Rewriting nuclear epigenetic scripts in mitochondrial diseases as a strategy for heteroplasmy control
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE300902 | GEO | 2025/07/02
REPOSITORIES: GEO
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Project description:Mitochondrial heteroplasmy, the presence of more than one mtDNA variant in a cell or individual is not as uncommon as previously thought. It is mostly due to the high mutation rate of the mtDNA and limited repair mechanisms present in the mitochondrion. The phenomenon has been studied mostly in human samples and in medical contexts. Heteroplasmy has also been researched in other species in fields such as forensics or genetic foot printing, but these studies usually focused on contained families within closely related species. Here we describe a large cross-species evaluation of heteroplasmy in mammals. We employed a novel approach to detect mitochondrial heteroplasmy in both novel and previously reported ChIP-sequencing datasets, which include concomitant mitochondrial DNA sequenced in the experiment. Here, we report novel ChIP-seq experiments for H3K4me1 and CEBPA across mammals, as well as some H3K4me3, H3K27ac and total histone H3 experiments. Most of the reported CEBPA experiments are good quality pull-downs, however the quality of many of the other experiments reported here has not been interrogated in detail. Whereas this does not affect the investigation of mitochondrial DNA pollution for the purposes of this study, both H3K4me1 and total histone H3 ChIP-seq datasets were often sequenced to relatively low depth and showed low ChIP enrichment compared to the other antibodies.
2016-05-13 | E-MTAB-3933 | biostudies-arrayexpress
Project description:Mitochondria generate signals of adaptation that regulate nuclear genes expression via retrograde signaling. But this phenomenon is complexified when qualitatively different mitochondria and mitochondrial DNA (mtDNA) coexist within cells. Although this cellular state of heteroplasmy leads to divergent phenotypes clinically, its consequences on cellular function and the cellular transcriptome are unknown. To interrogate this phenomenon, we generated somatic cell cybrids harboring increasing levels of a common mtDNA mutation (tRNALeu(UUR) 3243A>G) and mapped the resulting cellular phenotypes and transcriptional profiles across the complete range of heteroplasmy. Small increases in mutant mtDNAs caused relatively modest defect in mitochondrial oxidative capacity, but resulted in sharp transitions in mitochondrial ultrastructure and in the nuclear and mitochondrial transcriptomes, with the critical functional threshold corresponding to the induction of epigenetic regulatory systems. Principal component analysis underscores how each heteroplasmy level occupies a different "transcriptional space", with low levels heteroplasmy (20-30%) producing a dose-response linear progression in one direction, and mutationload of 50, 60 and 90% producing changes in the opposite direction. Hence, subtle changes in mitochondrial energetics can act through the epigenome to generate the phenotypes of the common “complex” diseases. Cells were generated by transferring the wildtype (3243A) and mutant (3243G) mtDNAs from a heteroplasmic 3243A>G patient’s lymphoblastoid cell line into 143B(TK-) mtDNA-deficient (ρo) cells and selected for transmitochondrial cybrids. Subsequent mtDNA depletion, reamplification, and cloning (Wiseman and Attardi, 1978) resulted in a series of stable cybrids harboring approximately 0, 20, 30, 50, 60, 90, and 100% 3243G mutant mtDNAs. Total RNA extracted from each cell line was then extracted, depleted of rRNA, and measured in sequenced in triplicates.
2014-09-02 | E-GEOD-56158 | biostudies-arrayexpress
Project description:Background: Cell free DNA (cfDNA) in plasma has received increasing attention and has been studied in a broad range of clinical conditions implicating inflammation, cancer, and aging. However, few studies have focused on mitochondrial DNA (mtDNA) in the cell free form. This study characterized the size distribution and sequence characteristics of plasma cell free mtDNA (cf mtDNA) in humans.Methods and Results: We optimized DNA isolation and next-generation sequencing library preparation protocols to better retain short DNA fragments from plasma, and applied these optimized methods to plasma samples from patients with sepsis. After massive parallel sequencing, we verified that our methods can retain substantially shorter DNA fragments than the standard isolation method, resulting in an average of 11.5 fold increase in short DNA fragments yield (DNA < 100bp). We report that cf mtDNA in plasma is highly enriched in short-size cfDNA (30 ~ 60 bp), which is much shorter than the value previously reported (~140 bp). Motivated by this unique size distribution, we size-selected short cfDNA fragments from the sequencing library, which further increased the mtDNA recovery rate by an average of 10.4 fold. Using this approach we detected mixtures of different mtDNA sequences, termed heteroplasmy, in plasma from 3 patients. In one patient who previously received bone marrow transplantation, different minor allele frequencies were observed between plasma and white blood cells (WBC) at heteroplasmic mtDNA sites, consistent with mixed-tissue origin for plasma DNA.Conclusion: mtDNA in plasma exists as very short fragments that exhibit mtDNA heteroplasmy distribution differences from that found in a single organ/tissue. This study is the first report of genome wide identification of mtDNA heteroplasmy in human plasma. Our optimized method can be used to investigate the potential utility of cf mtDNA fragments and heteroplasmy as biomarkers in various diseases.
2016-05-07 | GSE81178 | GEO
Project description:Mitochondrial DNA (mtDNA) 3243A>G tRNALeu(UUR) heteroplasmic mutation (m.3243A>G) exhibits clinically heterogeneous phenotypes. While the high mtDNA heteroplasmy exceeding a critical threshold causes mitochondrial encephalomyopathy, lactic acidosis with stroke-like episodes (MELAS) syndrome, the low mtDNA heteroplasmy causes maternally inherited diabetes with or without deafness (MIDD) syndrome. How quantitative differences in mtDNA heteroplasmy produces distinct pathological states has remained elusive. Here we show that despite striking similarities in the energy metabolic gene expression signature, the mitochondrial bioenergetics, biogenesis and fuel catabolic functions are distinct in cells harboring low or high levels of the m.3243A>G mutation compared to wild type cells. We further demonstrate that the low heteroplasmic mutant cells exhibit a coordinate induction of transcriptional regulators of the mitochondrial biogenesis, glucose and fatty acid metabolism pathways that lack in near homoplasmic mutant cells compared to wild type cells. Altogether, these results shed new biological insights on the potential mechanisms by which low mtDNA heteroplasmy may progressively cause diabetes mellitus.
2019-04-09 | GSE129091 | GEO
Project description:We evaluated here the physiological consequences of the generation of heteroplasmic embryos by mix of two wild type mtDNAs in the same zygote. In this animal model, mtDNA heteroplasmy is actively combated during germ-line transmission, embryonic development and somatic life of most differentiated cells. mtDNA heteroplasmy alone, even when both mtDNA types are individually non-pathogenic, causes a disease affecting mainly those tissues that are not able to reduce their heteroplasmy: heart, lung and skeletal muscle.
2020-01-01 | GSE93920 | GEO
Project description:Mitochondria generate signals of adaptation that regulate nuclear genes expression via retrograde signaling. But this phenomenon is complexified when qualitatively different mitochondria and mitochondrial DNA (mtDNA) coexist within cells. Although this cellular state of heteroplasmy leads to divergent phenotypes clinically, its consequences on cellular function and the cellular transcriptome are unknown. To interrogate this phenomenon, we generated somatic cell cybrids harboring increasing levels of a common mtDNA mutation (tRNALeu(UUR) 3243A>G) and mapped the resulting cellular phenotypes and transcriptional profiles across the complete range of heteroplasmy. Small increases in mutant mtDNAs caused relatively modest defect in mitochondrial oxidative capacity, but resulted in sharp transitions in mitochondrial ultrastructure and in the nuclear and mitochondrial transcriptomes, with the critical functional threshold corresponding to the induction of epigenetic regulatory systems. Principal component analysis underscores how each heteroplasmy level occupies a different "transcriptional space", with low levels heteroplasmy (20-30%) producing a dose-response linear progression in one direction, and mutationload of 50, 60 and 90% producing changes in the opposite direction. Hence, subtle changes in mitochondrial energetics can act through the epigenome to generate the phenotypes of the common “complex” diseases.
2014-09-02 | GSE56158 | GEO
Project description:Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled the precise introduction of base substitutions and the effective removal of genomes carrying harmful mutations. However, the reconstitution of mtDNA deletions responsible for severe mitochondrial disorders and age-related diseases has not yet been achieved in human cells. Here, we developed a method to engineer specific mtDNA deletions in human cells by co-expressing end-joining (EJ) machinery and targeted endonucleases. As a proof-of-concept, we used mito-EJ and mito-ScaI to generate a panel of clonal cell lines harboring a ~3.5 kb mtDNA deletion with the full spectrum of heteroplasmy. Investigating these isogenic cells revealed a critical threshold of ~75% deleted genomes, beyond which cells exhibited depletion of OXPHOS proteins, severe metabolic disruption, and impaired growth in galactose-containing media. Single-cell multiomic analysis revealed two distinct patterns of nuclear gene deregulation in response to mtDNA deletion accumulation; one triggered at the deletion threshold and another progressively responding to increasing heteroplasmy. In summary, the co-expression of mito-EJ and programable nucleases provides a powerful tool to model disease-associated mtDNA deletions in different cell types. Establishing a panel of cell lines with a large-scale deletion at varying levels of heteroplasmy is a valuable resource for understanding the impact of mtDNA deletions on diseases and guiding the development of potential therapeutic strategies.
2025-03-10 | GSE276718 | GEO
Project description:Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled the precise introduction of base substitutions and the effective removal of genomes carrying harmful mutations. However, the reconstitution of mtDNA deletions responsible for severe mitochondrial disorders and age-related diseases has not yet been achieved in human cells. Here, we developed a method to engineer specific mtDNA deletions in human cells by co-expressing end-joining (EJ) machinery and targeted endonucleases. As a proof-of-concept, we used mito-EJ and mito-ScaI to generate a panel of clonal cell lines harboring a ~3.5 kb mtDNA deletion with the full spectrum of heteroplasmy. Investigating these isogenic cells revealed a critical threshold of ~75% deleted genomes, beyond which cells exhibited depletion of OXPHOS proteins, severe metabolic disruption, and impaired growth in galactose-containing media. Single-cell multiomic analysis revealed two distinct patterns of nuclear gene deregulation in response to mtDNA deletion accumulation; one triggered at the deletion threshold and another progressively responding to increasing heteroplasmy. In summary, the co-expression of mito-EJ and programable nucleases provides a powerful tool to model disease-associated mtDNA deletions in different cell types. Establishing a panel of cell lines with a large-scale deletion at varying levels of heteroplasmy is a valuable resource for understanding the impact of mtDNA deletions on diseases and guiding the development of potential therapeutic strategies.
2025-03-10 | GSE276699 | GEO
Project description:Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled the precise introduction of base substitutions and the effective removal of genomes carrying harmful mutations. However, the reconstitution of mtDNA deletions responsible for severe mitochondrial disorders and age-related diseases has not yet been achieved in human cells. Here, we developed a method to engineer specific mtDNA deletions in human cells by co-expressing end-joining (EJ) machinery and targeted endonucleases. As a proof-of-concept, we used mito-EJ and mito-ScaI to generate a panel of clonal cell lines harboring a ~3.5 kb mtDNA deletion with the full spectrum of heteroplasmy. Investigating these isogenic cells revealed a critical threshold of ~75% deleted genomes, beyond which cells exhibited depletion of OXPHOS proteins, severe metabolic disruption, and impaired growth in galactose-containing media. Single-cell multiomic analysis revealed two distinct patterns of nuclear gene deregulation in response to mtDNA deletion accumulation; one triggered at the deletion threshold and another progressively responding to increasing heteroplasmy. In summary, the co-expression of mito-EJ and programable nucleases provides a powerful tool to model disease-associated mtDNA deletions in different cell types. Establishing a panel of cell lines with a large-scale deletion at varying levels of heteroplasmy is a valuable resource for understanding the impact of mtDNA deletions on diseases and guiding the development of potential therapeutic strategies.
2025-03-10 | GSE276696 | GEO
Project description:Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled the precise introduction of base substitutions and the effective removal of genomes carrying harmful mutations. However, the reconstitution of mtDNA deletions responsible for severe mitochondrial disorders and age-related diseases has not yet been achieved in human cells. Here, we developed a method to engineer specific mtDNA deletions in human cells by co-expressing end-joining (EJ) machinery and targeted endonucleases. As a proof-of-concept, we used mito-EJ and mito-ScaI to generate a panel of clonal cell lines harboring a ~3.5 kb mtDNA deletion with the full spectrum of heteroplasmy. Investigating these isogenic cells revealed a critical threshold of ~75% deleted genomes, beyond which cells exhibited depletion of OXPHOS proteins, severe metabolic disruption, and impaired growth in galactose-containing media. Single-cell multiomic analysis revealed two distinct patterns of nuclear gene deregulation in response to mtDNA deletion accumulation; one triggered at the deletion threshold and another progressively responding to increasing heteroplasmy. In summary, the co-expression of mito-EJ and programable nucleases provides a powerful tool to model disease-associated mtDNA deletions in different cell types. Establishing a panel of cell lines with a large-scale deletion at varying levels of heteroplasmy is a valuable resource for understanding the impact of mtDNA deletions on diseases and guiding the development of potential therapeutic strategies.
2025-03-10 | GSE276694 | GEO