Morphine Regulates Expression of m-Opioid Receptor MOR-1A, an Intron-Retention Carboxyl Terminal Splice Variant of the m-Opioid Receptor (OPRM1) Gene via miR-103/miR-107s

The m-opioid receptor (MOR-1) gene OPRM1 undergoes extensive alternative splicing, generating an array of splice variants. Of these variants, MOR-1A, an intron-retention carboxyl terminal splice variant identical to MOR-1 except for the terminal intracellular tail encoded by exon 3b, is quite abundant and conserved from rodent to humans. Increasing evidence indicates that miroRNAs (miRNAs) regulate MOR-1 expression and that m agonists such as morphine modulate miRNA expression. However, little is known about miRNA regulation of the OPRM1 splice variants. Using 39-rapid amplification cDNA end and Northern blot analyses, we identified the complete 39-untranslated region (39-UTR) for both mouse and human MOR-1A and their conserved polyadenylation site, and defined the role the 39-UTR in mRNA stability using a luciferase reporter assay. Computer models predicted a conserved miR-103/107 targeting site in the 39-UTR of both mouse and human MOR-1A. The functional relevance of miR-103/107 in regulating expression of MOR-1A protein through the consensus miR-103/107 binding sites in the 39-UTR was established by using mutagenesis and a miR-107 inhibitor in transfected human embryonic kidney 293 cells and Be (2)C cells that endogenously express human MOR-1A. Chronic morphine treatment significantly upregulated miR-103 and miR- 107 levels, leading to downregulation of polyribosome- associated MOR-1A in both Be(2)C cells and the striatum of a morphinetolerant mouse, providing a new perspective on understanding the roles of miRNAs and OPRM1 splice variants in modulating the complex actions of morphine in animals and humans.