Gene expression is a tightly regulated process that allows cells, tissues and ultimately whole organisms to thrive. At the heart of this regulation, transcription factors play a fundamental role in keeping things in check. However, over the last few decades, another level of transcriptional control, termed epigenetic regulation, has garnered extensive attention. In vertebrates, for example, epigenetic modification through cytosine methylation is thought to repress nuclear gene expression, and is implicated in the regulation of both normal and aberrant gene expression. However, most investigations to date have focused on epigenetic modifications of the nuclear genome, with little attention devoted to mitochondrial DNA. In their study, Lisa Shock, Prashant Thakkar, Erica Peterson, Richard Moran and Shirley Taylor, from the Virginia Commonwealth University, USA, specifically investigated the epigenetic control of mitochondrial gene expression. The team hypothesized that cytosine methylation of mitochondrial DNA plays an important role in the regulation of mitochondrial gene expression.

Initially, the authors examined the genomic region neighbouring the established coding sequence of an enzyme capable of methylating cytosine nucleotides in the nuclear genome, DNA methyltransferase 1 (DNMT1), to investigate whether this enzyme could be involved in mitochondrial DNA methylation. They discovered that a region upstream of the ‘nuclear form’ of DNMT1 encoded a sequence sufficient to import this protein into the mitochondrion, and that mammalian cells express both ‘nuclear’ and ‘mitochondrial’ DNMT1 transcripts and proteins.

Then the authors overexpressed two proteins (nuclear respiratory factor 1 and the peroxisome proliferator-activated γ coactivator-1α, which upregulate expression of nuclear encoded mitochondrial genes) in mammalian cells to explore the role of ‘mitochondrial’ DNMT1 in mitochondrial epigenetic regulation. The team found that when expressed together, these factors increased the abundance of DNMT1 in mitochondria fivefold, suggesting that ‘mitochondrial’ DNMT1 expression is responsive to endogenous factors regulating mitochondrial function.

In addition, the team looked at cells deficient in p53, a protein known to regulate mitochondrial respiration, to ensure that DNMT1 affected mitochondrial gene expression in a situation where mitochondrial function is altered. They observed a threefold increase in ‘mitochondrial’ DNMT1, which differentially affected mitochondrial gene expression, providing further evidence of a link between DNMT1 and changes in mitochondrial function. Then, the team used cells expressing DNMT1 with a tag allowing immunoprecipitation of the protein and found that DNMT1 coprecipitated with mitochondrial DNA, indicating their direct interaction in these cells. Finally, the authors established the presence of methylated nucleotides (5-methylcytosine and 5-hydroxymethylcytosine) in mitochondrial DNA, providing further confirmation of epigenetic regulation of the organelle.

Overall, these results confirm epigenetic modification of mitochondrial DNA through the action of DNMT1. In the mitochondria, this enzyme appears to methylate cytosine nucleotides, thereby affecting mitochondrial gene expression. In addition, this work adds to the complexity of the cross-talk between the nuclear and mitochondrial genomes necessary for cellular metabolic homeostasis. Although DNMT1 mitochondrial targeting sequence appears to be conserved across multiple mammalian species, it is not clear whether this conservation extends to other species. However, if this process is evolutionarily conserved, one can wonder how organisms that lack functional DNA methyltransferases (such as fruit flies) compensate for this deficiency.

L. S.
P. V.
E. J.
R. G.
S. M.
DNA methyltransferase 1, cytosine methylation, and cytosine hydroxymethylation in mammalian mitochondria
Proc. Natl. Acad. Sci. USA