Mitochondrial membrane hyperpolarization modulates nuclear DNA methylation and gene expression through phospholipid remodeling [RNA-Seq]
Maintenance of the mitochondrial inner membrane potential (??m) is critical for many aspects of mitochondrial function. While ??m loss and its consequences are well studied, little is known about the effects of mitochondrial hyperpolarization. In this study, we used cells deleted of ATP5IF1 (IF1), a natural inhibitor of the hydrolytic activity of the ATP synthase, as a genetic model of increased resting ??m. We found that the nuclear DNA hypermethylates when the ??m is chronically high, regulating the transcription of mitochondrial, carbohydrate and lipid genes. These effects can be reversed by decreasing the ??m and recapitulated in wild-type (WT) cells exposed to environmental chemicals that cause hyperpolarization. Surprisingly, phospholipid changes, but not redox or metabolic alterations, linked the ??m to the epigenome. Sorted hyperpolarized WT and ovarian cancer cells naturally depleted of IF1 also showed phospholipid remodeling, indicating this as an adaptation to mitochondrial hyperpolarization. These data provide a new framework for how mitochondria can impact epigenetics and cellular biology to influence health outcomes, including through chemical exposures and in disease states. Overall design: We aimed to investigate the transcriptome of a genetic model of mitochondrial membrane potential hyperpolarization (increased ??m). RNA from the following treatment groups were isolated from n=3 biologically independent replicates: 1) HEK293T cells containing a doxycycline-inducible dominant negative DNA polymerase gamma (WT); 2) the isogenic cell line knockout for the gene ATP5IF1 (IF1-KO_NOV0082); 3) IF1-KO expressing GFP (ATP_EVgfp), 4) the IF1-KO isogenic cell line overexpressing ectopic IF1 with GFP (ATP_cDNAgfp); 5) IF1-KO cells transfected with pcDNA3.1_empty vector (IF1-KO); 6) IF1_KO cells transfected with pcDNA3.1-UCP4(SLC25A27)-NE vector overexpressing UCP4 (IF1_KO_UCP4). RNA was quantified using Nanodropand, and for quality control, samples were re-assessed using Qubit 3.0. RNA integrity was confirmed using Agilent RNA 6000 Nano Kit in 2100 Bioanalyzer (Agilent).