Functional studies of a glucagon receptor isolated from frog Rana tigrina rugulosa: implications on the molecular evolution of glucagon receptors in vertebrates

In this report, the first amphibian glucagon receptor (GluR) cDNA was chara cterized from the liver of the frog Rana tigrina rugulosa. Functional expre ssion of the frog GluR in CHO and COS-7 cells showed a high specificity of the receptor towards human glucagon with an EC50 value of 0.8 +/- 0.5 nM, T he binding of radioiodinated human glucagon to GluR was displaced in a dose -dependent manner only with human glucagon and its antagonist (des-His(1)-[ Nle(9)-Ala(11)-Ala(16)]) with IC50 values of 12.0 +/- 3.0 and 7.8 +/- 1.0 n M, respectively. The frog GluR did not display any affinity towards fish an d human GLP-1s, and towards glucagon peptides derived from two species of t eleost fishes (goldfish, zebrafish), These fish glucagons contain substitut ions in several key residues that mere previously shown to be critical for the binding of human glucagon to its receptor. By RT-PCR, mRNA transcripts of frog GluR were located in the liver, brain, small intestine and colon. T hese results demonstrate a conservation of the functional characteristics o f the GluRs in frog and mammalian species and provide a framework for a bet ter understanding of the molecular evolution of the GluR and its physiologi cal function in vertebrates. (C) 1999 Federation of European Biochemical So cieties.