CD50 (ICAM-3) IS PHOSPHORYLATED ON TYROSINE AND IS ASSOCIATED WITH TYROSINE KINASE-ACTIVITY IN HUMAN NEUTROPHILS

CD50 (ICAM-3) is expressed at a high level on resting blood granulocyt es, monocytes, and lymphocytes. The constitutive high expression of CD 50 on resting leukocytes suggests that it is an important LFA-1 ligand in the initiation of the immune/inflammatory response. Using a radiol abeling technique initially designed to detect ecto-protein kinase act ivity, we found that CD50 mAbs immunoprecipitated a similar to 125- to 170-kDa phosphoprotein from human neutrophils. Phosphorylation was in creased after stimulation with the chemotactic agent FMLP, platelet-ac tivating factor, 12-O-tetradecanoyl-phorbol-13-acetate, and the calciu m ionophore A23187. This increase in phosphorylation was transient wit h the maximal phosphorylation, being observed by 1 min. Phosphoamino a cid analysis revealed that CD50 contained predominantly phosphotyrosin e. Although this assay system was designed initially to detect ecto-pr otein kinase activity, subsequent studies have shown that membrane pro teins can be phosphorylated on the cytoplasmic domain under these cond itions. When CD50 was immunoprecipitated from solubilized neutrophils, protein tyrosine kinase activity associated with CD50 was detected in the immunoprecipitate. The data suggest that phosphorylation of CD50 on tyrosine by an associated tyrosine kinase plays a role in the funct ion of CD50.