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Regulation of mitochondrial D‐loops by transcription factor A and single‐stranded DNA‐binding protein

Chihiro Takamatsu, Shuyo Umeda, Takashi Ohsato, Tetsuji Ohno, Yoshito Abe, Atsushi Fukuoh, Hideo Shinagawa, Naotaka Hamasaki, Dongchon Kang

Author Affiliations

  1. Chihiro Takamatsu1,
  2. Shuyo Umeda1,
  3. Takashi Ohsato1,
  4. Tetsuji Ohno1,
  5. Yoshito Abe1,
  6. Atsushi Fukuoh2,3,
  7. Hideo Shinagawa3,
  8. Naotaka Hamasaki1 and
  9. Dongchon Kang (kang{at}mailserver.med.kyushu-u.ac.jp)*,1
  1. 1 Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Graduate School of Medical Sciences, Fukuoka, 812‐8582, Japan
  2. 2 Department of Microbiology, Asahikawa Medical College, Midorigaoka‐Higashi, Asahikawa, 078‐8510, Japan
  3. 3 Department of Molecular Microbiology, Research Institute for Microbial Diseases, Osaka University, Suita, 565‐0871, Japan
  1. * Tel: +81 92 642 5749; Fax: +81 92 642 5772; E‐mail: kang{at}mailserver.med.kyushu-u.ac.jp

Abstract

During replication, mitochondrial DNA (mtDNA) takes on a triple‐stranded structure called a D‐loop. Although their physiological roles are not understood, D‐loops are implicated in replication and transcription of mtDNA. Little is known about the turnover of D‐loops. We investigated the effects of mitochondrial transcription factor A (TFAM) and single‐stranded DNA‐binding protein (mtSSB) on D‐loops. In human HeLa cells, TFAM and mtSSB are, respectively, 1700‐ and 3000‐fold more abundant than mtDNA. This level of TFAM is two orders of magnitude higher than reported previously and is sufficient to wrap human mtDNA entirely. TFAM resolves D‐loops in vitro if added in similar stoichiometries. mtSSB inhibits the resolution of mtDNA by TFAM but enhances resolution by RecG, a junction‐specific helicase from Escherichia coli. Hence, mtSSB functions in both stabilization and resolution. We propose that TFAM and mtSSB are cooperatively involved in stabilizing D‐loops and in the maintenance of mtDNA.

  • Received August 17, 2001.
  • Revision received January 22, 2002.
  • Accepted March 20, 2002.