Heterodimerization of gamma-aminobutyric acid B receptor subunits as revealed by the yeast two-hybrid system.
Several lines of evidence suggested that the first gamma-aminobutyric acid B receptor to be cloned required an additional factor for functional expression. GABA(B1) was retained within the endoplasmic reticulum and failed to couple to signal transduction pathways on stimulation with agonists. In radioligand binding experiments it was found that although the affinity of antagonists showed a close agreement between rat brain membranes and membranes expressing the cloned receptor, agonist ligands were significantly weaker at recombinant receptors. Using the C-terminal tail as bait, a yeast two-hybrid screen was run against a human brain cDNA library and identified a second receptor, GABA(B2), as a major interacting protein. This interaction was confirmed by coimmunoprecipitation as well as extensive colocalization studies. Coexpression of the two seven-transmembrane proteins generated a fully functional receptor, which was expressed at the cell surface confirming the importance of receptor heterodimerization for GABA(B) receptor activity.
White
Julia H
JH
Pathway Discovery, GlaxoSmithKline, Gunnels Wood Road, Stevenage, Hertfordshire SG1 2NY, UK. julia.h.white@gsk.com
Wise
Alan
A
Marshall
Fiona H
FH
D002478:Cells, Cultured; D019281:Dimerization; D005434:Flow Cytometry; D005614:Freeze Fracturing; D015723:Gene Library; D006031:Glycosylation; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D007668:Kidney / Q000166:cytology; D011233:Precipitin Tests; D018080:Receptors, GABA-B / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D013462:Sulfur Radioisotopes; D014162:Transfection; D020798:Two-Hybrid System Techniques; D015003:Yeasts; D001616:beta-Galactosidase
D018080:Receptors, GABA-B; D013462:Sulfur Radioisotopes; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D001616:beta-Galactosidase