Salmon growth hormone receptor: molecular cloning, ligand specificity, and response to fasting.
To better understand the role of growth hormone in regulating fish growth, the cDNA of growth hormone receptor (GHR) was cloned from the liver of masu salmon (Oncorhynchus masou) and characterized. The masu salmon GHR (msGHR) sequence revealed common features of a GHR, including a (Y/F)GEFS motif in the extracellular domain, a single transmembrane region, and Box 1 and Box 2 in the intracellular domain. However, the amino acid sequence identity was low (49%) compared to GHRs of other vertebrates including seven teleosts, and the putative msGHR protein lacked one pair of cysteine residues in the extracellular domain. To verify the identity of the msGHR, the recombinant protein of the extracellular domain was expressed with a histidine tag protein (His-msGHR-ECD), refolded and purified for analysis of its ligand specificity. In competition experiments, the specific binding between His-msGHR-ECD and radioiodine-labeled salmon GH was displaced completely by only salmon GH, and not by salmon prolactin or somatolactin. A real-time RT-PCR assay was used to measure salmon GHR mRNA in the liver of fed and fasted coho salmon (Oncorhynchus kisutch). The levels of hepatic GHR mRNA were lower in fasted fish compared to fed fish after 3 weeks, suggesting that GHR gene expression is reduced following a long-term fast. These results confirm the identity of the salmon GHR based on ligand specificity and response to fasting.
Fukada
Haruhisa
H
Division of Marine Bioscience, Graduate School of Fisheries Sciences, Hokkaido University, Hakodate, 041-8611, Japan.
Ozaki
Yuichi
Y
Pierce
Andrew L
AL
Adachi
Shinji
S
Yamauchi
Kohei
K
Hara
Akihiko
A
Swanson
Penny
P
Dickhoff
Walton W
WW
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D005215:Fasting / Q000378:metabolism*; D013006:Growth Hormone / Q000097:blood; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D008024:Ligands; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D017683:Oncorhynchus / Q000378:metabolism*; D017684:Oncorhynchus kisutch / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D011986:Receptors, Somatotropin / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014018:Tissue Distribution
D018076:DNA, Complementary; D008024:Ligands; D012333:RNA, Messenger; D011986:Receptors, Somatotropin; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone