cDNA cloning and functional expression of growth hormone receptor from soft-shelled turtle (Pelodiscus sinensis japonicus).
The growth hormone receptor (GHR) cDNA was cloned from the liver of soft-shelled turtle (Pelodiscus sinensis japonicus) using the polymerase chain reaction (PCR). Although GHR has been cloned from several mammalian and avian species, this is the first description of the reptilian receptor. As deduced from the nucleotide sequence, the precursor GHR of soft-shelled turtle (tGHR) is a protein of 615 amino acids which presents 72% identity with the chicken receptor and 57-64% identity with GHRs of several mammals. The tGHR expressed in COS-7 cells specifically bound human growth hormone (hGH) and was able to transduce an activation of transcription in the transfected cells. Binding of (125)I-hGH to the expressed receptor was decreased by the addition of excess unlabeled hGH, pig GH, and bream GH but not by pig insulin. The open reading frame of tGHR cDNA was inserted into the pSINrep/gfp (green fluorescence protein) vector and the tGHR-gfp fusion protein was stably expressed in baby hamster kidney (BHK) cells. Confocal imaging showed that tGHR-gfp was largely concentrated on the plasma membrane. Western blot analysis and deglycosylation treatment with PNGase F demonstrated that tGHR was a glycoprotein in BHK cells.
Zhang
X
X
Shanghai Institute of Biochemistry, Academia Sinica, Shanghai 200031, China.
Lu
X
X
Jing
N
N
Zhu
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D019556:COS Cells; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000235:genetics*; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D006801:Humans; D007668:Kidney; D008164:Luminescent Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D011986:Receptors, Somatotropin / Q000737:chemistry / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000032:analysis; D016415:Sequence Alignment; D014162:Transfection; D014426:Turtles / Q000235:genetics*
D018076:DNA, Complementary; D008164:Luminescent Proteins; D011986:Receptors, Somatotropin; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins