Mitofusin 1 overexpression rescues the abnormal mitochondrial dynamics caused by the Mitofusin 2 K357T mutation in vitro.

BACKGROUND AND AIMS: Mitofusin 1 (MFN1) and MFN2 are outer mitochondrial membrane fusogenic proteins regulating mitochondrial network morphology. MFN2 mutations cause Charcot-Marie-Tooth type 2A (CMT2A), an axonal neuropathy characterized by mitochondrial fusion defects, which in the case of a GTPase domain mutant, were rescued following wild type MFN1/2 (MFN1/2WT ) overexpression. In this study, we compared the therapeutic efficiency between MFN1WT and MFN2WT overexpression in correcting mitochondrial defects induced by the novel MFN2K357T mutation located in the highly conserved R3 region.

METHODS: Constructs expressing either MFN2K357T , MFN2WT , or MFN1WT under the ubiquitous chicken β-actin hybrid (CBh) promoter were generated. Flag or myc tag was used for their detection. Differentiated SH-SY5Y cells were single transfected with MFN1WT , MFN2WT , or MFN2K357T , as well as double transfected with MFN2K357T /MFN2WT or MFN2K357T /MFN1WT .

RESULTS: SH-SY5Y cells transfected with MFN2K357T exhibited severe perinuclear mitochondrial clustering with axon-like processes devoid of mitochondria. Single transfection with MFN1WT resulted in a more interconnected mitochondrial network than transfection with MFN2WT , accompanied by mitochondrial clusters. Double transfection of MFN2K357T with either MFN1WT or MFN2WT resolved the mutant-induced mitochondrial clusters and led to detectable mitochondria throughout the axon-like processes. MFN1WT showed higher efficacy than MFN2WT in rescuing these defects.

INTERPRETATION: These results further demonstrate the higher potential of MFN1WT over MFN2WT overexpression to rescue CMT2A-induced mitochondrial network abnormalities due to mutations outside the GTPase domain. This higher phenotypic rescue conferred by MFN1WT , possibly due to its higher mitochondrial fusogenic ability, may be applied to different CMT2A cases regardless of the MFN2 mutation type. This article is protected by copyright. All rights reserved.