2ND-MESSENGER REGULATION OF RECEPTOR ASSOCIATION WITH CLATHRIN-COATEDPITS - A NOVEL AND SELECTIVE MECHANISM IN THE CONTROL OF CD4 ENDOCYTOSIS

CD4, a member of the immunoglobulin superfamily, is not only expressed in T4 helper lymphocytes but also in myeloid cells. Receptor-mediated endocytosis plays a crucial role in the regulation of surface express ion of adhesion molecules such as CD4. In T lymphocytes p56(lck), a CD 4-associated tyrosine kinase, prevents CD4 internalization, but in mye loid cells p56(lck) is not expressed and CD4 is constitutively interna lized. In this study, we have investigated the role of cyclic AMP (cAM P) in the regulation of CD4 endocytosis in the myeloid cell line HL-60 . Elevations of cellular cAMP were elicited by 1) cholera toxin, 2) pe rtussis toxin, 3) forskolin and IBMX, 4) NaF, or 5) the physiological receptor agonist prostaglandin E-1. All five interventions led to an i nhibition of CD4 internalization. Increased cAMP levels did not inhibi t endocytosis per se, because internalization of insulin receptors and transferrin receptors and fluid phase endocytosis were either unchang ed or slightly enhanced. The mechanism of cAMP inhibition was further analyzed at the ultrastructural level. CD4 internalization, followed e ither by quantitative electron microscopy autoradiography or by immuno gold labeling, showed a rapid and temperature-dependent association of CD4 with clathrin-coated pits in control cells. This association was markedly inhibited in cells with elevated cAMP levels. Thus these find ings suggest a second-messenger regulation of CD4 internalization thro ugh an inhibition of CD4 association with clathrin-coated pits in p56( lck)-negative cells.