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Identification of three separate guanine nucleotide-binding proteins that interact with the delta-opioid receptor in NG108-15 neuroblastoma x glioma hybrid cells.
Mol Pharmacol. 1992 May; 41(5):822-31.MP

Abstract

Five separate guanine nucleotide-binding proteins (G proteins) were immunologically identified in membranes from neuroblastoma x glioma NG108-15 hybrid cells. These alpha subunit proteins were Gi2 alpha, two isoforms of Gi3 alpha, and two isoforms of Go alpha. The G proteins that interacted with delta-opioid receptors in these membranes were identified using cholera toxin (CTX)-induced ADP-ribosylation and antisera selective for various G protein alpha subunits. In the presence of delta-opioid agonists, CTX induced the incorporation of [32P]ADP-ribose into three pertussis toxin substrates. Using antisera generated against peptide sequences from G alpha subunits, these three pertussis toxin substrates were identified as Gi2 alpha, Go2 alpha, and one isoform of Gi3 alpha, which has yet to be identified. This CTX-induced labeling was demonstrated to be mediated via the delta-opioid receptor in these hybrid cells by the observation that delta agonists D-Ala2-D-Leu5-enkephalin (DA-DLE) and D-Pen2-D-Pen5-enkephalin, as well as the nonselective agonists etorphine and bremazocine, were active, but the mu agonist PL017 and the kappa agonist U-50-488H did not show this activity. This incorporation into all three substrates induced by DADLE was dose dependent, with EC50 (95% confidence interval) values ranging from 12 (3-52) to 183 (65-520) nM, which compared with the Kd value of 10 +/- 1.5 nM for this agonist, a dose that produces maximal inhibition of adenylate cyclase activity. Furthermore, pretreatment of the cells with pertussis toxin or treatment of the membranes with the antagonist naloxone blocked the incorporation induced by DADLE. Incorporation of [32P]ADP-ribose into all three substrates decreased 35-83% in membranes in which the receptors had been down-regulated by chronic treatment of the cells with DADLE. Thus, a single opioid receptor type can interact with three separate G proteins.

Authors+Show Affiliations

Department of Pharmacology, Louisiana State University School of Medicine, Shreveport 71130.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

1317000

Citation

Roerig, S C., et al. "Identification of Three Separate Guanine Nucleotide-binding Proteins That Interact With the Delta-opioid Receptor in NG108-15 Neuroblastoma X Glioma Hybrid Cells." Molecular Pharmacology, vol. 41, no. 5, 1992, pp. 822-31.
Roerig SC, Loh HH, Law PY. Identification of three separate guanine nucleotide-binding proteins that interact with the delta-opioid receptor in NG108-15 neuroblastoma x glioma hybrid cells. Mol Pharmacol. 1992;41(5):822-31.
Roerig, S. C., Loh, H. H., & Law, P. Y. (1992). Identification of three separate guanine nucleotide-binding proteins that interact with the delta-opioid receptor in NG108-15 neuroblastoma x glioma hybrid cells. Molecular Pharmacology, 41(5), 822-31.
Roerig SC, Loh HH, Law PY. Identification of Three Separate Guanine Nucleotide-binding Proteins That Interact With the Delta-opioid Receptor in NG108-15 Neuroblastoma X Glioma Hybrid Cells. Mol Pharmacol. 1992;41(5):822-31. PubMed PMID: 1317000.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of three separate guanine nucleotide-binding proteins that interact with the delta-opioid receptor in NG108-15 neuroblastoma x glioma hybrid cells. AU - Roerig,S C, AU - Loh,H H, AU - Law,P Y, PY - 1992/5/1/pubmed PY - 1992/5/1/medline PY - 1992/5/1/entrez SP - 822 EP - 31 JF - Molecular pharmacology JO - Mol Pharmacol VL - 41 IS - 5 N2 - Five separate guanine nucleotide-binding proteins (G proteins) were immunologically identified in membranes from neuroblastoma x glioma NG108-15 hybrid cells. These alpha subunit proteins were Gi2 alpha, two isoforms of Gi3 alpha, and two isoforms of Go alpha. The G proteins that interacted with delta-opioid receptors in these membranes were identified using cholera toxin (CTX)-induced ADP-ribosylation and antisera selective for various G protein alpha subunits. In the presence of delta-opioid agonists, CTX induced the incorporation of [32P]ADP-ribose into three pertussis toxin substrates. Using antisera generated against peptide sequences from G alpha subunits, these three pertussis toxin substrates were identified as Gi2 alpha, Go2 alpha, and one isoform of Gi3 alpha, which has yet to be identified. This CTX-induced labeling was demonstrated to be mediated via the delta-opioid receptor in these hybrid cells by the observation that delta agonists D-Ala2-D-Leu5-enkephalin (DA-DLE) and D-Pen2-D-Pen5-enkephalin, as well as the nonselective agonists etorphine and bremazocine, were active, but the mu agonist PL017 and the kappa agonist U-50-488H did not show this activity. This incorporation into all three substrates induced by DADLE was dose dependent, with EC50 (95% confidence interval) values ranging from 12 (3-52) to 183 (65-520) nM, which compared with the Kd value of 10 +/- 1.5 nM for this agonist, a dose that produces maximal inhibition of adenylate cyclase activity. Furthermore, pretreatment of the cells with pertussis toxin or treatment of the membranes with the antagonist naloxone blocked the incorporation induced by DADLE. Incorporation of [32P]ADP-ribose into all three substrates decreased 35-83% in membranes in which the receptors had been down-regulated by chronic treatment of the cells with DADLE. Thus, a single opioid receptor type can interact with three separate G proteins. SN - 0026-895X UR - https://www.unboundmedicine.com/medline/citation/1317000/Identification_of_three_separate_guanine_nucleotide_binding_proteins_that_interact_with_the_delta_opioid_receptor_in_NG108_15_neuroblastoma_x_glioma_hybrid_cells_ L2 - http://molpharm.aspetjournals.org/cgi/pmidlookup?view=long&pmid=1317000 DB - PRIME DP - Unbound Medicine ER -