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Optimized bisulfite sequencing analysis reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Mus musculus Homo sapiens

PROVIDER: GSE217117 | GEO | 2023/06/12

REPOSITORIES: GEO

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Optimized bisulfite sequencing analysis reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA.

Project description:BackgroundDNA methylation is one of the best characterized epigenetic modifications in the mammalian nuclear genome and is known to play a significant role in various biological processes. Nonetheless, the presence of 5-methylcytosine (5mC) in mitochondrial DNA remains controversial, as data ranging from the lack of 5mC to very extensive 5mC have been reported.ResultsBy conducting comprehensive bioinformatic analyses of both published and our own data, we reveal that previous observations of extensive and strand-biased mtDNA-5mC are likely artifacts due to a combination of factors including inefficient bisulfite conversion, extremely low sequencing reads in the L strand, and interference from nuclear mitochondrial DNA sequences (NUMTs). To reduce false positive mtDNA-5mC signals, we establish an optimized procedure for library preparation and data analysis of bisulfite sequencing. Leveraging our modified workflow, we demonstrate an even distribution of 5mC signals across the mtDNA and an average methylation level ranging from 0.19% to 0.67% in both cell lines and primary cells, which is indistinguishable from the background noise.ConclusionsWe have developed a framework for analyzing mtDNA-5mC through bisulfite sequencing, which enables us to present multiple lines of evidence for the lack of extensive 5mC in mammalian mtDNA. We assert that the data available to date do not support the reported presence of mtDNA-5mC.

| S-EPMC10403921 | biostudies-literature

Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [WGBS]

Project description:Whether 5mC exists in mitochondrial DNA is controversial, as data ranging from lack of 5mC to very extensive 5mC have been reported. By comprehensive bioinformatic analyses of published and our own data, we reveal that the observation of extensive and strand-biased mtDNA-5mC is artifact due to combinatorial effects of inefficient bisulfite conversion, extremely low sequencing reads in the L strand and interfering signals from nuclear mitochondrial DNA sequences (NUMTs).

2023-06-12 | GSE217116 | GEO

Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [TAPS]

2023-06-12 | GSE217114 | GEO

Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [TAPS]

Project description:Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [TAPS]

| PRJNA896966 | ENA

Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [WGBS]

Project description:Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [WGBS]

| PRJNA896967 | ENA

Optimized bisulfite sequencing analysis reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA

Project description:Optimized bisulfite sequencing analysis reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA

| PRJNA896964 | ENA

Accurate estimation of 5-methylcytosine in mammalian mitochondrial DNA.

Project description: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.

| S-EPMC5895755 | biostudies-literature

OxyBS: estimation of 5-methylcytosine and 5-hydroxymethylcytosine from tandem-treated oxidative bisulfite and bisulfite DNA.

Project description:UnlabelledThe use of sodium bisulfite (BS) treatment followed by hybridization to an Illumina Infinium BeadChip (HumanMethylation450 and MethylationEPIC) is a common method for interrogating 5-methylcytosine (5mC) at single nucleotide resolution. However, standard treatment of DNA with BS does not allow disambiguation of 5mC from an additional cytosine modification, 5-hydroxymethylcytosine (5hmC). Recently, it has been demonstrated that paired BS and oxidative bisulfite (oxBS) treatment on the same sample followed by hybridization to an Infinium microarray permits the differentiation of 5hmC from 5mC. Nevertheless, estimation of 5hmC and 5mC from tandem-treated arrays has been shown to produce irregular estimates of cytosine modifications.ResultsWe present a novel method using maximum likelihood estimation to accurately estimate the parameters of unmethylated cytosine (5C), 5mC and 5hmC from Infinium microarray data given the signal intensities from the oxBS and BS replicates.Availability and implementationOxyBS is an R package available on CRAN.ContactAndres.Houseman@oregonstate.eduSupplementary informationSupplementary data are available at Bioinformatics online.

| S-EPMC4978924 | biostudies-literature

Lack of 5-methylcytosine in Dictyostelium discoideum DNA.

Project description:We find no evidence for the presence of 5-methylcytosine in the DNA of Dictyostelium discoideum. Methylation was absent from CCGG sites in repetitive DNA and in DNA from the actin multigene family. Nor was 5-methylcytosine detected in total DNA when base composition was determined by means of h.p.l.c.

| S-EPMC1151219 | biostudies-other

oxBS-MLE: an efficient method to estimate 5-methylcytosine and 5-hydroxymethylcytosine in paired bisulfite and oxidative bisulfite treated DNA.

Project description:Motivation5-Methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) are important epigenetic regulators of gene expression. 5mC and 5hmC levels can be computationally inferred at single base resolution using sequencing or array data from paired DNA samples that have undergone bisulfite and oxidative bisulfite conversion. Current estimation methods have been shown to produce irregular estimates of 5hmC level or are extremely computation intensive.ResultsWe developed an efficient method oxBS-MLE based on binomial modeling of paired bisulfite and oxidative bisulfite data from sequencing or array analysis. Evaluation in several datasets showed that it outperformed alternative methods in estimate accuracy and computation speed.Availability and implementationoxBS-MLE is implemented in Bioconductor package ENmix.Contactniulg@ucmail.uc.eduSupplementary information: Supplementary data are available at Bioinformatics online.

| S-EPMC5181539 | biostudies-literature

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