Disease-causing MFN2 mutants impair mitochondrial fission dynamics by distinct DRP1 dysregulation

Mitochondria undergo fusion and fission. While DRP1 regulates fission, fusion is controlled by OPA1, MFN1, and MFN2. The balance between these processes and the crosstalk between machineries remains poorly understood. MFN2 mutations cause Charcot-Marie-Tooth disease type 2A (CMT2A), affecting mitochondrial fusion and morphology. However, their role in fission is unclear. Using skin fibroblasts from CMT2A patients (L248H and M376V MFN2 mutations) and wild-type mouse embryonic fibroblasts expressing these variants we studied how MFN2 mutations impact mitochondrial dynamics beyond fusion. We analysed mitochondrial morphology and dynamics, by live-cell confocal microscopy, and tested fusion/fission protein levels, oxygen consumption rate (OCR), extracellular acidification rate (ECAR), and oxidative phosphorylation complex subunits. MFN2 mutations impaired mitochondrial fusion and displayed distinct effects on fission and cellular metabolism. L248H-expressing cells showed hyper-elongated mitochondria, impaired fission, and increased OCR, while, M376V cells exhibited fragmentation, enhanced fission, and elevated ECAR. These effects correlated with differential Drp1 phosphorylation. Our findings demonstrate that MFN2 mutants differentially influences fission and metabolism, highlighting the need to consider these effects in therapies aimed at modulating mitochondrial dynamics.