title: Regulation of AT1R expression through HuR by insulin.
abstract: Angiotensin II type 1 receptor (AT1R) has a pathophysiological role in hypertension, atherosclerosis and heart failure. Type 2 diabetes is hyperinsulinemic state and a major risk factor for atherosclerosis and hypertension. It is known that hyperinsulinemia upregulates AT1R expression post-transcriptionally by increasing the half-life of AT1R mRNA, but little is known about the mechanism of this effect. In the present study, we first identified AT1R 3'-UTR as a mediator of insulin effect. Using 3'-UTR as a bait, we identified through analysis of insulin-stimulated cell lysates by affinity purification and mass spectrometry HuR as an insulin-regulated AT1R mRNA binding protein. By ribonucleoprotein immunoprecipitation, we found HuR binding to AT1R to be increased by insulin. Overexpression of HuR leads to increased AT1R expression in a 3'-UTR-dependent manner. Both insulin and HuR overexpression stabilize AT1R 3'-UTR and their responsive element within 3'-UTR are located within the same region. Cell fractionation demonstrated that insulin induced HuR translocation from nucleus to cytoplasm increased HuR binding to cytoplasmic AT1R 3'-UTR. Consistent with HuR translocation playing a mechanistic role in HuR effect, a reduction in the cytoplasmic levels of HuR either by silencing of HuR expression or by inhibition of HuR translocation into cytoplasm attenuated insulin response. These results show that HuR translocation to cytoplasm is enhanced by insulin leading to AT1R upregulation through HuR-mediated stabilization of AT1R mRNA.
authors.affiliation: Research Program for Molecular Medicine, Biomedicum Helsinki, FIN-00014 University of Helsinki, Finland.
authors.forename: Kirsi
authors.initials: K
authors.lastname: Paukku
authors.forename: Michael
authors.initials: M
authors.lastname: Backlund
authors.forename: Rudolf A
authors.initials: RA
authors.lastname: De Boer
authors.forename: Nisse
authors.initials: N
authors.lastname: Kalkkinen
authors.forename: Kimmo K
authors.initials: KK
authors.lastname: Kontula
authors.forename: Jukka Y A
authors.initials: JY
authors.lastname: Lehtonen
mesh_terms: D020413:3' Untranslated Regions; D001665:Binding Sites; D002478:Cells, Cultured; D051959:ELAV Proteins / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D021381:Protein Transport / Q000187:drug effects; D020871:RNA Stability; D012333:RNA, Messenger / Q000378:metabolism*; D044140:Receptor, Angiotensin, Type 1 / Q000235:genetics* / Q000378:metabolism
chemical_list: D020413:3' Untranslated Regions; D051959:ELAV Proteins; D005231:Fatty Acids, Unsaturated; D007328:Insulin; D012333:RNA, Messenger; D044140:Receptor, Angiotensin, Type 1; C038753:leptomycin B