RT Journal Article
SR Electronic
T1 Human 5-Hydroxytryptamine<sub>5A</sub> Receptors Activate Coexpressed G<sub>i</sub> and G<sub>o</sub> Proteins in<em>Spodoptera frugiperda</em> 9 Cells
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 1034
OP 1044
VO 57
IS 5
A1 Bart J. B. Francken
A1 Katty Josson
A1 Peter Lijnen
A1 Mirek Jurzak
A1 Walter H. M. L. Luyten
A1 Josée E. Leysen
YR 2000
UL http://molpharm.aspetjournals.org/content/57/5/1034.abstract
AB The ability of the human 5-hydroxytryptamine serotonin type 5A (h5-ht5A) receptor to couple to G proteins from distinct families was investigated through the simultaneous infection ofSpodoptera frugiperda 9 insect cells with recombinant baculoviruses encoding the various proteins. Expression of G proteins was demonstrated in immunoblots. Receptor-G protein coupling was monitored by high-affinity agonist binding and agonist-induced stimulation of [35S]guanosine-5′-O-(3-thio) triphosphate binding to membranes. Receptors expressed alone displayed low-affinity agonist binding, and endogenous G proteins were only poorly stimulated on the addition of 5-hydroxytryptamine. When receptors were coexpressed with mammalian Gi/Go proteins (Gαi or Gαo plus Gβ1γ2), the coupled phenotype was achieved: agonists bound with high affinity in a guanosine-5′-(β,γ-imido)triphosphate-sensitive manner and stimulated [35S]guanosine-5′-O-(3-thio)triphosphate binding to high levels. These effects were not observed on coexpression with Gz/Gs/Gq/11/16 or G12/13. Various ligands were evaluated for their agonistic, antagonistic, or inverse agonistic behavior in both receptor binding and activation assays. Although Go displayed different receptor coupling characteristics than Gi proteins, no clear coupling preference was evident. Coexpression of receptors and Gαi subunits without Gβ1γ2produced increases in both agonist affinity and maximum G protein activation that were smaller than those in the presence of Gβ1γ2, suggesting that Gβ1γ2 coexpression improves receptor-G protein coupling. Similarly, coexpression of receptors with Gβ1γ2 alone resulted in an improved interaction with endogenous G proteins. Our results demonstrate that h5-ht5A receptors expressed in Spodoptera frugiperda 9 cells selectively and functionally couple to coexpressed mammalian Gi and Go proteins. The American Society for Pharmacology and Experimental Therapeutics