Cellular expression of the ␤ 2 -adrenergic receptor (␤ 2 -AR) is suppressed at the translational level by 3-untranslated region (UTR) sequences. To test the possible role of 3-UTR-binding proteins in translational suppression of ␤ 2 -AR mRNA, we expressed the full-length 3-UTR or the adenylate/uridylate-rich (A؉U-rich element (ARE)) RNA from the 3-UTR sequences of ␤ 2 -AR in cell lines that endogenously express this receptor. Reversal of ␤ 2 -adrenergic receptor translational repression by retroviral expression of 3-UTR sequences suggested that ARE RNA-binding proteins are involved in translational suppression of ␤ 2 -adrenergic receptor expression. Using a 20-nucleotide ARE RNA from the receptor 3-UTR as an affinity ligand, we purified the proteins that bind to these sequences. T-cell-restricted intracellular antigen-related protein (TIAR) was one of the strongly bound proteins identified by this method. UV-catalyzed cross-linking experiments using in vitro transcribed 3-UTR RNA and glutathione S-transferase-TIAR demonstrated multiple binding sites for this protein on ␤ 2 -AR 3-UTR sequences. The distal 340-nucleotide region of the 3-UTR was identified as a target RNA motif for TIAR binding by both RNA gel shift analysis and immunoprecipitation experiments. Overexpression of TIAR resulted in suppression of receptor protein synthesis and a significant shift in endogenously expressed ␤ 2 -AR mRNA toward low molecular weight fractions in sucrose gradient polysome fractionation. Taken together, our results provide the first evidence for translational control of ␤ 2 -AR mRNA by TIAR.1 are low abundance membrane-integrated G-protein-coupled receptors that are activated by catecholamines at the cell surface. ␤ 2 -AR mRNA is transcribed from a single intronless gene (1-3), and both transcriptional (4) and post-transcriptional mechanisms (5-10) are known to regulate receptor expression. Post-transcriptional mechanisms include regulation at the level of receptor mRNA translation (5-7) as well as regulation of mRNA stability (8, 9). A highly conserved 5Ј-leader cistron present in the 5Ј-untranslated region (UTR) of this receptor inhibits the translation of receptor mRNA (5, 6). ␤ 2 -Adrenergic receptors from several mammalian species have a highly conserved 3Ј-untranslated region (10) with multiple adenine/uridine-rich (ARE) regions (11-13). The 3Ј-untranslated region of the ␤ 2 -adrenergic receptor contains regulatory elements that can alter stability of the receptor transcript in response to agonist challenge (11-13). Recently, we demonstrated that the 3Ј-UTR sequences negatively regulate the translation of the receptor mRNA (7).The modulation of translational efficiency is an important post-transcriptional control mechanism for eukaryotic gene expression. In most cases, translational control results from interaction of RNA-binding proteins with the 5Ј-and/or 3Ј-UTR sequences (14 -28). Many sequence-specific RNA-binding proteins that bind to 3Ј-UTR of mRNAs have also been identified (29 -33). These interacting proteins ...

␤ 2 -Adrenergic receptors (␤ 2 -ARs) are low abundance integral membrane proteins that mediate the effects of catecholamines at the cell surface. Post-transcriptional regulation of ␤ 2 -AR is dependent, in part, on sequences within the 5-and 3-untranslated regions (UTRs) of the receptor mRNA. In this work, we demonstrate that 3-UTR sequences regulate the translation of the receptor mRNA. Deletion of the 3-UTR sequences resulted in 2-2.5-fold increases in receptor expression. The steadystate levels of ␤ 2 -AR mRNA did not change significantly in the presence or absence of the 3-UTR, suggesting that the translation of the receptor mRNA is suppressed by 3-UTR sequences. Introduction of the receptor 3-UTR sequences into the 3-UTR of a heterologous reporter gene (luciferase) resulted in a 70% decrease in reporter gene expression without significant changes in luciferase mRNA levels. Sucrose density gradient fractionation of cytoplasmic extracts from Chinese hamster ovary cells transfected with full-length receptor cDNA demonstrated that the receptor transcripts were distributed between polysomal and non-polysomal fractions. Deletion of 3-UTR sequences from the receptor cDNA resulted in a clear shift in the distribution of receptor mRNA toward the polysomal fractions, favoring increased translation. The 3-UTR sequences of the receptor mRNA were sufficient to shift the distribution of luciferase mRNA from predominantly polysomal fractions toward non-polysomal fractions in cells transfected with the chimeric luciferase construct. Taken together, our results provide the first evidence for translational control of ␤ 2 -AR expression by 3-UTR sequences. Presumably, this occurs by affecting the receptor mRNA localization.

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