Segregation of mitochondrial DNA heteroplasmy through a developmental genetic bottleneck in human embryos
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina MiSeq
ORGANISM(S): Homo Sapiens
SUBMITTER: University of Cambridge
PROVIDER: PRJEB56092 | EVA | 2022-09-18
REPOSITORIES: EVA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| Single_cell_deep_sequencing_merged.vcf.csi | Other | |||
| Single_cell_deep_sequencing_merged.vcf.gz | Vcf | |||
| Single_cell_deep_sequencing_merged.vcf.gz.csi | Vcf |
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Project description:Mitochondrial DNA (mtDNA) mutations cause inherited diseases and are implicated in the pathogenesis of common late-onset disorders, but it is not clear how they arise and propagate in the humans. Here we show that mtDNA mutations are present in primordial germ cells (PGCs) within healthy female human embryos. Close scrutiny revealed the signature of selection against non-synonymous variants in the protein-coding region, tRNA gene variants, and variants in specific regions of the non-coding D-loop. In isolated single PGCs we saw a profound reduction in the cellular mtDNA content, with discrete mitochondria containing ~5 mtDNA molecules during early germline development. Single cell deep mtDNA sequencing showed rare variants reaching higher heteroplasmy levels in later PGCs, consistent with the observed genetic bottleneck, and predicting >80% levels within isolated organelles. Genome-wide RNA-seq showed a progressive upregulation of genes involving mtDNA replication and transcription, linked to a transition from glycolytic to oxidative metabolism. The metabolic shift exposes deleterious mutations to selection at the organellar level during early germ cell development. In this way, the genetic bottleneck prevents the relentless accumulation of mtDNA mutations in the human population predicted by Muller’s ratchet. Mutations escaping this mechanism will, however, show massive shifts in heteroplasmy levels within one human generation, explaining the extreme phenotypic variation seen in human pedigrees with inherited mtDNA disorders.
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Project description:We investigated the contentious role of mitochondrial reactive oxygen species (ROS) on mitochondrial DNA (mtDNA) quality and quantity in female reproductive aging. By conditionally knocking out the Sod2 gene in female mouse germline, we observed increased mitochondrial ROS and decreased oocyte quality, primarily due to impacts on OXPHOS complex II and mtDNA encoded mRNA levels. Interestingly, we found no increased mtDNA mutations, but alterations in mtDNA quantity, indicating the susceptibility of mtDNA to the mitochondrial ROS during reproductive aging. Notably, when we further decreased the basal level of mtDNA quantity by deactivating the mtSSB protein in Sod2 conditional knockout females, we observed an exacerbation of reproductive aging effects. This highlights the crucial role of mtDNA quantity in mitigating the impact of oxidative stress on fertility.
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Project description:We studied global distribution of hydroxymethylation (5hmC) and DNA methylation (5mC) in during normal B-cell development and CLL maturation. To derive disease-specific epigenetic changes we have compared CLL subgroups to their corresponding cell of origin. Our data showed that naïve B-cells, compared to non-class switched and class switched memory B-cells, had higher levels of 5-hmC and 5-mC. By comparing with ChIP sequencing data of H3K27Ac and H3K4me1 modifications of CLL samples with 5hmC and 5mC levels, we observed an increase of 5-hmC at overall gene body regions, CpG island shores and regulatory elements. Comparing 5-hmC distribution with RNA sequencing we showed that genes with higher transcriptional activity had higher 5-hmC and vice versa. Thus, our study suggests that the global loss of 5-hmC levels, with significant increase at gene regulatory regions, constitute a novel hallmark of CLL pathogenesis.
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Project description:This study characterized variations in the methylation profile of mitochondrial DNA (mtDNA) during initial bovine embryo development and correlated the presence of methylation with mtDNA transcription. Bovine oocytes were obtained from abattoir ovaries and submitted to in vitro culture procedures. Oocytes and embryos were collected at various stages (immature oocyte, IM; mature oocyte, MII; zygote, ZY; 4-cells, 4C; 16-cells, 16C and blastocysts, BL). Total DNA (including mtDNA) was used for Whole Genome Enzymatic Methyl Sequencing and for quantification of mtDNA copy number. Extracted RNA was used for quantification of mitochondrial transcripts (ND6, CYTB, tRNA-Phe and tRNA-Gln) using Droplet Digital PCR. The number of mtDNA copies per oocyte/embryo was found to be similar, while methylation levels in mtDNA varied among stages. Higher total methylation levels were found mainly at 4C and 16C. In specific gene regions, higher methylation levels were also observed at 4C and 16C (ND6, CYTB and tRNA-Phe), as well as an inverse correlation with the quantity of transcripts for these regions. This is a first description of epigenetic changes occurring in mtDNA during early embryonic development. Our results indicate that methylation might regulate the mtDNA transcription at a local level, particularly around the time of embryonic genome activation.
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Project description:Background: Low birth weight is associated with an increased adult metabolic disease risk. It is widely discussed that poor intrauterine conditions could induce long-lasting epigenetic modifications, leading to systemic changes in regulation of metabolic genes. In a unique cohort of 17 monozygotic (MZ) monochorionic female twins very discordant for birth weight (relative differences ranging from 21.3-35.7%), we examined if adverse prenatal growth conditions experienced by the smaller co-twins lead to systemic long-lasting DNA methylation changes. Genome-wide DNA methylation profiles were acquired from saliva DNA using the Infinium HumanMethylation450 BeadChip, targeting ~2% of all CpGs in the genome. Results: Overall, co-twins showed very similar genome-wide DNA methylation profiles. Since observed differences were almost exclusively caused by variable cellular composition, an original marker-based adjustment strategy was developed to eliminate such variation at affected CpGs. Among adjusted and unchanged CpGs 3153 were differentially methylated between the heavy and light co-twins at nominal significance (p<0.01), of which 45 showed absolute mean β-value differences >0.05 (max=0.08). Deep bisulfite sequencing of eight such loci revealed that differences remained in the range of technical variation, arguing against a reproducible biological effect. Analysis of methylation in repetitive elements using methylation-dependent primer extension assays also indicated no significant intra-pair differences. Conclusions: Severe intrauterine growth differences observed within these MZ twins are not associated with long-lasting DNA methylation differences in cells composing saliva, detectable with up-to-date technologies. Additionally, our results indicate that uneven cell type composition can lead to spurious results and should be addressed in epigenomic studies. DNA methylation profiles of saliva from 17 Adult Female MZ MC Twins discordant for birth weight.
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