TY - JOUR
T1 - Accurate estimation of 5-methylcytosine in mammalian mitochondrial DNA
AU - Matsuda, Shigeru
AU - Yasukawa, Takehiro
AU - Sakaguchi, Yuriko
AU - Ichiyanagi, Kenji
AU - Unoki, Motoko
AU - Gotoh, Kazuhito
AU - Fukuda, Kei
AU - Sasaki, Hiroyuki
AU - Suzuki, Tsutomu
AU - Kang, Dongchon
N1 - Funding Information:
We thank Dr. H. Koseki, RIKEN for E14 ESCs; Dr. M. Okano, Kumamoto Univ. for TKO ESCs; Prof. K. Hayashi, Kyushu Univ. for STO cell line; Ms. N. Nomiyama, Kyushu Univ. for technical assistance; Dr. G. Nagamatsu, Kyushu Univ. for technical advice and Dr. H. Toh, Kyushu Univ. for bioinformatics analysis. This work was supported in part by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science [JSPS KAKENHI grant numbers JP17K07504 to T.Y., JP25112010 to H.S. and JP17H01550 to D.K.] and by the Cooperative Research Project Program of the Medical Institute of Bioregulation, Kyushu University to T.Y.
Publisher Copyright:
© 2018 The Author(s).
PY - 2018/12/1
Y1 - 2018/12/1
N2 - Whilst 5-methylcytosine (5mC) is a major epigenetic mark in the nuclear DNA in mammals, whether or not mitochondrial DNA (mtDNA) receives 5mC modification remains controversial. Herein, we exhaustively analysed mouse mtDNA using three methods that are based upon different principles for detecting 5mC. Next-generation bisulfite sequencing did not give any significant signatures of methylation in mtDNAs of liver, brain and embryonic stem cells (ESCs). Also, treatment with methylated cytosine-sensitive endonuclease McrBC resulted in no substantial decrease of mtDNA band intensities in Southern hybridisation. Furthermore, mass spectrometric nucleoside analyses of highly purified liver mtDNA preparations did not detect 5-methyldeoxycytidine at the levels found in the nuclear DNA but at a range of only 0.3-0.5% of deoxycytidine. Taken together, we propose that 5mC is not present at any specific region(s) of mtDNA and that levels of the methylated cytosine are fairly low, provided the modification occurs. It is thus unlikely that 5mC plays a universal role in mtDNA gene expression or mitochondrial metabolism.
AB - Whilst 5-methylcytosine (5mC) is a major epigenetic mark in the nuclear DNA in mammals, whether or not mitochondrial DNA (mtDNA) receives 5mC modification remains controversial. Herein, we exhaustively analysed mouse mtDNA using three methods that are based upon different principles for detecting 5mC. Next-generation bisulfite sequencing did not give any significant signatures of methylation in mtDNAs of liver, brain and embryonic stem cells (ESCs). Also, treatment with methylated cytosine-sensitive endonuclease McrBC resulted in no substantial decrease of mtDNA band intensities in Southern hybridisation. Furthermore, mass spectrometric nucleoside analyses of highly purified liver mtDNA preparations did not detect 5-methyldeoxycytidine at the levels found in the nuclear DNA but at a range of only 0.3-0.5% of deoxycytidine. Taken together, we propose that 5mC is not present at any specific region(s) of mtDNA and that levels of the methylated cytosine are fairly low, provided the modification occurs. It is thus unlikely that 5mC plays a universal role in mtDNA gene expression or mitochondrial metabolism.
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U2 - 10.1038/s41598-018-24251-z
DO - 10.1038/s41598-018-24251-z
M3 - Article
C2 - 29643477
AN - SCOPUS:85045269740
SN - 2045-2322
VL - 8
JO - Scientific Reports
JF - Scientific Reports
IS - 1
M1 - 5801
ER -