Genome-wide Analysis, Computational Modeling and Competition Assay of the Intermolecular Interaction Profile Between Human SH2-ome and the Putative Phosphorylation Sites of PDGFRA C-terminal Tail

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Human platelet-derived growth factor receptor alpha (PDGFRA) is a central regulator of physiological fibroblast functions and also plays an important role in the pathological fibrosis of many otolaryngological diseases. Previous studies have primarily addressed the extracellular immunoglobulin-like domain repeats, transmembrane domain and split kinase domain of PDGFRA, while its carboxy-terminal tail (CT-tail) extends from the kinase domain, still remains largely unexplored to date. The CT-tail is composed of [Formula: see text]150 amino acids and exists in an intrinsically disordered state, containing six putative tyrosine phosphorylation sites that serve as the potential targets of phosphotyrosine-recognition Src homology 2 (SH2) domains to orchestrate downstream cell signaling networks. In this study, we characterized the intermolecular binding events of the six putative phosphotyrosine sites in PDGFRA’s CT-tail to all the 120 SH2 domains in the human genome, i.e. the human SH2-ome, consequently creating a domain-phosphopeptide interaction (DPI) profile. A number of promising binding partner hits were identified from the profile, which cover previously reported binders and also many new potential partners. We demonstrated that the SH2 domains of both NCK1 and NCK2 isoforms can effectively recognize and interact with their cognate pTyr958 site in a nonspecific manner. In contrast, the cognate pTyr944 site can only be selectively targeted by the second SH2 domain of PLCG1 isoform in all four phospholipase C-gamma (PLCG) SH2 domains, but, as might be expected, the PLCG1(2) SH2 domain cannot effectively interact with the NCK-cognate pTyr944 site. In addition, we also found that a natural phosphopeptide derived from the PDGFRB pTyr1021 site is a good binder of PLCG1(2) SH2 domain, which can competitively disrupt the native interaction between the domain and its cognate pTyr944 site in PDGFRA’s CT-tail with a high competitive potency.

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