Random mtDNA mutagenesis maps genotype-phenotype associations across metabolic contexts and lineages
- 1Memorial Sloan Kettering Cancer Center
Abstract
Mitochondrial DNA (mtDNA) mutations cause inherited disease, accumulate with age, and recur in tumors, yet their effects remain unresolved because existing models interrogate one engineered allele at a time. We developed mtDIVE, a random C[->]T mutagenesis platform for pooled mitochondrial forward genetics in human epithelial cells and zebrafish embryos. mtDIVE generates genotypes ranging from single substitutions to >100 mutations per molecule, impairs respiration, and causes lineage-specific developmental delay. Coupling single-cell transcriptomics with mtDNA genotyping revealed that selection on mutant genomes and their transcriptional consequences depend on metabolic state and developmental lineage. Obligate OXPHOS eliminates highly mutated cells by apoptosis, whereas glycolytic conditions promote their competitive displacement. Individual loci exert modest but non-redundant effects that predict cell state beyond aggregate mutation burden, and cancer-recurrent variants are enriched among mutations linked to reduced mitochondrial transcription. Thus, mtDIVE links mtDNA genotypes to cellular phenotypes, separating the consequences of mutation identity from mutational load.
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BibTeX
@misc{giladi2026random,
title = {{Random mtDNA mutagenesis maps genotype-phenotype associations across metabolic contexts and lineages}},
author = {A. Giladi and R. Baik and C.-Y. Chung and M. Kim and J. Adams and R. Saha and M. Land and R. Cui and E. DeBitetto and M. E. Kelley and C. B. Thompson and E. Reznik and R. Sharma and R. Challinge and S. R. Isaac and J. Peterson and A. van Oudenaarden and A. Sfeir},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.08.757705},
url = {https://www.biorxiv.org/content/10.64898/2026.10.08.757705v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Random mtDNA mutagenesis maps genotype-phenotype associations across metabolic contexts and lineages
AU - A. Giladi
AU - R. Baik
AU - C.-Y. Chung
AU - M. Kim
AU - J. Adams
AU - R. Saha
AU - M. Land
AU - R. Cui
AU - E. DeBitetto
AU - M. E. Kelley
AU - C. B. Thompson
AU - E. Reznik
AU - R. Sharma
AU - R. Challinge
AU - S. R. Isaac
AU - J. Peterson
AU - A. van Oudenaarden
AU - A. Sfeir
PY - 2026
DA - 2026/10/08
PB - bioRxiv
DO - 10.64898/2026.10.08.757705
UR - https://www.biorxiv.org/content/10.64898/2026.10.08.757705v1
AB - Mitochondrial DNA (mtDNA) mutations cause inherited disease, accumulate with age, and recur in tumors, yet their effects remain unresolved because existing models interrogate one engineered allele at a time. We developed mtDIVE, a random C[->]T mutagenesis platform for pooled mitochondrial forward genetics in human epithelial cells and zebrafish embryos. mtDIVE generates genotypes ranging from single substitutions to >100 mutations per molecule, impairs respiration, and causes lineage-specific developmental delay. Coupling single-cell transcriptomics with mtDNA genotyping revealed that selection on mutant genomes and their transcriptional consequences depend on metabolic state and developmental lineage. Obligate OXPHOS eliminates highly mutated cells by apoptosis, whereas glycolytic conditions promote their competitive displacement. Individual loci exert modest but non-redundant effects that predict cell state beyond aggregate mutation burden, and cancer-recurrent variants are enriched among mutations linked to reduced mitochondrial transcription. Thus, mtDIVE links mtDNA genotypes to cellular phenotypes, separating the consequences of mutation identity from mutational load.
ER -