Mitochondrial DNA copy number reduction via in vitro TFAM knockout remodels the nuclear epigenome and transcriptome

Mitochondrial DNA copy number (mtDNA-CN) is associated with several age-related chronic diseases and is a predictor of all-cause mortality. Here, we examine site-specific differential nuclear DNA (nDNA) methylation and differential gene expression resulting from in vitro reduction of mtDNA-CN to uncover shared genes and biological pathways mediating the effect of mtDNA-CN on disease. Epigenome and transcriptome profiles were generated for three independent human embryonic kidney (HEK293T) cell lines harboring a mitochondrial transcription factor A (TFAM) knockout generated via CRISPR-Cas9, and matched control lines. We identified 2924 differentially methylated sites, 67 differentially methylated regions, and 102 differentially expressed genes associated with mtDNA-CN. Integrated analysis uncovered 24 Gene-CpG pairs. GABAA receptor genes and related pathways, the neuroactive ligand signaling pathway, ABCD1/2 gene activity, and cell signaling processes were overrepresented, providing insight into the underlying biological mechanisms facilitating these associations. We also report evidence implicating chromatin state regulatory mechanisms as modulators of mtDNA-CN effect on gene expression. We demonstrate that mitochondrial DNA variation signals to the nuclear DNA epigenome and transcriptome and may lead to nuclear remodeling relevant to development, aging, and complex disease. The amount of mitochondrial DNA, called mitochondrial DNA copy number (mtDNA-CN), has been linked with risk for various health conditions and diseases. To better understand this relationship, we studied the association between mtDNA-CN reduction and other DNA changes within the cell, specifically those in the cell nucleus. Using a cell model, we compared changes in cells with lower mtDNA-CN to normal cells. We noticed several patterns of changes that point to different genomic regions, communication processes, and signaling pathways within the cells that may be involved in the association of mtDNA-CN with disease development. Our findings help explain how the mitochondria interact with the nucleus in the context of complex disease.