Properties of a kappa-opioid receptor expressed in CHO cells: interaction with multiple G-proteins is not specific for any individual G alpha subunit and is similar to that of other opioid receptors.
The purpose of the present study was to examine the coupling pattern of a recently cloned kappa-opioid receptor stably transfected in CHO cells to individual G alpha subunits with subsequent comparison to that observed previously for delta- and mu-opioid receptors. Data presented in the current study indicate the successful stable expression of a kappa-opioid receptor in CHO cells. This is supported by experiments in which ligands with selectivity for kappa-, but not delta- or mu-opioid receptors demonstrated high affinity for the expressed receptor and were able to potently and efficaciously produce inhibition of adenylyl cyclase activity. In addition, only kappa-opioid agonists were able to induce dose-dependent increases in the incorporation of [32P]azidoanilido-GTP into four G alpha subunits, three of which were identified as Gi3 alpha, Gi2 alpha and Go2 alpha. Further, the amount of kappa-opioid agonists required to induce 50% maximal labeling of any individual G alpha subunit was similar. Although kappa-opioid agonists produced equivalent maximal labelling of Gi3 alpha, Gi2 alpha and Go2 alpha, significantly less agonist-induced labeling was observed for an unknown G-protein designated as G? alpha. Although these results are slightly different than those observed previously for both delta- and mu-opioid receptors, it appears that all opioid receptors stably transfected in CHO cells interact with multiple G-proteins and that this coupling is not selective for any individual G alpha subunit.
Prather
P L
PL
Department of Pharmacology, School of Medicine, University of Minnesota, Minneapolis, USA.
McGinn
T M
TM
Claude
P A
PA
Liu-Chen
L Y
LY
Loh
H H
HH
Law
P Y
PY
D000262:Adenylyl Cyclases / Q000032:analysis; D000345:Affinity Labels; D000818:Animals; D001345:Autoradiography; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D019204:GTP-Binding Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D017465:Receptors, Opioid, delta / Q000096:biosynthesis / Q000235:genetics*; D017473:Receptors, Opioid, kappa / Q000096:biosynthesis / Q000235:genetics*; D017450:Receptors, Opioid, mu / Q000096:biosynthesis / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection
D000345:Affinity Labels; D010446:Peptide Fragments; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D011994:Recombinant Proteins; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases