MITOL Regulates Endoplasmic Reticulum-Mitochondria Contacts via Mitofusin2

• Published in: Molecular cell 2013-07, Vol.51 (1), p.20-34
• Main Authors: Sugiura, Ayumu, Nagashima, Shun, Tokuyama, Takeshi, Amo, Taku, Matsuki, Yohei, Ishido, Satoshi, Kudo, Yoshihisa, McBride, Heidi M., Fukuda, Toshifumi, Matsushita, Nobuko, Inatome, Ryoko, Yanagi, Shigeru
• Format: Article
• Language: English
• Subjects: Animals; centrifugation; endoplasmic reticulum; Endoplasmic Reticulum - metabolism; GTP Phosphohydrolases - analysis; GTP Phosphohydrolases - metabolism; GTP Phosphohydrolases - physiology; guanosine triphosphate; guanosinetriphosphatase; HeLa Cells; Humans; hydrolysis; Membrane Proteins; Mice; mitochondria; Mitochondria - metabolism; Mitochondrial Proteins - analysis; Mitochondrial Proteins - metabolism; Mitochondrial Proteins - physiology; mutation; polyacrylamide gel electrophoresis; ubiquitin-protein ligase; Ubiquitin-Protein Ligases - metabolism; Ubiquitin-Protein Ligases - physiology; Ubiquitination
• ISSN: 1097-2765; 1097-4164
• DOI: 10.1016/j.molcel.2013.04.023

The mitochondrial ubiquitin ligase MITOL regulates mitochondrial dynamics. We report here that MITOL regulates mitochondria-associated endoplasmic reticulum (ER) membrane (MAM) domain formation through mitofusin2 (Mfn2). MITOL interacts with and ubiquitinates mitochondrial Mfn2, but not ER-associated Mfn2. Mutation analysis identified a specific interaction between MITOL C-terminal domain and Mfn2 HR1 domain. MITOL mediated lysine-63-linked polyubiquitin chain addition to Mfn2, but not its proteasomal degradation. MITOL knockdown inhibited Mfn2 complex formation and caused Mfn2 mislocalization and MAM dysfunction. Sucrose-density gradient centrifugation and blue native PAGE retardation assay demonstrated that MITOL is required for GTP-dependent Mfn2 oligomerization. MITOL knockdown reduced Mfn2 GTP binding, resulting in reduced GTP hydrolysis. We identified K192 in the GTPase domain of Mfn2 as a major ubiquitination site for MITOL. A K192R mutation blocked oligomerization even in the presence of GTP. Taken together, these results suggested that MITOL regulates ER tethering to mitochondria by activating Mfn2 via K192 ubiquitination. •MITOL directly interacts with and ubiquitinates mitochondrial Mfn2•MITOL regulates the GTP-binding activity and oligomerization of Mfn2•MITOL regulates the interaction between ER and mitochondria via Mfn2