Molecular identification and functional expression of mu 3, a novel alternatively spliced variant of the human mu opiate receptor gene.
Studies from our laboratory have revealed a novel mu opiate receptor, mu 3, which is expressed in both vascular tissues and leukocytes. The mu 3 receptor is selective for opiate alkaloids and is insensitive to opioid peptides. We now identify the mu 3 receptor at the molecular level using a 441-bp conserved region of the mu 1 receptor. Sequence analysis of the isolated cDNA suggests that it is a novel, alternatively spliced variant of the mu opiate receptor gene. To determine whether protein expressed from this cDNA exhibits the biochemical characteristics expected of the mu 3 receptor, the cDNA clone was expressed in a heterologous system. At the functional level, COS-1 cells transfected with the mu 3 receptor cDNA exhibited dose-dependent release of NO following treatment with morphine, but not opioid peptides (i.e., Met-enkephalin). Naloxone was able to block the effect of morphine on COS-1 transfected cells. Nontransfected COS-1 cells did not produce NO in the presence of morphine or the opioid peptides at similar concentrations. Receptor binding analysis with [(3)H]dihydromorphine further supports the opiate alkaloid selectivity and opioid peptide insensitivity of this receptor. These data suggest that this new mu opiate receptor cDNA encodes the mu 3 opiate receptor, since it exhibits biochemical characteristics known to be unique to this receptor (opiate alkaloid selective and opioid peptide insensitive). Furthermore, using Northern blot, RT-PCR, and sequence analysis, we have demonstrated the expression of this new mu variant in human vascular tissue, mononuclear cells, polymorphonuclear cells, and human neuroblastoma cells.
Cadet
Patrick
P
Neuroscience Research Institute, State University of New York, Old Westbury, NY 11568, USA.
Mantione
Kirk J
KJ
Stefano
George B
GB
D017398:Alternative Splicing / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive / Q000235:genetics; D015152:Blotting, Northern; D019556:COS Cells; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D015870:Gene Expression / Q000502:physiology; D015723:Gene Library; D006801:Humans; D008969:Molecular Sequence Data; D009292:Narcotic Antagonists / Q000378:metabolism / Q000494:pharmacology; D009928:Organ Specificity / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D014162:Transfection
D009292:Narcotic Antagonists; C485764:OPRM1 protein, human; D020033:Protein Isoforms; D017450:Receptors, Opioid, mu