Down-regulation of soluble guanylyl cyclase expression by cyclic AMP is mediated by mRNA-stabilizing protein HuR.
We analyzed whether the cyclic AMP induced down-regulation of the nitric oxide (NO) receptor soluble guanylyl cyclase (sGC) is mediated by the mRNA-protecting protein HuR. Exposure (up to 24 h) of isolated rat aortic segments to the activator of adenylyl cyclase, forskolin (10 microM), and to both activators of cAMP-stimulated protein kinase (PKA), 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole-3',5'-cyclic monophosphorothioate, Spisomer (Sp-5,6-DCl-cBIMPS; 400 nM), and N6-phenyl-cAMP (10 microM), strongly reduced sGCalpha1beta1 and HuR protein and mRNA expression in a time-dependent and actinomycin D (10 microM)-sensitive fashion. In vitro degradation of sGCalpha1 and beta1 poly(A)+ mRNA by native rat aortic protein was markedly increased by pretreatment of intact aortas with forskolin. Native protein extract from rat aorta shifted the electrophoretic mobility of biotin-labeled riboprobes from the 3'-untranslated region of sGCalpha1 and beta1 mRNA, and these bands was supershifted by a monoclonal antibody directed against the mRNA-stabilizing protein HuR. Forskolin decreased the HuR-sGCalpha1 and beta1 mRNA interaction and HuR protein expression in rat aorta, and this was prevented by the PKA inhibitory cAMP analog 3',5'-cyclic monophosphorothioate, Rp-isomer (Rp-cAMPS). In cultured smooth muscle cells from rat aorta, forskolin induced a rapid increase in Fos/p-Fos protein levels and activator protein 1 (AP-1) binding activity. Inhibition of this transcription factor by an AP-1 decoy prevented the forskolin-induced down-regulation of HuR. We conclude that forskolin/cAMP decrease the expression of heterodimeric sGC in rat aortic smooth muscle cells via activation of Fos/AP-1, which decreases the expression of HuR and thus destabilizes the sGCalpha1 and beta1 mRNA.
Klöss
Stephan
S
Institut für Kardiovaskuläre Physiologie, Johann Wolfgang v. Goethe-Universität, Frankfurt/Main, Germany. s.kloess@em.uni-frankfurt.de
Srivastava
Rashi
R
Mülsch
Alexander
A
D020413:3' Untranslated Regions / Q000187:drug effects / Q000378:metabolism; D000262:Adenylyl Cyclases; D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism*; D001011:Aorta; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology*; D003609:Dactinomycin / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D004347:Drug Interactions; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D015870:Gene Expression / Q000187:drug effects*; D005786:Gene Expression Regulation; D016762:Genes, fos; D006162:Guanylate Cyclase / Q000235:genetics / Q000378:metabolism*; D008297:Male; D020871:RNA Stability / Q000187:drug effects / Q000502:physiology*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D018808:Transcription Factor AP-1 / Q000378:metabolism
D020413:3' Untranslated Regions; D000954:Antigens, Surface; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D016601:RNA-Binding Proteins; D018808:Transcription Factor AP-1; D003609:Dactinomycin; D005576:Colforsin; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D006162:Guanylate Cyclase