Complex I heteroplasmy in MT-ND1 or MT-ND5 is enough to drop neuronal membrane potential and stall commitment

Verdict. Pathogenic heteroplasmy in mitochondrial ND1 or ND5, installed with a cytosine base editor, barely touches energy metabolism in stem cells. In induced neurons the same alleles drop membrane potential and push transcription away from neuronal commitment, even when heteroplasmy is relatively low.

Metal sites and redox

Complex I (NADH:ubiquinone oxidoreductase) carries FMN and eight iron-sulfur clusters in the N- and Q-modules. MT-ND1 and MT-ND5 encode proton-pumping ND subunits on the transmembrane arm, not those Fe-S proteins. This abstract does not name a cluster, a core oxidation level, or NAD+/NADH as the measured couple. The neuronal chemistry they report is loss of mitochondrial membrane potential and a transcriptional reprogramming of impaired commitment. Do not invent [4Fe-4S] involvement at ND1 or ND5. If Complex I electron transfer is the lesion, the Fe-S enzymes are in the matrix arm, upstream of the ND subunits that were edited.

What the authors report

Mitochondrial DNA variants, inherited or somatic, are particularly hard on cells that rely on mitochondrial metabolism, including neurons. Phenotype is thought to depend on heteroplasmy, but patient-derived cells cannot pin a result on one variant, and conventional genome editors do not work on mtDNA. The authors use DddA-derived cytosine base editor (DdCBE) to introduce pathogenic heteroplasmic variants into Neurogenin 2-inducible human iPSCs. They target two Complex I genes, MT-ND1 and MT-ND5, a frequent contributor to human disease including Leigh syndrome. Edited iPSC lines show low off-target editing and keep similar heteroplasmy after subclonal isolation and differentiation into NGN2-induced neurons. The Complex I variants do not substantially affect mitochondrial energy metabolism in undifferentiated iPSCs. They do reduce mitochondrial membrane potential in induced neurons, where they drive a transcriptional reprogramming indicative of impaired neuronal commitment. Heteroplasmic Complex I variants are therefore sufficient to disrupt neural fate specification even at relatively low heteroplasmy. More broadly, mtDNA editing in human iPSCs is offered as a platform for how mitochondrial fitness and heteroplasmy influence development and disease.

What was measured

DdCBE installation of MT-ND1 and MT-ND5 variants, off-target editing, heteroplasmy after subcloning and neuronal differentiation, mitochondrial energy metabolism in iPSCs, membrane potential in induced neurons, and the neuronal-commitment transcriptome. Exact heteroplasmy percentages and the metabolic assay in iPSCs are not named.

Caveats

Abstract only. Preprint. "Relatively low heteroplasmy" is unquantified here. iPSC metabolism is a negative result without a named assay. This is not a patient-derived Leigh line. Whether both ND alleles are phenocopies is unstated.

What to do with it

Treat Complex I ND heteroplasmy as a neurodevelopmental commitment problem that can be silent in iPSCs. The molecular test is the heteroplasmy fraction at which induced neurons lose membrane potential and leave the neuronal transcriptional program, and whether that fraction differs between ND1 and ND5.

Bibliographic record

Abstract-based editorial synthesis by Mitos. Not peer review.