title: A Comprehensive Immunoreceptor Phosphotyrosine-based Signaling Network Revealed by Reciprocal Protein-Peptide Array Screening.
abstract: Cells of the immune system communicate with their environment through immunoreceptors. These receptors often harbor intracellular tyrosine residues, which, when phosphorylated upon receptor activation, serve as docking sites to recruit downstream signaling proteins containing the Src Homology 2 (SH2) domain. A systematic investigation of interactions between the SH2 domain and the immunoreceptor tyrosine-based regulatory motifs (ITRM), including inhibitory (ITIM), activating (ITAM), or switching (ITSM) motifs, is critical for understanding cellular signal transduction and immune function. Using the B cell inhibitory receptor CD22 as an example, we developed an approach that combines reciprocal or bidirectional phosphopeptide and SH2 domain array screens with in-solution binding assays to identify a comprehensive SH2-CD22 interaction network. Extending this approach to 194 human ITRM sequences and 78 SH2 domains led to the identification of a high-confidence immunoreceptor interactome containing 1137 binary interactions. Besides recapitulating many previously reported interactions, our study uncovered numerous novel interactions. The resulting ITRM-SH2 interactome not only helped to fill many gaps in the immune signaling network, it also allowed us to associate different SH2 domains to distinct immune functions. Detailed analysis of the NK cell ITRM-mediated interactions led to the identification of a network nucleated by the Vav3 and Fyn SH2 domains. We showed further that these SH2 domains have distinct functions in cytotoxicity. The bidirectional protein-peptide array approach described herein may be applied to the numerous other peptide-binding modules to identify potential protein-protein interactions in a systematic and reliable manner.
authors.affiliation: From the ‡Department of Biochemistry and the Siebens-Drake Medical Research Institute, Schulich School of Medicine and Dentistry, Western University, London, Ontario N6A 5C1;
authors.forename: Huadong
authors.initials: H
authors.lastname: Liu
authors.affiliation: From the ‡Department of Biochemistry and the Siebens-Drake Medical Research Institute, Schulich School of Medicine and Dentistry, Western University, London, Ontario N6A 5C1;
authors.forename: Lei
authors.initials: L
authors.lastname: Li
authors.affiliation: From the ‡Department of Biochemistry and the Siebens-Drake Medical Research Institute, Schulich School of Medicine and Dentistry, Western University, London, Ontario N6A 5C1;
authors.forename: Courtney
authors.initials: C
authors.lastname: Voss
authors.affiliation: §Department of Chemistry, University of Waterloo, Waterloo, ON, N2L 3G1, Canada.
authors.forename: Feng
authors.initials: F
authors.lastname: Wang
authors.affiliation: §Department of Chemistry, University of Waterloo, Waterloo, ON, N2L 3G1, Canada.
authors.forename: Juewen
authors.initials: J
authors.lastname: Liu
authors.affiliation: From the ‡Department of Biochemistry and the Siebens-Drake Medical Research Institute, Schulich School of Medicine and Dentistry, Western University, London, Ontario N6A 5C1; sli@uwo.ca.
authors.forename: Shawn Shun-Cheng
authors.initials: SS
authors.lastname: Li
mesh_terms: D020816:Amino Acid Motifs; D006801:Humans; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D019000:Phosphotyrosine / Q000378:metabolism*; D040081:Protein Array Analysis; D011485:Protein Binding; D011971:Receptors, Immunologic / Q000378:metabolism*; D015398:Signal Transduction; D020298:U937 Cells; D018909:src Homology Domains
chemical_list: D010455:Peptides; D011971:Receptors, Immunologic; D019000:Phosphotyrosine