Nitric oxide increases the decay of matrix metalloproteinase 9 mRNA by inhibiting the expression of mRNA-stabilizing factor HuR.
Dysregulation of extracellular matrix turnover is an important feature of many inflammatory processes. Rat renal mesangial cells express high levels of matrix metalloproteinase 9 (MMP-9) in response to inflammatory cytokines such as interleukin-1 beta. We demonstrate that NO does strongly destabilize MMP-9 mRNA, since different luciferase reporter gene constructs containing the MMP-9 3' untranslated region (UTR) displayed significant reduced luciferase activity in response to the presence of NO. Moreover, by use of an in vitro degradation assay we found that the cytoplasmic fractions of NO-treated cells contained a higher capacity to degrade MMP-9 transcripts than those obtained from control cells. An RNA electrophoretic mobility shift assay demonstrated that three of four putative AU-rich elements present in the 3' UTR of MMP-9 were constitutively occupied by the mRNA-stabilizing factor HuR and that the RNA binding was strongly attenuated by the presence of NO. The addition of recombinant glutathione transferase-HuR prevented the rapid decay of MMP-9 mRNA, whereas the addition of a neutralizing anti-HuR antibody caused an acceleration of MMP-9 mRNA degradation. Furthermore, the expression of HuR mRNA and protein was significantly reduced by exogenously and endogenously produced NO. These inhibitory effects were mimicked by the cGMP analog 8-bromo-cGMP and reversed by LY-83583, an inhibitor of soluble guanylyl cyclase. These results demonstrate that NO acts in a cGMP-dependent mechanism to inhibit the expression level of HuR, thereby reducing the stability of MMP-9 mRNA.
Akool
El-Sayed
el-S
Pharmazentrum Frankfurt, Klinikum der Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany.
Kleinert
Hartmut
H
Hamada
Farid M A
FM
Abdelwahab
Mohamed H
MH
Förstermann
Ulrich
U
Pfeilschifter
Josef
J
Eberhardt
Wolfgang
W
D020413:3' Untranslated Regions; D000634:Aminoquinolines / Q000494:pharmacology; D000818:Animals; D000954:Antigens, Surface; D001483:Base Sequence; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000494:pharmacology; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors / Q000378:metabolism; D007375:Interleukin-1 / Q000494:pharmacology; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000235:genetics* / Q000378:metabolism; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D009569:Nitric Oxide / Q000378:metabolism* / Q000494:pharmacology; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009603:Nitroso Compounds / Q000494:pharmacology; D020871:RNA Stability / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid; D012261:Ribonucleoproteins / Q000378:metabolism
D020413:3' Untranslated Regions; D000634:Aminoquinolines; D000954:Antigens, Surface; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D004791:Enzyme Inhibitors; D007375:Interleukin-1; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; D003609:Dactinomycin; C016276:8-bromocyclic GMP; D009569:Nitric Oxide; C041715:6-anilino-5,8-quinolinedione; D020780:Matrix Metalloproteinase 9; D006162:Guanylate Cyclase; D006152:Cyclic GMP