Characterization of CB1 cannabinoid receptors using receptor peptide fragments and site-directed antibodies.
The mechanism by which CB1 cannabinoid receptors are coupled to the Gi/Go class of G proteins was studied. A peptide representing the juxtamembrane carboxyl terminus robustly stimulated guanosine-5'-O-(3-thio)triphosphate binding. Peptides simulating subdomains of the third intracellular loop (IL3) activated minimally when present alone but produced additive effects when present in combination. Peptides representing the amino-side IL3 and the juxtamembrane carboxyl terminus autonomously inhibited adenylate cyclase, and this response was not significantly augmented or inhibited by peptides representing other intracellular domains. Site-directed antipeptide antibodies developed against the domains of the amino terminus, first extracellular loop, amino-side IL3, and juxtamembrane carboxyl terminus of CB1 receptors failed to influence binding of [3H]CP-55940. However, IgG raised against the amino-side IL3 diminished the agonist-dependent inhibition of adenylate cyclase. These experiments suggest that the juxtamembrane carboxyl terminus is critical for G protein activation by CB1 cannabinoid receptors and that the amino-side IL3 also may interact with Gi proteins leading to inhibition of adenylate cyclase.
Howlett
A C
AC
Department of Pharmacological and Physiological Science, Saint Louis University School of Medicine, St. Louis, Missouri 63104, USA. howletta@slu.edu
Song
C
C
Berglund
B A
BA
Wilken
G H
GH
Pigg
J J
JJ
D000067956:Adenylyl Cyclase Inhibitors; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019204:GTP-Binding Proteins / Q000502:physiology; D007106:Immune Sera / Q000276:immunology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology
D000067956:Adenylyl Cyclase Inhibitors; D007106:Immune Sera; D010446:Peptide Fragments; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug; D019204:GTP-Binding Proteins