Hee Jung Choi1, So Dam Jin1, Deivendran Rengaraj1, Jin Hwa Kim1, Bertrand Pain2, Jae Yong Han3,4. 1. Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea. 2. Univ Lyon, Universite ́Lyon 1, INSERM, INRAE, Stem Cell and Brain Research Institute, U1208, USC1361, 69500, Bron, France. 3. Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea. jaehan@snu.ac.kr. 4. Institute for Biomedical Sciences, Shinshu University, Minamiminowa, Nagano, 399-4598, Japan. jaehan@snu.ac.kr.
Abstract
BACKGROUND: NANOG is a core transcription factor (TF) in embryonic stem cells (ESCs) and primordial germ cells (PGCs). Regulation of the NANOG gene by TFs, epigenetic factors, and autoregulatory factors is well characterized in ESCs, and transcriptional regulation of NANOG is well established in these cells. Although NANOG plays a key role in germ cells, the molecular mechanism underlying its transcriptional regulation in PGCs has not been studied. Therefore, we investigated the mechanism that regulates transcription of the chicken NANOG (cNANOG) gene in PGCs and ESCs. RESULTS: We first identified the transcription start site of cNANOG by 5'-rapid amplification of cDNA ends PCR analysis. Then, we measured the promoter activity of various 5' flanking regions of cNANOG in chicken PGCs and ESCs using the luciferase reporter assay. cNANOG expression required transcriptional regulatory elements, which were positively regulated by POU5F3 (OCT4) and SOX2 and negatively regulated by TP53 in PGCs. The proximal region of the cNANOG promoter contains a positive transcriptional regulatory element (CCAAT/enhancer-binding protein (CEBP)-binding site) in ESCs. Furthermore, small interfering RNA-mediated knockdown demonstrated that POU5F3, SOX2, and CEBP played a role in cell type-specific transcription of cNANOG. CONCLUSIONS: We show for the first time that different trans-regulatory elements control transcription of cNANOG in a cell type-specific manner. This finding might help to elucidate the mechanism that regulates cNANOG expression in PGCs and ESCs.
BACKGROUND:NANOG is a core transcription factor (TF) in embryonic stem cells (ESCs) and primordial germ cells (PGCs). Regulation of the NANOG gene by TFs, epigenetic factors, and autoregulatory factors is well characterized in ESCs, and transcriptional regulation of NANOG is well established in these cells. Although NANOG plays a key role in germ cells, the molecular mechanism underlying its transcriptional regulation in PGCs has not been studied. Therefore, we investigated the mechanism that regulates transcription of the chicken NANOG (cNANOG) gene in PGCs and ESCs. RESULTS: We first identified the transcription start site of cNANOG by 5'-rapid amplification of cDNA ends PCR analysis. Then, we measured the promoter activity of various 5' flanking regions of cNANOG in chicken PGCs and ESCs using the luciferase reporter assay. cNANOGexpression required transcriptional regulatory elements, which were positively regulated by POU5F3 (OCT4) and SOX2 and negatively regulated by TP53 in PGCs. The proximal region of the cNANOG promoter contains a positive transcriptional regulatory element (CCAAT/enhancer-binding protein (CEBP)-binding site) in ESCs. Furthermore, small interfering RNA-mediated knockdown demonstrated that POU5F3, SOX2, and CEBP played a role in cell type-specific transcription of cNANOG. CONCLUSIONS: We show for the first time that different trans-regulatory elements control transcription of cNANOG in a cell type-specific manner. This finding might help to elucidate the mechanism that regulates cNANOGexpression in PGCs and ESCs.
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