Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Genomic Organization and Promoter Characterization of the Murine Glial Cell-derived Neurotrophic Factor Inducible Transcription Factor (mGIF) Gene

생쥐 신경교세포 유래 신경영양인자 유도성 전사인자 (mGIF) 유전자의 유전체 구조 및 프로모터 특성 분석

  • Kim, Ok-Soo (Department of Pharmaceutical Engineering, College of Medical Life Science, University) ;
  • Kim, Yong-Man (FCB-Pharmicell Co., LTD) ;
  • Kim, Nam-Young (Department of Pharmaceutical Engineering, College of Medical Life Science, University) ;
  • Lee, Eo-Jin (Department of Pharmaceutical Engineering, College of Medical Life Science, University) ;
  • Jang, Min-Kyung (Department of Pharmaceutical Engineering, College of Medical Life Science, University) ;
  • Lee, Dong-Geun (Department of Pharmaceutical Engineering, College of Medical Life Science, University) ;
  • Lee, Sang-Hyeon (Department of Pharmaceutical Engineering, College of Medical Life Science, University)
  • 김옥수 (신라대학교 의생명과학대학 제약공학과) ;
  • 김용만 (FCB-파미셀(주)) ;
  • 김남영 (신라대학교 읫애명과학대학 제약공학과) ;
  • 이어진 (신라대학교 의생명과학대학 제약공학과) ;
  • 장민경 (신라대학교 의생명과학대학 제약공학과) ;
  • 이동근 (신라대학교 의생명과학대학 제약공학과) ;
  • 이상현 (신라대학교 의생명과학대학 제약공학과)
  • Published : 2007.02.28
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Abstract

To study the transcriptional mechanisms by which expression of the murine glial cell-derived neurotrophic factor inducible transcription factor (mGIF) gene is regulated, a murine genomic clone was iso-lated using a mGIF cDNA as probe. A 13-kb genomic fragment, which comprises 4-kb upstream of the transcription initiation site was sequenced. The promoter region lacks a TATA box and CAAT box, is rich in G+C content, and has multiple putative binding sites for the transcription factor Spl. The mGIF gene also has consensus sequences for AP2 binding sites. The transcriptional activity of five deletion mutants of a 2.1-kb fragment was analyzed by modulating transcription of the heterologous luciferase gene in the promoterless plasmid pGL2-Basic. All mutants showed significant transcriptional activity in the murine neuroblastoma cell line NB41A3. Transient expression assays suggested the presence of a positive regulator between -213 and -129 while a negative regulator was found in the region between -806 and -214. Relatively strong transcriptional activity was observed in neuronal NB41A3, glial C6 cells and hepatic HepG2, but very weak activity in skeletal muscle C2C12 cells. These findings confirm the tissue-specific activity of the mGIF promoter and suggest that this gene shares structural and functional similarities with the dopamine receptor genes that it regulates.

생쥐 신경교세포 유래 신경영양인자 유도성 전사인자(mGIF)의 발현조절에 필요한 전사기작을 연구하기 위하여 mGIF cDNA를 탐침자로 이용하여 genomic clone을 분리하였다. 전체 유전자 13-kb 영역 중 전사개시점에서 4-kb 상류영역의 유전자 서열을 파악한 결과, 프로모터 영역에서 TATA box와 CAAT box는 발견할 수 없었으며 G+C content는 높은 것으로 나타났고 여러 개의 Sp1 전사인자 결합영역이 있었다. 또한 mGIF 유전자는 AP2 결합에 필요한 보존적 영역이 있었다. mGIF 유전자의 프로모터 영역의 단편들을 프로모터가 없는 pGL2-Basic 플라스미드의 luciferase 유전자의 상류에 연결하여 서로 다른 5종류의 결손 돌연변이체를 제조하고 NB41A3 세포주를 이용하여 전사활성을 측정하였다. Transient expression assays 결과, 모든 결손 돌연변이체에서 전사활성이 나타났으며 -213과 -129사이에 전사촉진 영역이 존재하며 -806과 -214사이에 전사억제 영역이 있는 것으로 나타났다. 신경세포주인 NB41A3과 신경교세포주인 C6 그리고 간세포주인 HepG2에서 mGIF 유전자 프로모터의 높은 활성이 관찰되었으며, 근육세포주인 C2C12에서는 낮은 활성이 관찰되었다. 따라서 mGIF 유전자는 조직특이적으로 발현하며 도파민 수용체 유전자와 구조적, 기능적 유사성이 있는 것을 알 수 있었다.

Keywords

References

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