AGONIST-INDUCED DOWN-REGULATION OF PLATELET-ACTIVATING-FACTOR RECEPTOR GENE-EXPRESSION IN U937 CELLS

Prolonged exposure (8-24 h) of human promonocytic U937 cells to 100 nM 1-O-hexadecyl-2-N-methylcarbamyl-sn-glycero-3- phosphocholine (carbar myl-PAF), a non-metabolizable analogue of platelet-activating factor ( PAF), reduced the numbers of PAF receptors by 50-75%, as determined by the radioligand-binding assay. To clarify whether the down-regulation of receptor numbers is due to decreased expression level of the PAF-r eceptor gene, the effect of carbamyl-PAF on the steady-state level of PAF-receptor mRNA was examined by a highly sensitive reverse-transcrip tase PCR method. A 50% decline in the level of PAF-receptor mRNA was o bserved in U937 cells pretreated with 100 nM carbamyl-PAF for 24 h. Th e effect of carbamyl-PAF was dose-dependent, with an EC(50) value arou nd 10 nM. A PAF-receptor antagonist, SRI-63675, was able to attenuate the effect of carbamyl-PAF. Furthermore lysoPAF, at 1 mu M, was unable to induce a significant decrease in PAF-receptor mRNA after incubatio n for 24 h, indicating that the effect of carbamyl-PAF was specific. T he half-life of the PAF-receptor mRNA measured in the presence of acti nomycin D was unaffected by carbamyl-PAF treatment. In contrast, nucle ar run-off experiments demonstrated that the transcription rate of the PAF-receptor gene in carbamyl-PAF-treated cells was about 65% of that in control cells. These results suggest that the PAF receptor in U937 cells is subject to down-regulation by agonist, at least partly, at t he transcriptional level.