RT Journal Article SR Electronic T1 MFN2 Influences Amyotrophic Lateral Sclerosis Pathology JF bioRxiv FD Cold Spring Harbor Laboratory SP 2021.10.30.466517 DO 10.1101/2021.10.30.466517 A1 Kristi L. Russell A1 Jonathan M. Downie A1 Summer B. Gibson A1 Karla P. Figueroa A1 Cody J. Steely A1 Mark B. Bromberg A1 L. Charles Murtaugh A1 Lynn B. Jorde A1 Stefan M. Pulst YR 2021 UL http://biorxiv.org/content/early/2021/10/31/2021.10.30.466517.abstract AB Objective To better understand the pathology of amyotrophic lateral sclerosis, we used sequence data from patients seen at the University of Utah to identify novel disease-associated loci. We utilized both in vitro and in vivo studies to determine the biological effect of patient mutations in MFN2.Method Sequence data for a total of 140 patients were run through VAAST and Phevor to determine genes that were more burdened with rare, nonsynonymous variants compared to control longevity cohort. Variants identified in MFN2 were expressed in Mfn2 knockout cells to determine if mutant MFN2 could rescue mitochondrial morphology defects. We identified additional rare, nonsynonymous variants in MFN2 in ALSdb that were expressed in knockout mouse embryonic fibroblasts (MEFs). Membrane potential was measured to quantify mitochondrial health upon mutant MFN2 expression. mfn2 knockout zebrafish were used to examine movement compared to wildtype and protein aggregation in brain.Results MFN2 mutations identified in ALS patients from our University of Utah cohort and ALSdb were defective in rescuing morphological defects in Mfn2 knockout MEFs. Selected mutants showed decreased membrane potential compared to wildtype MFN2 expression. Zebrafish heterozygous and homozygous for loss of mfn2 showed increased TDP-43 levels in their hindbrain and cerebellum.Conclusion In total, 21 rare, deleterious mutations in MFN2 were tested in Mfn2 knockout MEFs. Mutant MFN2 expression was not able to rescue the knockout phenotype, though at differing degrees of severity. Decreased membrane potential also argues for inhibited mitochondrial function. Increased TDP-43 levels in mutant zebrafish illustrates MFN2’s function in ALS pathology. MFN2 variants influence ALS pathology and highlight the importance of mitochondria in neurodegeneration.Competing Interest StatementK.L. Russell, J.M. Downie, K.P. Figueroa, C.J. Steely, L. Charles Murtaugh, and M.B. Bromberg report no disclosures relevant to the manuscript. S.B. Gibson reports the following competing interests: Recursion Pharmaceuticals, shareholder and Cytokinetics, advisory board. S.M. Pulst reports the following competing interests: Progenitor Life Sciences, shareholder, Cedars Sinai, royalties, University of Utah, royalties, and Ataxion Therapeutics, consultant. L.B. Jorde reports receiving royalties from Elsevier, Inc for his Medical Genetics textbook.