Cloning and characterization of the promoter region of the mouse mu opioid receptor gene.
Opioid compounds have potent analgesic and euphoric properties. They act with specific cell-membrane receptors which have been pharmacologically defined into three major classes, mu, kappa and delta. These receptors are highly regulated with respect to their gene expression, resulting in a temporally and spatially specific pattern of distribution for each receptor. To characterize the promoter sequence of the mu opioid receptor (MOR) gene, a mouse genomic DNA library was screened under high stringency with a rat MOR (MOR-1) cDNA probe and genomic sequences for the mouse MOR gene were isolated. From one genomic clone, a 2.3-kb EcoRI fragment, which hybridized to the 5'-end of the rat MOR-1 cDNA 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 initiation site 793 bp upstream from the translation start site. Chimeric constructs of mouse MOR deletion fragments fused to a luciferase reporter gene were transfected into a human neuroblastoma cell line, SK-N-SH, which constitutively expresses endogenous MOR. These transient expression studies indicated that the 0.2-kb region upstream from the transcription initiation site possesses a functional promoter, which directs the expression of the reporter gene in vitro and may possess promoter activity for the mouse MOR gene in vivo.
Liang
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Y
Department of Pharmacology and Toxicology, Indianapolis, IN 46202-5121, USA.
Mestek
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A
Yu
L
L
Carr
L G
LG
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D005786:Gene Expression Regulation / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D010442:Peptide Chain Initiation, Translational; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D017450:Receptors, Opioid, mu / Q000235:genetics*; D014162:Transfection
D017450:Receptors, Opioid, mu