Ala/Thr(201) in extracellular loop 2 and Leu/Phe(290) in transmembrane domain 6 of type 1 frog gonadotropin-releasing hormone receptor confer differential ligand sensitivity and signal transduction.
Recently, we have identified three distinct types of bullfrog GnRH receptor (designated bfGnRHR-1, bfGnRHR-2, and bfGnRHR-3). In the present study, we have isolated three GnRHR clones in Rana dybowskii (dyGnRHR-1, dyGnRHR-2, and dyGnRHR-3). Despite high homology of dyGnRHRs with the corresponding bfGnRHRs, dyGnRHRs revealed different signaling pathways and ligand sensitivity compared with the bfGnRHR counterparts. Activation of dyGnRHRs with GnRH stimulated cAMP-mediated gene expression. However, dyGnRHR-3 but not dyGnRHR-1 and -2 induced c-fos promoter-driven gene expression. Consistently, dyGnRHR-1 and dyGnRHR-2 were not able to increase GnRH-induced inositol phosphate accumulation, whereas all bfGnRHRs and dyGnRHR-3 were, indicating that dyGnRHR-1 and dyGnRHR-2 are coupled to solely G(s), whereas all bfGnRHRs and dyGnRHR-3 are coupled to both G(s) and G(q/11). Moreover, dyGnRHR-1 and dyGnRHR-2 showed about 10-fold less sensitivity to each ligand than that of the bfGnRHR counterparts. Using type 1 chimeric and point-mutated receptors, we further elucidated that specific amino acids, Ala/Thr(201) in extracellular loop 2 and Leu/Phe(290) in transmembrane domain 6 of the type 1 receptor, are responsible for ligand sensitivity and signal transduction pathway. Particularly, substitution of Leu(290) to Phe in dyGnRHR-1 increased GnRH-induced inositol phosphate production as well as c-fos promoter-driven gene expression whereas substitution of Phe(290) to Leu in bfGnRHR-1 decreased those activities. Collectively, these results demonstrate the presence of three types of GnRHR in amphibians, and suggest species- and type-specific ligand recognition and different signaling pathways in frog GnRHRs.
Seong
Jae Young
JY
Hormone Research Center, Chonnam National University, Kwangju, 500-757, Korea.
Wang
Li
L
Oh
Da Young
DY
Yun
Oim
O
Maiti
Kaushik
K
Li
Jian Hua
JH
Soh
Jae Mok
JM
Choi
Hueng Sik
HS
Kim
Kyungjin
K
Vaudry
Hubert
H
Kwon
Hyuk Bang
HB
D000595:Amino Acid Sequence; D000818:Animals; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D016296:Mutagenesis / Q000502:physiology; D017354:Point Mutation / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011898:Ranidae; D011966:Receptors, LHRH / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D013045:Species Specificity
D008024:Ligands; D011966:Receptors, LHRH; D011993:Recombinant Fusion Proteins