CD63 Associates with Tyrosine Kinase Activity and CD11/ CD18, and Transmits an Activation Signal in Neutrophils

As a member of the tetraspan family, it has been hypothesized that CD63 may be associated with signal transduction; however, its role in leukocyte function is unknown. To examine the potential ability of CD63 to activate neutrophils, the effects of five CD63 mAbs, AHN-16, -16.1, -16.2, -16.3, and -16.5, were examined for their ability to alter neutrophil adhesion to HUVEC monolayers. These CD63 Abs increased neutrophil adhesion to resting and TNF-stimulated HUVEC monolayers. This increase in neutrophil adhesion caused by CD63 Abs was blocked by a CD18 Ab and was associated with up-regulation of CD11/CD18 and down-regulation of CD62L on the neutrophil surface. CD11/CD18 was also found to be associated with CD63. This increase in neutrophil adhesion required physiologic extracellular calcium concentrations at or near the time of CD63 Ab binding. The incubation of CD63 Abs with cells in the absence of calcium for 10 min before repletion of calcium resulted in no increase in neutrophil adhesion. Protein kinase activity was detected in neutrophils associated with CD63. Most of the protein kinase activity associated with these Ags was tyrosine kinase activity, with a lesser amount of threonine and serine kinase activities. Src family kinases Lyn and Hck accounted for much of the associated tyrosine kinase activity. The data suggest that CD63 Ab binding to the neutrophil surface triggers a transient activation signal that requires extracellular calcium and regulates the adhesive activity of CD11/CD18. Associated protein kinase activity may play a role in signal transduction by CD63 to regulate other cell functions.