CLONING AND CHARACTERIZATION OF THE PROMOTER REGION OF THE MOUSE MU-OPIOID RECEPTOR GENE

Opioid compounds have potent analgesic and euphoric properties. They a ct with specific cell-membrane receptors which have been pharmacologic ally defined into three major classes, mu, kappa and delta. These rece ptors are highly regulated with respect to their gene expression, resu lting in a temporally and spatially specific pattern of distribution f or each receptor. To characterize the promoter sequence of the mu opio id receptor (MOR) gene, a mouse genomic DNA library was screened under high stringency with a rat MOR (MOR-1) cDNA probe and genomic sequenc es for the mouse MOR gene were isolated. From one genomic clone, a 2.3 -kb EcoRI fragment, which hybridized to the S-end of the rat MOR-1 cDN A probe, was subcloned and sequenced. This fragment contains 1.3 kb of sequence upstream of the initiation codon, extends downstream through exon 1 and includes a portion of intron 1. Primer extension analysis using mouse brain poly (A)(+) RNA identified a transcription initiatio n site 793 bp upstream from the translation start site. Chimeric const ructs of mouse MOR deletion fragments fused to a luciferase reporter g ene were transfected into a human neuroblastoma cell line, SK-N-SH, wh ich constitutively expresses endogenous MOR. These transient expressio n studies indicated that the 0.2-kb region upstream from the transcrip tion initiation site possesses a functional promoter, which directs th e expression of the reporter gene in vitro and may possess promoter ac tivity for the mouse MOR gene in vivo.