Identification of promoter region of ghrelin gene in human medullary thyroid carcinoma cell line.
Ghrelin, an endogenous ligand for growth hormone (GH) secretagogue receptor, stimulates GH secretion. The ghrelin gene is expressed most abundantly in stomach. The mRNA is also detected in other tissues and cell lines. However, the mechanism of the transcriptional regulation of the ghrelin gene has not yet been clarified. In the present study, we have investigated the regulatory region of the ghrelin gene expression in the human medullary thyroid carcinoma cell line (TT cells). PCR analysis of the 5'-region for human ghrelin gene revealed the presence of the first exon corresponding to the short non-coding first exon of the mouse ghrelin gene. The first exon is located at the 502 bp upstream from the 5'-end of the formerly reported human ghrelin gene. RT-PCR analysis showed the expression of the first exon in the stomach and TT cells. The expression of the first exon in the human stomach was confirmed by 5'-RACE method. Significant level of promoter activity was observed in the 1225-1107 bp up-stream region of the translation initiation site by luciferase assay. Specific protein binding to the promoter region of -1129 to -1100 was detected by electrophoretic mobility shift assay with nuclear extract from TT cells. These results suggest that the ghrelin gene expression in TT cells might be regulated by the upstream region of the first exon.
Department of Biochemistry, Faculty of Medicine, Mie University, 2-174 Edobashi, Tsu, Mie 514-8507, Japan. naoyan@doc.medic.mie-u.ac.jp
Naoya
N
Nakai
Masabumi
M
Kaneko
Nobuhiro
N
Nakao
Takahiko
T
Fujikawa
Kunio
K
Nakashima
Masato
M
Ogata
Minoru
M
Tanaka
D024506:5' Flanking Region / Q000235:genetics; D001483:Base Sequence; D018276:Carcinoma, Medullary / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D024202:Electrophoretic Mobility Shift Assay; D005091:Exons / Q000235:genetics; D054439:Ghrelin; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D036361:Peptide Hormones / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013961:Thyroid Gland / Q000378:metabolism*; D014158:Transcription, Genetic
D054439:Ghrelin; D009687:Nuclear Proteins; D036361:Peptide Hormones; D012333:RNA, Messenger; D008156:Luciferases