DREF [DRE (DNA replication-related element) binding factor] is an 80 kDa polypeptide homodimer which plays an important role in regulating cell proliferation-related genes. Both DNA binding and dimer formation activities are associated with residues 16–115 of the N-terminal region. However, the mechanisms by which DREF dimerization and DNA binding are regulated remain unknown. Here, we report that the DNA binding activity of DREF is regulated by a redox mechanism, and that the cysteine residues are involved in this regulation. Electrophoretic mobility shift analysis using Drosophila Kc cell extracts or recombinant DREF proteins indicated that the DNA binding domain is sufficient for redox regulation. Site-directed mutagenesis and transient transfection assays showed that Cys59 and/or Cys62 are critical both for DNA binding and for redox regulation, whereas Cys91 is dispensable. In addition, experiments using Kc cells indicated that the DNA binding activity and function of DREF are affected by the intracellular redox state. These findings give insight into the exact nature of DREF function in the regulation of target genes by the intracellular redox state.
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March 2004
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Research Article|
March 15 2004
Redox regulation of DNA binding activity of DREF (DNA replication-related element binding factor) in Drosophila
Tae-Yeong CHOI;
Tae-Yeong CHOI
*Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735, Korea
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S. Young PARK;
S. Young PARK
*Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735, Korea
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Ho-Sung KANG;
Ho-Sung KANG
*Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735, Korea
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Jae-Hun CHEONG;
Jae-Hun CHEONG
*Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735, Korea
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Han-Do KIM;
Han-Do KIM
*Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735, Korea
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Bok Luel LEE;
Bok Luel LEE
†College of Pharmacy, Pusan National University, Busan 609-735, Korea
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Fumiko HIROSE;
Fumiko HIROSE
‡Department of Life Science, Graduate School of Science, Himeji Institute of Technology, 3-2-1 Koto, Kamigori, Hyogo 678-1297, Japan
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Masamitsu YAMAGUCHI;
Masamitsu YAMAGUCHI
§Division of Biotechnology, Faculty of Textile Sciences, Kyoto Institute of Technology, Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan
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Mi-Ae YOO
Mi-Ae YOO
1
*Department of Molecular Biology, College of Natural Science, Pusan National University, Busan 609-735, Korea
1To whom correspondence should be addressed (e-mail mayoo@pusan.ac.kr).
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Publisher: Portland Press Ltd
Received:
October 21 2003
Accepted:
December 02 2003
Accepted Manuscript online:
December 02 2003
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2004
2004
Biochem J (2004) 378 (3): 833–838.
Article history
Received:
October 21 2003
Accepted:
December 02 2003
Accepted Manuscript online:
December 02 2003
Citation
Tae-Yeong CHOI, S. Young PARK, Ho-Sung KANG, Jae-Hun CHEONG, Han-Do KIM, Bok Luel LEE, Fumiko HIROSE, Masamitsu YAMAGUCHI, Mi-Ae YOO; Redox regulation of DNA binding activity of DREF (DNA replication-related element binding factor) in Drosophila. Biochem J 15 March 2004; 378 (3): 833–838. doi: https://doi.org/10.1042/bj20031601
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