Activation via multiple signaling pathways induces down-regulation of platelet-activating factor receptors on human B lymphocytes

Platelet-activating factor receptor (PAFR) has been identified in B cell li nes and primary human B cells, but the regulation of PAFR during B cell act ivation has not been completely elucidated. In the present study, we have i nvestigated the effects of B cell activation on PAFR binding parameters, PA FR mRNA and PAF-triggered intracellular calcium mobilization, The human B l ymphoid cell line LA350 was shown to exhibit high levels of PAFR (48,550 +/ - 4,310 sites/cell) as determined by radio-ligand binding assay with PAFR a ntagonist [H-3]WEB2086. Treatment with phorbol, 12,13-dibutyrate caused a b iphasic reduction of PAFR binding. The early phase was inhibited by the pro tein kinase C inhibitor bisindolylmaleimide I (BIM), whereas the late phase was not blocked by BIM, protein tyrosine kinase inhibitor genistein, or th e mitogen-activated protein kinase/extracellular signal-related kinase inhi bitor PD98059. However, staurosporine, a broad-spectrum protein kinase inhi bitor, completely inhibited the late phase down-regulation, Ionomycin also decreased [H-3]WEB2086 binding sites, whereas the combination of PDB and io nomycin induced a greater reduction than either agent alone. Cross-linking of B cell receptor by anti-IgM Ab also induced down-regulation of PAFR, whi ch was abolished by genistein or PD98059, but not by BIM or staurosporine. The decrease in surface PAFR number was closely paralleled by the reduction in PAFR mRNA both in LA350 cells and human tonsillar B cells, and was asso ciated with decreased response to PAF indicated by decreased intracellular calcium mobilization. These data show that multiple signaling pathways are involved in down-regulating PAFR expression during B cell activation and de velopment.