Abstract
Mitofusins are members of the dynamin-related protein family of large GTPases that harness the energy from nucleotide hydrolysis to remodel membranes. Mitofusins possess four structural domains, including a GTPase domain, two extended helical bundles (HB1 and HB2), and a transmembrane region. We have characterized four Charcot-Marie-Tooth type 2A-associated variants with amino acid substitutions in Mfn2 that are proximal to the hinge that connects HB1 and HB2. A functional defect was not apparent in cells as the mitochondrial morphology of Mfn2-null cells was restored by expression of any of these variants. However, a significant fusion deficiency was observed in vitro, which was improved by the addition of crude cytosol extract or soluble Bax. All four variants had reduced nucleotide-dependent assembly in cis, but not trans, and this was also improved by the addition of Bax. Together, our data demonstrate an important role for this region in Mfn2 GTP-dependent oligomerization and membrane fusion and is consistent with a model where cytosolic factors such as Bax are masking molecular defects associated with Mfn2 disease variants in cells.
Highlights
Mitochondrial dynamics have become increasingly recognized as an important indicator of and contributor to both cellular health and death
Mitochondria are trafficked on microtubules and the overall shape and connectivity of the mitochondrial network is maintained or modified through mitochondrial fusion and division, which are mediated by membrane remodeling large GTPase proteins of the dynamin related protein (DRP) family (Labbeet al, 2014)
In a structural model of Mfn2 based on the closed structure of bacterial dynamin like protein (BDLP) (PDB 2J69), S378 resides in HB1, C390 is within HB2, and A383 and Q386 are within a loop connecting HB1 and HB2 (Fig 1A)
Summary
Mitochondrial dynamics have become increasingly recognized as an important indicator of and contributor to both cellular health and death. When mitochondrial fusion occurs more frequently than division, the result is a more connected network comprising longer mitochondria, which is associated with increased ATP production, such as during a cellular stress response. The importance of these processes and their regulation is highlighted by the association of dysregulated mitochondrial dynamics with various diseases such as Parkinson’s disease, diabetes, and peripheral neuropathies (Züchner et al, 2004; Vital & Vital, 2012; Celardo et al, 2014; Wada & Nakatsuka, 2016; Rovira-Llopis et al, 2017)
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