Anti-AP-1 activity of all-trans retinoic acid in glomerular mesangial cells.
Functional antagonism between retinoic acid (RA) receptors and activator protein-1 (AP-1) transcription factors might regulate expression of genes involved in the response to injury in the kidney. We designed experiments to analyze the mechanisms by which RA inhibits AP-1-directed transcriptional responses in glomerular mesangial cells. RA inhibited serum-stimulated mesangial cell proliferation as assessed by measurements of [3H]thymidine uptake and cell number. In transient transfection assays with a chloramphenicol acetyltransferase reporter, RA completely blocked transcription directed by an AP-1 cis-element in cells stimulated by serum. AP-1 DNA binding was analyzed in electrophoretic gel mobility shift assays using nuclear extracts from control or RA-pretreated cells stimulated with serum. RA did not abolish AP-1 DNA binding activity under the conditions of this assay. The apparent equilibrium dissociation constant, maximal density of binding, and association rate for the AP-1-DNA interaction were similar in serum-stimulated cells or RA-pretreated cells stimulated with serum. RA repressed serum-stimulated induction of the immediate early genes c-fos and c-jun, whose protein products dimerize to form AP-1. Repression was relatively selective for c-fos/c-jun; induction of other immediate early transcription factors (junB, c-myc, and egr-1) was not downregulated by RA. That repression of c-fos by RA might contribute to anti-AP-1 activity was suggested by experiments with an antisense c-fos expression vector, which demonstrated that c-fos induction was required for serum-stimulated AP-1 activity. Together, these data demonstrate that RA antagonizes AP-1-directed transcription without inhibiting AP-1 DNA-binding in mesangial cells. Selective repression of c-fos and c-jun might contribute to the anti-AP-1 activity of RA.
Simonson
M S
MS
Department of Medicine, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106.
D000818:Animals; D015152:Blotting, Northern; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D003470:Culture Media; D004247:DNA / Q000096:biosynthesis*; D004305:Dose-Response Relationship, Drug; D016232:Endothelins / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects*; D005786:Gene Expression Regulation / Q000187:drug effects; D017781:Genes, Immediate-Early / Q000187:drug effects / Q000502:physiology*; D005920:Glomerular Mesangium / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D013936:Thymidine / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000037:antagonists & inhibitors*; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology*; D014316:Tritium
D003470:Culture Media; D016232:Endothelins; D018808:Transcription Factor AP-1; D014316:Tritium; D014212:Tretinoin; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D013936:Thymidine