Regulation of nuclear epigenome by mitochondrial DNA heteroplasmy

Significance Mitochondrial DNA (mtDNA) mutations have been associated with common metabolic and degenerative disease phenotypes, implying a bioenergetic etiology for these diseases. For example, the mtDNA tRNALeu(UUR) m.3243A > G mutation manifests as diabetes, neurodegenerative disease, or lethal pediatric disease, depending on the percentage of mutant mtDNAs within the cell (heteroplasmy). Cultured cybrid cell lines harboring 3243G heteroplasmy levels corresponding to the different clinical phenotypes have distinct transcriptional profiles. Exhaustive metabolomic and histone posttranscriptional modification analysis of these 3243G cybrids revealed that changes in mtDNA heteroplasmy cause changes in mitochondrial intermediates and redox state, which result in distinctive histone modification changes. Thus, changes in the mitochondrial genotype change mitochondrial metabolism, which change the epigenome and transcriptome, which induce distinct clinical phenotypes.
Diseases associated with mitochondrial DNA (mtDNA) mutations are highly variable in phenotype, in large part because of differences in the percentage of normal and mutant mtDNAs (heteroplasmy) present within the cell. For example, increasing heteroplasmy levels of the mtDNA tRNALeu(UUR) nucleotide (nt) 3243A > G mutation result successively in diabetes, neuromuscular degenerative disease, and perinatal lethality. These phenotypes are associated with differences in mitochondrial function and nuclear DNA (nDNA) gene expression, which are recapitulated in cybrid cell lines with different percentages of m.3243G mutant mtDNAs. Using metabolic tracing, histone mass spectrometry, and NADH fluorescence lifetime imaging microscopy in these cells, we now show that increasing levels of this single mtDNA mutation cause profound changes in the nuclear epigenome. At high heteroplasmy, mitochondrially derived acetyl-CoA levels decrease causing decreased histone H4 acetylation, with glutamine-derived acetyl-CoA compensating when glucose-derived acetyl-CoA is limiting. In contrast, α-ketoglutarate levels increase at midlevel heteroplasmy and are inversely correlated with histone H3 methylation. Inhibition of mitochondrial protein synthesis induces acetylation and methylation changes, and restoration of mitochondrial function reverses these effects. mtDNA heteroplasmy also affects mitochondrial NAD+/NADH ratio, which correlates with nuclear histone acetylation, whereas nuclear NAD+/NADH ratio correlates with changes in nDNA and mtDNA transcription. Thus, mutations in the mtDNA cause distinct metabolic and epigenomic changes at different heteroplasmy levels, potentially explaining transcriptional and phenotypic variability of mitochondrial disease.