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 (carbarmyl-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-receptor gene, the effect of carbamyl-PAF on the steady-state level of PAF-receptor mRNA was examined by a highly sensitive reverse-transcriptase PCR method. A 50% decline in the level of PAF-receptor mRNA was observed in U937 cells pretreated with 100 nM carbamyl-PAF for 24 h. The effect of carbamyl-PAF was dose-dependent, with an EC50 value around 10 nM. PAF-receptor antagonist, SRI-63675, was able to attenuate the effect of carbamyl-PAF. Furthermore lysoPAF, at 1 uM, was unable to induce a significant decrease in PAF-receptor mRNA after incubation for 24 h, indicating that the effect of carbamyl-PAF was specific. The half-life of the PAF-receptor mRNA measured in the presence of actinomycin D was unaffected by carbamyl-PAF treatment. In contrast, nuclear 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 the transcriptional level.
Chau
L Y
LY
Division of Cardiovascular Research, Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan, Republic of China.
Peck
K
K
Yen
H H
HH
Wang
J Y
JY
D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D015536:Down-Regulation; D005786:Gene Expression Regulation / Q000187:drug effects*; D006207:Half-Life; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010972:Platelet Activating Factor / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D014158:Transcription, Genetic
D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C039351:platelet activating factor receptor; C083283:1-O-hexadecyl-2-N-methylcarbamol -sn-glycerol-3-phosphocholine; D012194:RNA-Directed DNA Polymerase