration; Cell Line; Computer science; Cytokines; Extracellular signal-regulated kinase; Fyn protein; Humans; Intracellular Signaling Peptides and Proteins - metabolism; Itk protein; Kinases; Kinetics; Labeling; Lck protein; Leukocytes; Lymphocytes; Lymphocytes T; Membrane Proteins - metabolism; Molecular biology; Mutagenesis; Negative feedback; Peptides; Phosphatase; Phosphoproteins - genetics; Phosphoproteins - metabolism; Phosphorylation; Phosphotransferases; Protein Processing, Post-Translational; Proteomics; Receptors, Antigen, T-Cell - metabolism; Regulations; Scaffolding; Signal Transduction; Signaling; Site-directed mutagenesis; SLP-76 protein; Src protein; src-Family Kinases - metabolism; T cell receptors; T cells; T-cell receptor; T-Lymphocytes - metabolism; Tyrosine; White blood cells
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0046725

The SH2-domain-containing leukocyte protein of 76 kDa (SLP-76) plays a critical scaffolding role in T cell receptor (TCR) signaling. As an adaptor protein that contains multiple protein-binding domains, SLP-76 interacts with many signaling molecules and links proximal receptor stimulation to downstream effectors. The function of SLP-76 in TCR signaling has been widely studied using the Jurkat human leukaemic T cell line through protein disruption or site-directed mutagenesis. However, a wide-scale characterization of SLP-76-dependant phosphorylation events is still lacking. Quantitative profiling of over a hundred tyrosine phosphorylation sites revealed new modes of regulation of phosphorylation of PAG, PI3K, and WASP while reconfirming previously established regulation of Itk, PLCγ, and Erk phosphorylation by SLP-76. The absence of SLP-76 also perturbed the phosphorylation of Src family kinases (SFKs) Lck and Fyn, and subsequently a large number of SFK-regulated signaling molecules. Altogether our data suggests unique modes of regulation of positive and negative feedback pathways in T cells by SLP-76, reconfirming its central role in the pathway.