Systematic analysis of NDUFAF6 in complex I assembly and mitochondrial disease

Andrew Y Sung; Rachel M Guerra; Laura H Steenberge; Charlotte L Alston; Kei Murayama; Yasushi Okazaki; Masaru Shimura; Holger Prokisch; Daniele Ghezzi; Alessandra Torraco; Rosalba Carrozzo; Agnès Rötig; Robert W Taylor; James L Keck; David J Pagliarini

doi:10.1038/s42255-024-01039-2

Systematic analysis of NDUFAF6 in complex I assembly and mitochondrial disease

Nat Metab. 2024 May 8. doi: 10.1038/s42255-024-01039-2. Online ahead of print.

Authors

Andrew Y Sung¹, Rachel M Guerra², Laura H Steenberge³, Charlotte L Alston^{4

5}, Kei Murayama^{6

7}, Yasushi Okazaki⁷, Masaru Shimura^{6

8}, Holger Prokisch^{8

9}, Daniele Ghezzi^{10

11}, Alessandra Torraco¹², Rosalba Carrozzo¹², Agnès Rötig¹³, Robert W Taylor^{4

5}, James L Keck¹, David J Pagliarini^{14

15

16}

Affiliations

¹ Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA.
² Department of Cell Biology and Physiology, Washington University School of Medicine in St. Louis, St. Louis, MO, USA.
³ Department of Biochemistry, University of Wisconsin-Madison, Madison, WI, USA.
⁴ Mitochondrial Research Group, Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK.
⁵ NHS Highly Specialised Service for Rare Mitochondrial Disorders, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK.
⁶ Department of Metabolism, Chiba Children's Hospital, Chiba, Japan.
⁷ Diagnostics and Therapeutic of Intractable Diseases, Intractable Disease Research Center, Graduate School of Medicine, Juntendo University, Tokyo, Japan.
⁸ Institute of Neurogenomics, Computational Health Center, Helmholtz Zentrum München, Neuherberg, Germany.
⁹ School of Medicine, Institute of Human Genetics, Technical University of Munich, Munich, Germany.
¹⁰ Department of Pathophysiology and Transplantation, University of Milan, Milan, Italy.
¹¹ Medical Genetics and Neurogenetics Unit, Fondazione IRCCS Instituto Neurologico Carlo Besta, Milan, Italy.
¹² Unit of Cell Biology and Diagnosis of Mitochondrial Disorders, Laboratory of Medical Genetics, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy.
¹³ Université Paris Cité, Imagine Institute, INSERM UMR 1163, Paris, France.
¹⁴ Department of Cell Biology and Physiology, Washington University School of Medicine in St. Louis, St. Louis, MO, USA. pagliarini@wustl.edu.
¹⁵ Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO, USA. pagliarini@wustl.edu.
¹⁶ Department of Genetics, Washington University School of Medicine, St. Louis, MO, USA. pagliarini@wustl.edu.

PMID: 38720117
DOI: 10.1038/s42255-024-01039-2

Abstract

Isolated complex I (CI) deficiencies are a major cause of primary mitochondrial disease. A substantial proportion of CI deficiencies are believed to arise from defects in CI assembly factors (CIAFs) that are not part of the CI holoenzyme. The biochemistry of these CIAFs is poorly defined, making their role in CI assembly unclear, and confounding interpretation of potential disease-causing genetic variants. To address these challenges, we devised a deep mutational scanning approach to systematically assess the function of thousands of NDUFAF6 genetic variants. Guided by these data, biochemical analyses and cross-linking mass spectrometry, we discovered that the CIAF NDUFAF6 facilitates incorporation of NDUFS8 into CI and reveal that NDUFS8 overexpression rectifies NDUFAF6 deficiency. Our data further provide experimental support of pathogenicity for seven novel NDUFAF6 variants associated with human pathology and introduce functional evidence for over 5,000 additional variants. Overall, our work defines the molecular function of NDUFAF6 and provides a clinical resource for aiding diagnosis of NDUFAF6-related diseases.

Abstract

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