TAILIEUCHUNG - Báo cáo khoa học: Identification of NF1 as a silencer protein of the human adenine nucleotide translocase-2 gene

The human adenine nucleotide translocase-2 (ANT2) pro-moter contains a silencer region that confers partial repres-sion on the heterologous herpes simplex virus thymidine kinase (HSVtk) promoter [Barath, P., Albert-Fournier, B., Luciakova, K., Nelson, . (1999)J. Biol. , 3378–3384]. Two sequences in the silencer (Site-2 and Site-3) are protected in the DNase I assayin vitro, and one of these is a repeated GTCCTG element previously shown to act as theactive repressor element. | Eur. J. Biochem. 271 1781-1788 2004 FEBS 2004 doi Identification of NF1 as a silencer protein of the human adenine nucleotide translocase-2 gene Peter Barath1 2 Daniela Poliakova1 2 Katarina Luciakova1 2 and B. Dean Nelson1 1Department of Biochemistry and Biophysics Arrhenius Laboratories Stockholm University Sweden 2Cancer Research Institute Slovak Academy of Sciences Bratislava Slovak Republic The human adenine nucleotide translocase-2 ANT2 promoter contains a silencer region that confers partial repression on the heterologous herpes simplex virus thymidine kinase HSVtk promoter Barath P. Albert-Fournier B. Luciakova K. Nelson . 1999 J. Biol. Chem. 274 3378-3384 . Two sequences in the silencer Site-2 and Site-3 are protected in the DNase I assay in vitro and one of these is a repeated GTCCTG element previously shown to act as the active repressor element. We have now purified the DNA binding protein and identified it using MALDI-TOF MS as a 33-kDa member of the nuclear factor 1 NF1 family of transcription factors. NF1 purified from rat liver and HeLa cell nuclei bind to both silencer Site-2 and Site-3 resulting in a DNase I footprint identical to that obtained with purified recombinant NF1. Furthermore transient transfection experiments with reporter constructs containing mutated silencer Site-2 and or Site-3 show that both sites contribute to repression of the HSVtk promoter. Finally chromatin immunoprecipitation analysis reveals that NF1 is bound to both elements on the endogenous HeLa cell ANT2 promoter. Our data support the belief that NF1 acts as a repressor when bound to silencing Site-2 and Site-3 of the ANT2 gene. Keywords adenine nucleotide translocase NF1 promoter regulation silencer protein transcription. Two major isoforms of the adenine nucleotide translocase ANT are expressed in mammalian cells. Both catalyse the exchange of mitochondrial ATP for cytosolic ADP thereby playing key roles in maintaining the cytosolic

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