Retinoic acid induction of human tissue-type plasminogen activator gene expression via a direct repeat element (DR5) located at -7 kilobases.
All-trans-retinoic acid (RA) and retinoids induce synthesis of tissue-type plasminogen activator (t-PA) in endothelial and neuroblastoma cells in vitro and in rats in vivo. In HT1080 fibrosarcoma cells, induction of t-PA-related antigen secretion and t-PA mRNA steady state levels by RA were found to depend on de novo protein and mRNA synthesis. Fragments derived from the 5'-flanking region of the t-PA gene (+197 to -9578 base pairs (bp)) were linked to the chloramphenicol acetyltransferase gene. Transfection studies demonstrated that the region spanning bp -7145 to -9578 mediated induction by RA. A functional retinoic acid response element (RARE), consisting of a direct repeat of the GGGTCA motif spaced by 5 nucleotides (t-PA/DR5), was localized at -7.3 kilobases. The t-PA/DR5 element interacted with the heterodimer composed of retinoic acid receptor alpha and retinoid X receptor alpha in vitro, whereas its mutation abolished induction by RA in transient expression. In human EA.hy926 hybrid endothelial and in SK-N-SH neuroblastoma cells, the activity of t-PA/DR5 was found to be independent of the intervening sequence (-632 to -7144 bp) and of its distance from the transcription initiation site. Staurosporine, an inhibitor of protein kinase activity, inhibited induction by RA, suggesting that it required protein phosphorylation.
Bulens
F
F
Center for Molecular and Vascular Biology, University of Leuven, Belgium.
Ibañez-Tallon
I
I
Van Acker
P
P
De Vriese
A
A
Nelles
L
L
Belayew
A
A
Collen
D
D
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D004247:DNA / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004727:Endothelium / Q000201:enzymology; D004790:Enzyme Induction; D005354:Fibrosarcoma; D015870:Gene Expression / Q000187:drug effects*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009447:Neuroblastoma; D009838:Oligodeoxyribonucleotides; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D055503:Protein Multimerization; D051381:Rats; D018168:Receptors, Retinoic Acid / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis; D012091:Repetitive Sequences, Nucleic Acid / Q000187:drug effects*; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D017384:Sequence Deletion; D019311:Staurosporine; D010959:Tissue Plasminogen Activator / Q000096:biosynthesis* / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D000470:Alkaloids; D009838:Oligodeoxyribonucleotides; C000607071:RARA protein, human; C000607073:Rara protein, rat; D018168:Receptors, Retinoic Acid; D011994:Recombinant Proteins; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D014157:Transcription Factors; D014212:Tretinoin; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D011493:Protein Kinase C; D010959:Tissue Plasminogen Activator; D019311:Staurosporine