Selective stimulation by ceramide of the expression of the alpha isoform of retinoic acid and retinoid X receptors in osteoblastic cells. A role of sphingosine 1-phosphate-mediated AP-1 in the ligand-dependent transcriptional activity of these receptors.
Recent studies have demonstrated that sphingosine 1-phosphate (SPP) plays a functional role as a signaling molecule in gene expression in several kinds of cells. The present study demonstrates selective expression by ceramide of retinoic acid receptor-alpha (RAR-alpha) and retinoic X receptor-alpha (RXR-alpha) in osteoblastic MC3T3-E1 cells and a functional role of SPP-mediated AP-1 in the signaling mechanism of ligand-dependent transcriptional activity of heterodimers of these receptors in the cells. C(2)- and C(6)-ceramides selectively stimulated the expression of RAR-alpha and RXR-alpha genes, but not that of beta and gamma isoform genes of RAR and RXR, in the cells. The C(2)-ceramide-induced stimulation was clearly inhibited by dl-threo-dihydrosphingosine, an inhibitor of sphingosine kinase. SPP also selectively stimulated the expression of both receptors and increased the specific binding of the nuclear proteins to direct repeat 5 (DR-5), a consensus sequence of RAR-RXR. In addition, SPP markedly stimulated transient chloramphenicol acetyltransferase (CAT) activity of retinoic acid-dependent transcriptional activity in the cells transfected with a DR-5-CAT reporter gene. The SPP stimulation was activation protein-1 (AP-1)-dependent, because the SPP stimulatory action toward these nuclear gene expressions and the transient CAT activity were inhibited by antisense c-fos and c-jun oligonucleotides. We observed that SPP actually stimulated AP-1 transcriptional activity in the cells. This study suggests an important role of SPP-mediated AP-1 in the selective expression of RAR-alpha and RXR-alpha in osteoblastic cells via the sphingosine pathway.
Department of Oral Microbiology, Meikai University School of Dentistry, Keyakidai, Sakado City, Saitama 350-0283, Japan.
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Takeshita
A
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Watanabe
Y
Y
Takada
S
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Hanazawa
D000818:Animals; D002518:Ceramides / Q000494:pharmacology*; D019281:Dimerization; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D016762:Genes, fos / Q000235:genetics; D016758:Genes, jun / Q000235:genetics; D008246:Lysophospholipids; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D015398:Signal Transduction / Q000187:drug effects; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013379:Substrate Specificity; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014212:Tretinoin / Q000494:pharmacology
D002518:Ceramides; D008246:Lysophospholipids; D009687:Nuclear Proteins; D020033:Protein Isoforms; C000607072:Rara protein, mouse; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D018808:Transcription Factor AP-1; D014157:Transcription Factors; C060506:sphingosine 1-phosphate; D014212:Tretinoin; D013108:Sphingomyelin Phosphodiesterase; D013110:Sphingosine