Many cell surface receptors that mediate the actions of hormones and neurotransmitters are coupled to the activation or inhibition of intracellular enzymes or of ion channels by GTP-binding regulatory proteins (G proteins). The activated G proteins thereby transmit information about receptor occupancy to the Interior of cells. The long-term goal of the research of this section has been to understand in detail the mechanism of action of the research of this section has been to understand in detail the mechanism of action of receptors for the opiates and their close relatives. The neuroblastoma x glioma hybrid cell line, NG 108- 15, is richly endowed with opiate receptors, and is a particularly good source of this protein since it expresses only the delta type of opiate receptor. In the past year we have Isolated many peptides believed to be fragments of opiate receptors and determined their amino acid sequences. In the past year we, and others, have successfully obtained a clone of the delta opiate receptor of NG 108-15 cells. The clone which we isolated is apparently identical to that reported by Evans et al. (Science 258: 1952-1955 (1992), and essentially identical with that of Kieffer et al. (Proc. Natl. Acad. Sci. USA 89: 12048-12052 (1992). We have found full length transcripts of the gene in mouse brain but in no other tissues examined. With In the brain the gene is expressed at low levels in many regions but transcripts are found in particularly large amounts in the anterior pituitary and pineal glands. Since these tissues are located outside the blood brain barrier, opioid peptides easily can reach receptors in these areas from the blood. The gene, which is present as a single copy, has been mapped to the distal region of mouse chromosome 4. which corresponds to human chromosome ID. Even though there is only a single gene specifying the delta opiate receptor, there are 5 size species of mRNA corresponding to this gene in NG108-15 cells and all of them are regulated by opiate agonists. There is clearly much to be learned about the regulation of receptor gene transcription and translation. A third new approach which is made possible by the availability of clones is an analysis of the structural basis of agonist efficacy by site- directed mutagenesis. Such studies were started two years ago with a clone of alpha2-adrenergic receptor with the aim of examining the roles of conformational flexibility in determining the affinity and efficacy of various hormones and drugs, and the first results to emerge are of real interest, and will be discussed below. Related studies have also been undertaken with the newly cloned opiate receptor.
