CMT2A alleles with different effects on mitochondrial fusion share altered mitochondrial distribution and mitofusin turnover

Charcot-Marie-Tooth type 2A neuropathy (CMT2A) is caused by nearly one hundred missense mutations in the mitochondrial fusion factor mitofusin (Mfn). We showed that mutation hotspots target different structural motifs with functional significance for Mfn2. We then asked whether these mutations could differentially affect Mfn activity by comparing fourteen CMT2A alleles in drosophila. Based on specific changes in mitochondrial architecture, we grouped these alleles into four phenotypic classes. In particular, we identified dominant-negative alleles of variable strength, generally associated with catalytic motifs and hydrophobic cores, and active alleles favouring mitochondrial fusion, mostly located at the interface between domains. The cytoplasmic distribution of mitochondria was disturbed to varying degrees by the CMT2A alleles, associated with reduced endoplasmic reticulum-mitochondria overlap and fewer mitochondria at neuromuscular junctions. The heterogeneity of mitochondrial repartition was a major driver of neurotoxicity, correlating with the level of locomotor impairment measured in mutant flies and the disease severity in patients. At the molecular scale, most mutants showed reduced ubiquitination and increased basal levels of Mfn, likely contributing to their dominant properties. In conclusion, the CMT2A alleles induce an unexpected diversity of Mfn alterations in flies with general consequences on mitochondrion distribution and mutant protein level.