Post-transcriptional regulation of soluble guanylyl cyclase expression in rat aorta.
We investigated the molecular mechanism of cyclic GMP-induced down-regulation of soluble guanylyl cyclase expression in rat aorta. 3-(5'-Hydroxymethyl-2'-furyl)-1-benzyl indazole (YC-1), an allosteric activator of this enzyme, decreased the expression of soluble guanylyl cyclase alpha(1) subunit mRNA and protein. This effect was blocked by the enzyme inhibitor 4H-8-bromo-1,2,4-oxadiazolo(3,4-d)benz(b-1,4)oxazin-1-one (NS2028) and by actinomycin D. Guanylyl cyclase alpha(1) mRNA-degrading activity was increased in protein extracts from YC-1-exposed aorta and was attenuated by pretreatment with actinomycin D and NS2028. Gelshift and supershift analyses using an adenylate-uridylate-rich ribonucleotide from the 3'-untranslated region of the alpha(1) mRNA and a monoclonal antibody directed against the mRNA-stabilizing protein HuR revealed HuR mRNA binding activity in aortic extracts, which was absent in extracts from YC-1-stimulated aortas. YC-1 decreased the expression of HuR, and this decrease was prevented by NS2028. Similarly, down-regulation of HuR by RNA interference in cultured rat aortic smooth muscle cells decreased alpha(1) mRNA and protein expression. We conclude that HuR protects the guanylyl cyclase alpha(1) mRNA by binding to the 3'-untranslated region. Activation of guanylyl cyclase decreases HuR expression, inducing a rapid degradation of guanylyl cyclase alpha(1) mRNA and lowering alpha(1) subunit expression as a negative feedback response.
Kloss
Stephan
S
Institut für Kardiovaskuläre Physiologie, Johann Wolfgang Goethe-Universität, Theodor Stern-Kai 7, Frankfurt/Main D60590, Germany.
Furneaux
Henry
H
Mülsch
Alexander
A
D020413:3' Untranslated Regions; D000818:Animals; D000954:Antigens, Surface; D001011:Aorta / Q000201:enzymology* / Q000378:metabolism / Q000473:pathology; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D003609:Dactinomycin / Q000494:pharmacology; D015536:Down-Regulation; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D020536:Enzyme Activators / Q000494:pharmacology; D006162:Guanylate Cyclase; D007191:Indazoles / Q000494:pharmacology; D008297:Male; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000166:cytology; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010069:Oxadiazoles / Q000494:pharmacology; D010078:Oxazines / Q000494:pharmacology; D011061:Poly A / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D018160:Receptors, Cytoplasmic and Nuclear / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071756:Soluble Guanylyl Cyclase; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection
D020413:3' Untranslated Regions; D000954:Antigens, Surface; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D020536:Enzyme Activators; D007191:Indazoles; C111257:NS 2028; D019384:Nucleic Acid Synthesis Inhibitors; D010069:Oxadiazoles; D010078:Oxazines; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D018160:Receptors, Cytoplasmic and Nuclear; C090937:3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole; D003609:Dactinomycin; D011061:Poly A; D012313:RNA; D006162:Guanylate Cyclase; D000071756:Soluble Guanylyl Cyclase