Intra-individual purifying selection on mitochondrial DNA variants during human oogenesis

Abstract

Study question: Does selection for mtDNA mutations occur in human oocytes?

Summary answer: We provide statistical evidence in favor of the existence of purifying selection for mtDNA mutations in human oocytes acting between the expulsion of the first and second polar bodies (PBs).

What is known already: Several lines of evidence in Metazoa, including humans, indicate that variation within the germline of mitochondrial genomes is under purifying selection. The presence of this internal selection filter in the germline has important consequences for the evolutionary trajectory of mtDNA. However, the nature and localization of this internal filter are still unclear while several hypotheses are proposed in the literature.

Study design, size, duration: In this study, 60 mitochondrial genomes were sequenced from 17 sets of oocytes, first and second PBs, and peripheral blood taken from nine women between 38 and 43 years of age.

Participants/materials, setting, methods: Whole genome amplification was performed only on the single cell samples and Sanger sequencing was performed on amplicons. The comparison of variant profiles between first and second PB sequences showed no difference in substitution rates but displayed instead a sharp difference in pathogenicity scores of protein-coding sequences using three different metrics (MutPred, Polyphen and SNPs&GO).

Main results and the role of chance: Unlike the first, second PBs showed no significant differences in pathogenic scores with blood and oocyte sequences. This suggests that a filtering mechanism for disadvantageous variants operates during oocyte development between the expulsion of the first and second PB.

Large scale data: N/A.

Limitations, reasons for caution: The sample size is small and further studies are needed before this approach can be used in clinical practice. Studies on a model organism would allow the sample size to be increased.

Wider implications of the findings: This work opens the way to the study of the correlation between mtDNA mutations, mitochondrial capacity and viability of oocytes.

Study funding/competing interest(s): This work was supported by a SISMER grant. Laboratory facilities and skills were freely provided by SISMER, and by the Alma Mater Studiorum, University of Bologna. The authors have no conflict of interest to disclose.

Keywords: first and second polar body; heteroplasmy; human oocytes; mtDNA variants; purifying selection.

Figure 1

Polar bodies (PBs) have the same relative variant profile as blood with the exception of ND5. G-test statistics for each mitochondrial gene of PB changes using as expectation the relative change distribution among blood samples. Dashed line indicates 5% risk level with a Bonferroni correction, while dotted line indicates an uncorrected 5% risk level. Negative signs on top of bars indicate that observed values were lower than expected. Gray bar indicates noncoding or synonymous changes, while black bars indicate non-synonymous changes. The name of the genomic region follows the names used on accession NC_012920.1, with the only exception of HSV-I, HSV-II, HSV-III and D-Loop that split the original region annotated as D-Loop in the three hyper-variable regions and the remnant of the D-Loop, respectively.

Figure 2

Schematic interpretation of results. The different cell lines that lead to Oocyte (Ovo) and Blood have both, since their start, consistent amount of heteroplasmy. The different mitochondrial molecules have different number of substitutions (different line thickness) and level of functionality (high pathogenic scores darker lines). Molecules with high pathogenic scores are not allowed to reach mature oocyte by some selection mechanism (dashed line). Sequencing hide low heteroplasmy level per site in tissues with high molecular counts (gray disk).