Abstract
Comparison of mitochondrial DNA (mtDNA) methylation patterns in oocytes, blastocysts and ovarian granulosa cells indicates hitherto unsuspected dynamics. Oocytes and blastocysts recovered from cows subjected to ovarian stimulation and from non-stimulated abattoir ovaries were analyzed using bisulphite transformation of DNA followed by whole genome sequencing. The cow is a recognized as a good model for human oocyte and pre-implantation development. The number of mtDNA copies is high in oocytes (200,000–400,000) and early embryos, resulting in very high coverage (>3000x) and very low p values for each of 716 cytosine-based nucleosides. Methylation ratio was lowest in oocytes, following by blastocysts then granulosa cells and was not restricted to CG sites but was found also at CHG and CHH sites. The initial methylation pattern is conserved during the first week of life but not in somatic cells. RNA analysis of mitochondria encoded genes showed a significant inverse correlation between methylation and expression for almost all sequences. Methylation was more extensive in somatic tissues from mature animals than in immature pre-pubertal animals. Our findings suggest that mtDNA methylation might play a programming role during gametogenesis and would be subject to epigenetic regulation according to environment and/or maternal maturity.
Highlights
Comparison of mitochondrial DNA methylation patterns in oocytes, blastocysts and ovarian granulosa cells indicates hitherto unsuspected dynamics
The results indicate that bovine mitochondrial DNA shows a unique signature in oocytes and blastocysts compared to the associated somatic cells; an inverted methylation-expression relation; and a specific pattern of mtDNA methylation associated with oocyte status/quality that may reflect programming of embryonic development potential
Because of the size of mitochondrial DNA, we obtained significantly higher coverage of each cytosine (719 nucleoside sites) above 3,000-fold in oocytes samples compared to what is typically obtained for somatic cells (4-fold; only 38% of the cytosine were covered)
Summary
Comparison of mitochondrial DNA (mtDNA) methylation patterns in oocytes, blastocysts and ovarian granulosa cells indicates hitherto unsuspected dynamics. Mitochondria play an important role in all animal cells as a metabolic energy provider, as well as a regulator of many cellular processes including apoptosis or cell suicide[1] They are present in all tissues, including in oocytes where they go through a selection bottleneck to ensure the passage of the most functional organelles to the generation[2]. In the case of mammalian cells, mitochondrial DNA (mtDNA) encodes 13 protein components of the oxidative phosphorylation system (subunits of complexes I, III, IV, and V), as well as mitochondria-specific small and large ribosomal RNA fragments and an array of 22 transfer RNA molecules[6]. Www.nature.com/scientificreports reproductive tissues, they are rarely studied except in association with specific mitochondrial dysfunctions or mutations in humans[11]
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