Figure 1
Mitochondrial kinetics in the female germline. Throughout germline development, the number of mitochondrial DNA (mtDNA) molecules per cell varies from 105 - 106 in mature oocytes (before fertilization), 102 - 103 in primordial germ cells (PGCs) and 105 - 106 back to mature oocytes. This variation in copy number accounts for the mitochondrial genetic bottleneck, which forces segregation of mtDNA molecules. In line with this, the mitochondrial network is fragmented in oocytes, allowing efficient partitioning of mitochondria among hundreds of cells until embryonic implantation. In addition, only few cells in the embryo differentiate into PGCs, supporting a sampling effect towards selection of a single mtDNA genotype to populate the future oocyte.
Figure 2
Mitochondrial DNA inheritance in somatic and germ cells. Different mitochondria in a single somatic cell (A) are interconnected by constant events of fusion and fission, allowing them to share membranes, solutes, metabolites, proteins, RNAs and DNA (mitochondrial DNA – mtDNA). Hence, when a mutation in mtDNA arises, it can rapidly spread throughout the mitochondrial network. In this case, mutant (red circles) and wild-type (green circles) mtDNAs may co-exist, which is known as heteroplasmy. In comparison, homoplasmic mitochondria contain a single mtDNA genotype, either mutant or wild-type. Unless the mutation level exceeds a critical threshold necessary to cause a biochemical defect (i.e., above 60-90%; red mitochondria), the mutation effect will be masked by wild-type molecules (green mitochondria with both mutant and wild-type mtDNA). In germ cells (B), downregulation of fusion likely minimizes heteroplasmy within mitochondria, enhancing selection at the organellar level (i.e, stronger association between mitochondrial genotype and phenotype). In addition, decreased fusion leads to mitochondrial fragmentation, enhancing mtDNA segregation among embryonic cells. Hence, decreased levels of mtDNA in primordial germ cells (PGCs) makes possible selection at the cellular level (i.e., stronger association between mitochondrial genotype and cellular phenotype). Thus, as a result of selection against deleterious mutations, mature oocyte from the next generation may contain lower levels of mutant mtDNA.
Figure 3
New technologies for preventing inheritance of mitochondrial diseases. The mitochondrial replacement therapy (MRT; A) proposes the replacement of a patient’s mitochondria in oocytes by donor mitochondria. Towards that, mature oocytes arrested at the metaphase-II stage are collected from the patient and a donor. While the patient’s oocytes are supposed to contain mutant (red) mitochondrial DNA (mtDNA), donor oocytes should contain only wild-type (green) mtDNA. Next, the spindle is removed from the patient’s oocyte (donor karyoplast) to be injected into the donor oocyte from which the spindle was previously removed (donor cytoplast). Fertilization of the reconstructed oocyte should lead to a blastocyst, which can be used for embryonic stem cell (ESC) derivation. Although MRT allows transplantation of karyoplast with minimal carryover (~1%) of mutant mtDNA, recent data have provided evidence of a reversal in ESCs back to 100% mutant mtDNA (Hyslop et al., 2016Hyslop LA, Blakeley P, Craven L, Richardson J, Fogarty NME, Fragouli E, Lamb M, Wamaitha SE, Prathalingam N, Zhang Q et al. (2016) Towards clinical application of pronuclear transfer to prevent mitochondrial DNA disease. Nature 534:383–386.; Kang et al., 2016Kang E, Wu J, Gutierrez NM, Koski A, Tippner-Hedges R, Agaronyan K, Platero-Luengo A, Martinez-Redondo P, Ma H, Lee Y et al. (2016) Mitochondrial replacement in human oocytes carrying pathogenic mitochondrial DNA mutations. Nature 540:270–275.). An alternative strategy to MRT is the nuclease-mediated elimination of mutant mtDNA (B), which relies on the use of mitochondrial-targeted restriction endonucleases (mito-TALENs). These nucleases are designed to selectively cut mutant mtDNA, but not wild-type molecules. However, ~10% of targeted molecules were shown to be left uncut in newborns after use of mito-TALENs (Reddy et al., 2015Reddy P, Ocampo A, Suzuki K, Luo J, Bacman SR, Williams SL, Sugawara A, Okamura D, Tsunekawa Y, Wu J et al. (2015) Selective elimination of mitochondrial mutations in the germline by genome editing. Cell 161:459).
