CD31 (PECAM-1), CDw32 (FcγRII), and anti-HLA class I monoclonal antibodies recognize phosphotyrosine-containing proteins on the surface of human neutrophils

Keith M Skubitz, Said A. Goueli

Research output: Contribution to journalArticlepeer-review

22 Scopus citations

Abstract

Although most studies of protein phosphorylation have focused on intracellular reactions, studies have provided evidence for the existence of ectoprotein kinase activity on the surface of some cells including human neutrophils. The identification and characterization of physiologic substrates of ectoprotein kinase activity should aid the understanding of the role of this enzyme activity in cell function. Immunoprecipitation and subsequent gel electrophoresis of proteins from neutrophils labeled with [γ- 32P]ATP under conditions initially designed to detect ectoprotein kinase activity revealed that CD31, CDw32, and anti-HLA class I mAbs specifically recognize phosphoproteins on the surface of human neutrophils. Phosphorylation of these proteins was inhibited by pretreatment of cells with an impermeant sulfhydryl reagent before radiolabeling. Phosphoamino acid analysis of the proteins revealed that they contained predominantly phosphotyrosine. However, controlled proteolysis of intact cells and purified HLA class I revealed that the HLA class I heavy chain was phosphorylated on the cytoplasmic domain. These results suggest that the molecules recognized by CD31 (PECAM-1) and CDw32 (FcγRII) Abs may also be phosphorylated on cytoplasmic domains under conditions originally designed to detect ectoprotein kinase activity. Phosphorylation of CD31 (PECAM-1), FcγRII, and HLA class I heavy chain on tyrosine may play a role in regulating their function. These results emphasize that the demonstration that a membrane protein is an ectokinase substrate is complex and requires the definitive localization of the phosphorylated residue to the extracellular domain of the protein.

Original languageEnglish (US)
Pages (from-to)5902-5911
Number of pages10
JournalJournal of Immunology
Volume152
Issue number12
StatePublished - Jun 15 1994

Fingerprint Dive into the research topics of 'CD31 (PECAM-1), CDw32 (FcγRII), and anti-HLA class I monoclonal antibodies recognize phosphotyrosine-containing proteins on the surface of human neutrophils'. Together they form a unique fingerprint.

Cite this