Recruitment of SHP-2 by FcRL6 is dependent on tyrosine 371. (A) IP/Western analysis of pervanadate-treated primary NK cells shows phosphotyrosines in samples immunoprecipitated with anti-FcRL6 antibody, but not anti-CD56 control. (B) Phosphotyrosine-specific antibodies detected a strong 66-kDa band in immunoprecipitates from pervanadate-treated (PV), but not untreated (UT), NK92 cells stably expressing wild-type FcRL6. In addition, SHP-2 is recruited by FcRL6 in pervanadate-treated cells. Mutation of tyrosine 371, but not tyrosine 356, to phenylalanine leads to an abrogation of FcRL6 phosphorylation and a failure to recruit SHP-2. ERK immunoblot is shown as loading control. This result has been repeated twice. (C) Recruitment of SHP-2 by phosphorylated NKG2A in pervanadate-treated NK92 cells is shown as positive control. NKG2A recruits more SHP-2 than FcRL6, suggesting low stoichiometry of interaction between FcRL6 and SHP-2. Samples immunoprecipitated from untreated or pervanadate-treated NK92 cells with anti-NKG2A or a control antibody were immunoblotted with phosphotyrosine- and SHP-2–specific antibodies. ERK immunoblot is shown as loading control.