title: MicroRNA let-7 establishes expression of beta2-adrenergic receptors and dynamically down-regulates agonist-promoted down-regulation.
abstract: Although β(2)-adrenergic receptors (β(2)AR) are expressed on most cell types, mechanisms that establish expression levels and regulate expression by chronic agonist remain unclear. The 3' UTR of ADRB2 has a conserved 8-nucleotide seed region that we hypothesized is targeted by the let-7 family of miRNAs leading to translational repression. In luciferase assays with transfected cells, luc-β(2)WT3'UTR had decreased expression when cotransfected with let-7f, but a mutated luc-β(2)3'UTR lacking the seed was unaffected by let-7f; a mutated let-7f also had no effect on luc-β(2)WT3'UTR expression. ADRB2 mRNA was in greater abundance in immunoprecipitates of Ago2, a core component of the miRNA-induced silencing complex, when cells were transfected with let-7f, but not with a mutated let-7f, indicating a direct interaction with the silencing mechanism. H292 cells transfected with let-7f caused ∼60% decrease in native β(2)AR expression, but transfection with let-7f-specific locked nucleic acid anti-miRNA increased β(2)AR expression by ∼twofold. We considered that an increase in let-7f leading to greater repression of translation contributes to agonist-promoted down-regulation. Paradoxically, in cells and in lungs from mice treated in vivo, an ∼50% decrease in let-7f occurs during long-term agonist exposure, indicating a counterregulatory event. Consistent with this notion, let-7f locked nucleic acid transfection caused depressed agonist-promoted down-regulation. Thus, let-7f miRNA regulates baseline β(2)AR expression and decreases in let-7f evoked by agonist attenuate down-regulation. This positive feedback loop has not previously been described for a G protein-coupled receptor and its miRNA. Methods to decrease let-7f expression in targeted cells may increase therapeutic responses to β-agonist by increasing β(2)AR expression or minimizing tachyphylaxis.
authors: Department of Medicine, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors: Wayne C H
authors: WC
authors: Wang
authors: Aster H
authors: AH
authors: Juan
authors: Alfredo
authors: A
authors: Panebra
authors: Stephen B
authors: SB
authors: Liggett
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D015536:Down-Regulation; D006801:Humans; D051379:Mice; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D034622:RNA Interference; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics*
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D035683:MicroRNAs; D018343:Receptors, Adrenergic, beta-2; C487931:mirnlet7 microRNA, human