Specificity of the SH2 domains of SHP-1 in the interaction with the immunoreceptor tyrosine-based inhibitory motif-bearing receptor gp49B

Inhibitory receptors on hemopoietic cells critically regulate cellular func tion, Despite their expression on a variety of cell types, these inhibitory receptors signal through a common mechanism involving tyrosine phosphoryla tion of the immunoreceptor tyrosine-based inhibitory motif (ITIM), which en gages Src homology 2 (SH2) domain-containing cytoplasmic tyrosine or inosit ol phosphatases. In this study, we have investigated the proximal signal-tr ansduction pathway of an ITIM-bearing receptor, gp49B, a member of a newly described family of murine NK and mast cell receptors, We demonstrate that the tyrosine residues within the ITIMs are phosphorylated and serve for the association and activation of the cytoplasmic tyrosine phosphatase SHP-1. Furthermore, we demonstrate a physiologic association between gp49B and SHP -1 by coimmunoprecipitation studies from NK cells, To address the mechanism of binding between gp49B and SHP-1, binding studies involving glutathione S-transferase SHP-1 mutants were performed. Utilizing the tandem SH2 domain s of SHP-1, we show that either SH2 domain ran interact with phosphorylated gp49B, Full-length SHP-1, with an inactivated amino SH2 domain, also retai ned gp49B binding. However, binding to gp49B was disrupted by inactivation of the carboxyl SH2 domain of full-length SHP-1, suggesting that in the pre sence of the phosphatase domain, the carboxyl SH2 domain is required for th e recruitment of phosphorylated gp49B. Thus, gp49B signaling involves SHP-1 , and this association is dependent on tyrosine phosphorylation of the gp49 B ITIMs, and an intact SHP-1 carboxyl SH2 domain.