Mitochondrial DNA haplotypes induce differential patterns of DNA methylation that result in differential chromosomal gene expression patterns

Justin C St. John,Xin Sun,Te-Sha Tsai,Ian A Trounce,Jacqueline L Johnson,Daniel J Gough,Matthew McKenzie,William T Lee,Daniel J Garama,Jodee A Gould

doi:10.1038/cddiscovery.2017.62

Abstract

Mitochondrial DNA copy number is strictly regulated during development as naive cells differentiate into mature cells to ensure that specific cell types have sufficient copies of mitochondrial DNA to perform their specialised functions. Mitochondrial DNA haplotypes are defined as specific regions of mitochondrial DNA that cluster with other mitochondrial sequences to show the phylogenetic origins of maternal lineages. Mitochondrial DNA haplotypes are associated with a range of phenotypes and disease. To understand how mitochondrial DNA haplotypes induce these characteristics, we used four embryonic stem cell lines that have the same set of chromosomes but possess different mitochondrial DNA haplotypes. We show that mitochondrial DNA haplotypes influence changes in chromosomal gene expression and affinity for nuclear-encoded mitochondrial DNA replication factors to modulate mitochondrial DNA copy number, two events that act synchronously during differentiation. Global DNA methylation analysis showed that each haplotype induces distinct DNA methylation patterns, which, when modulated by DNA demethylation agents, resulted in skewed gene expression patterns that highlight the effectiveness of the new DNA methylation patterns established by each haplotype. The haplotypes differentially regulate α-ketoglutarate, a metabolite from the TCA cycle that modulates the TET family of proteins, which catalyse the transition from 5-methylcytosine, indicative of DNA methylation, to 5-hydroxymethylcytosine, indicative of DNA demethylation. Our outcomes show that mitochondrial DNA haplotypes differentially modulate chromosomal gene expression patterns of naive and differentiating cells by establishing mitochondrial DNA haplotype-specific DNA methylation patterns.

Highlights

The murine mitochondrial genome is a double-stranded, ~ 16.3 kb, circular genome.[1]
We have investigated whether chromosomal gene expression patterns can be altered in a haplotype-specific manner due to modulation of global DNA methylation patterns
We examined mitochondrial malate dehydrogenase 2 (MDH2) activity, as MDH2 occurs before α-KG in the tricarboxylic acid (TCA) cycle, and observed a corresponding fold change decrease in CC9dunni and CC9pahari cells with vitamin C (VitC) treatment (Figure 6f) but there was no change for 5-Aza (Figure 6g)

Summary

Introduction

The murine mitochondrial genome (mtDNA) is a double-stranded, ~ 16.3 kb, circular genome.[1]. Whilst the majority of the subunits of the electron transfer chain are encoded by the nuclear genome, each of the complexes, except for complex II, has one or more of its proteins encoded by mtDNA.[2] mtDNA encodes two rRNAs and 22 tRNAs and has one major non-coding region, the D-loop. The D-loop possesses two hypervariable regions that identify maternal relatives,[3] and is the site of interaction for the nuclear-encoded transcription and replication factors.[4]. MtDNA copy number is strictly regulated during development and differentiation.[5] The primordial germ cells possess ~ 200 copies of mtDNA,[6] which exponentially increase during oogenesis until the mature, fertilisable oocyte has 4150 000 copies.[7,8]. There is active reduction of mtDNA copy number until the blastocyst stage, the final stage of preimplantation development.[6,8] Whilst the blastocyst’s outer ring of cells, the trophectodermal cells, replicate mtDNA as they differentiate into the trophectoderm,[8] the inner cell mass cells, which form the embryo proper and are the source of embryonic stem (ES) cells, further reduce mtDNA copy number to establish the mtDNA set point.[8,9]

Methods

Results

Conclusion