Abstract
A hallmark of mammals is a diploid genome. Despite constraints from dosage compensation and imprinting, haploid embryonic stem cells can be established. However, rapid diploidization is observed in such cultures from mice, rats, and humans, limiting their use and indicating counterselection of a haploid genome. Here, we use metabolic profiling to discover that diploidization is triggered by an imbalance that arises from a smaller cytoplasmic volume and increased mitochondrial density. Reduced respiration causes a change in redox potential, leading to increased NADPH. Conversely, we demonstrate that NADPH oxidation in the mitochondria is sufficient to stabilize the haploid genome. We further show that the redox change leads to reduced AURORA kinase activation on chromosomes, connecting metabolic state to mitotic regulation. Our data, therefore, identify a mitochondrial metabolic imbalance as the root cause of diploidization and connect redox dysregulation to karyotypic instability.
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Data availability
The RNA seq data of haplod and diploid ESC samples generated in this study have been deposited in the Sequence Read Archive (SRA) database under accession code PRJNA1390253. The raw data underlying the figures are provided as a Source data file. G.D.M. or A.W. may be contacted for additional details on the protocols that support the findings of this study. Source data are provided with this paper.
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Acknowledgements
We acknowledge the Functional Genomics Center Zurich (FGCZ) and the Flow Cytometry Core Facility (FCCF) for technical support. We thank C. Ebner for the help with reagent preparations. We thank F. Mair, I. Vgenopoulou, A. Grison, and J. Corn for their helpful discussions. GDM was supported by the ETH Zurich Postdoctoral Fellowship Program as well as the Marie Curie Actions for People COFUND Program. This work was supported by grants from the Swiss National Science Foundation (SNF grants 31003A_152814/1 and 31003A_175643/1). T.K. was supported by the SNF grant 310030_215716.
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G.D.M., A.W. designed the research. G.D.M., A.R., K.H., T.F., T.K., and N.Z. collected the data. G.D.M., T.F., T.K., and N.Z. conducted software analysis. G.D.M., A.W. supervised the work. G.D.M., A.R., K.H., T.F., T.K., N.Z., and A.W. prepared and revised the manuscript.
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Di Minin, G., Rüegg, A.B., Halter, K. et al. Mitochondrial metabolic imbalance drives diploidization in mouse haploid embryonic stem cells via NADPH overload. Nat Commun (2026). https://doi.org/10.1038/s41467-026-70939-6
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DOI: https://doi.org/10.1038/s41467-026-70939-6
