Restoring mitofusin balance prevents axonal degeneration in a Charcot-Marie-Tooth type 2A model
Yueqin Zhou, Sharon Carmona, A.K.M.G. Muhammad, Shaughn Bell, Jesse Landeros, Michael Vazquez, Ritchie Ho, Antonietta Franco, Bin Lu, Gerald W. Dorn II, Shaomei Wang, Cathleen M. Lutz, Robert H. Baloh
Yueqin Zhou, Sharon Carmona, A.K.M.G. Muhammad, Shaughn Bell, Jesse Landeros, Michael Vazquez, Ritchie Ho, Antonietta Franco, Bin Lu, Gerald W. Dorn II, Shaomei Wang, Cathleen M. Lutz, Robert H. Baloh
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Research Article Neuroscience

Restoring mitofusin balance prevents axonal degeneration in a Charcot-Marie-Tooth type 2A model

Abstract

Mitofusin-2 (MFN2) is a mitochondrial outer-membrane protein that plays a pivotal role in mitochondrial dynamics in most tissues, yet mutations in MFN2, which cause Charcot-Marie-Tooth disease type 2A (CMT2A), primarily affect the nervous system. We generated a transgenic mouse model of CMT2A that developed severe early onset vision loss and neurological deficits, axonal degeneration without cell body loss, and cytoplasmic and axonal accumulations of fragmented mitochondria. While mitochondrial aggregates were labeled for mitophagy, mutant MFN2 did not inhibit Parkin-mediated degradation, but instead had a dominant negative effect on mitochondrial fusion only when MFN1 was at low levels, as occurs in neurons. Finally, using a transgenic approach, we found that augmenting the level of MFN1 in the nervous system in vivo rescued all phenotypes in mutant MFN2R94Q-expressing mice. These data demonstrate that the MFN1/MFN2 ratio is a key determinant of tissue specificity in CMT2A and indicate that augmentation of MFN1 in the nervous system is a viable therapeutic strategy for the disease.

Authors

Yueqin Zhou, Sharon Carmona, A.K.M.G. Muhammad, Shaughn Bell, Jesse Landeros, Michael Vazquez, Ritchie Ho, Antonietta Franco, Bin Lu, Gerald W. Dorn II, Shaomei Wang, Cathleen M. Lutz, Robert H. Baloh

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Figure 2

Widespread axon degeneration and gliosis without neuronal cell loss in MFN2R94Q mice.

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Widespread axon degeneration and gliosis without neuronal cell loss in M...
(A) Degenerating axons labeled with Fluoro-Jade staining in the brain pyramidal tracts of the medulla and the spinal cord of MFN2R94Q mice (5 months old). Scale bar: 50 μM. Graph represents the quantification of Fluoro-Jade–positive signal as pixel2 per mm2 ROI subtracted from background. Data are represented as mean ± SEM. n = 3 mice/genotype. Student’s t test, 2-tailed for pyramidal track and 1-tailed for spinal cord. *P < 0.05. (B) Toluidine blue–stained plastic sections of spinal cord and tibial nerves from 14-month-old nTg or MFN2WT and MFN2R94Q mice. Scale bars: 10 μM (spinal cord); 50 μM (tibial nerve). Arrows indicate degenerating axons in the spinal cord. Tibial nerve showed axonal atrophy without axonal loss. n = 3 mice/genotype. (C) Size frequency histogram of tibial nerve axons (n = 3 mice/group) showing axonal atrophy in MFN2R94Q mice (Kolmogorov-Smirnov test, P < 0.0001). (D) RGC axons stained with neurofilament antibody (RT97) in retinal whole-mount preparations. Scale bars: 50 μM. Arrowheads indicate beaded axons, and arrows indicate atrophic RGCs, seen only in MFN2R94Q mice. n = 2 mice/genotype. (E) Neurofilament in cross sections of optic nerve stained with SMI32 antibody. Scale bar: 50 μM. Axonal spheroids (white arrowheads) were present in the optic nerves of MFN2R94Q mouse (3 months old). Graph represents the quantification of axonal spheroids. Data are represented as mean ± SEM. n = 4 /genotype. Student’s t test, 2-tailed. ****P < 0.0001. (F) Astrocyte activation was present in regions of MFN2R94Q mouse brains (brain stem shown here) in areas of projection neurons that contained mitochondrial aggregates (5-month-old mice). GFAP (green); Flag (red); DAPI (blue). Scale bar: 50 μM. n = 3 mice/genotype. (G) Microgliosis in regions of MFN2R94Q mouse brains (cortex shown here) (5-month-old mice). Iba1 (green); Flag (red); DAPI (blue). Scale bar: 100 μM. n = 3 mice/genotype.