Vol. 60, Issue 6, 1308-1324, December 2001

Agonist-Mediated Down-Regulation of Rat ₁-Adrenergic Receptor Transcripts: Role of Potential Post-Transcriptional Degradation Factors

Philbert Kirigiti, Ying Bai, Yong-Feng Yang, Xiaorong Li,¹ Biao Li,² Gary Brewer, and Curtis A. Machida

Division of Neuroscience, Oregon Regional Primate Research Center, Oregon Health & Science University West Campus, Beaverton, Oregon (P.K., Y.B., Y.-F.Y., X.L., B.L., C.A.M.); Department of Molecular Genetics and Microbiology, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway, New Jersey (G.B.); Department of Biochemistry and Molecular Biology and Graduate Program in Neuroscience, Oregon Health & Science University, Portland, Oregon (C.A.M.)

The human ₁-adrenergic receptor (AR) and hamster ₂-AR transcripts can be post-transcriptionally regulated at the level of mRNA stability and undergo accelerated agonist-mediated degradation via interaction of their 3' untranslated regions (UTR) with RNA binding proteins. Using RNase protection assays, we have determined that chronic isoproterenol exposure of rat C6 glioma cells results in the accelerated reduction of ₁-AR mRNAs. To determine the role of cellular environment on the agonist-independent and agonist-mediated degradation of ₁-AR mRNAs, we transfected rat ₁-AR expression recombinants into both hamster DDT₁MF2 cells and rat L6 cells. The rat ₁-AR mRNAs in the two transfectant cell pools retain longer agonist-independent half-lives than in the C6 environment and undergo accelerated degradation upon chronic agonist exposure. Using UV-cross-linking/immunoblot and immunoprecipitation analyses, we have determined that the rat ₁-AR 3' UTR recognizes a predominant M_r 39,000 component, identified as the mammalian elav-like protein HuR, and several other minor components, including the heteronuclear protein hnRNP A1. HuR levels are more highly expressed in C6 cells than in DDT₁MF2 and L6 cells and are induced after chronic isoproterenol treatment. Furthermore, C6 transfectants containing an HuR expression recombinant exhibit reduced ₁-AR mRNA half-lives that were statistically comparable with half-lives identified in isoproterenol-treated C6 cells. These results imply that HuR plays a potential role in the agonist-independent and agonist-mediated down-regulation of ₁-AR mRNAs.