MITOL regulates endoplasmic reticulum-mitochondria contacts via Mitofusin2

Mol Cell. 2013 Jul 11;51(1):20-34. doi: 10.1016/j.molcel.2013.04.023. Epub 2013 May 30.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Endoplasmic Reticulum / metabolism*
  • GTP Phosphohydrolases / analysis
  • GTP Phosphohydrolases / metabolism*
  • GTP Phosphohydrolases / physiology*
  • HeLa Cells
  • Humans
  • Membrane Proteins
  • Mice
  • Mitochondria / metabolism*
  • Mitochondrial Proteins / analysis
  • Mitochondrial Proteins / metabolism*
  • Mitochondrial Proteins / physiology*
  • Ubiquitin-Protein Ligases / metabolism
  • Ubiquitin-Protein Ligases / physiology*
  • Ubiquitination

Substances

  • Membrane Proteins
  • Mitochondrial Proteins
  • MARCH5 protein, human
  • Ubiquitin-Protein Ligases
  • GTP Phosphohydrolases
  • MFN2 protein, human