Phosphorylation of mitochondrial polyubiquitin by PINK1 promotes Parkin mitochondrial tethering

PLoS Genet. 2014 Dec 4;10(12):e1004861. doi: 10.1371/journal.pgen.1004861. eCollection 2014 Dec.

Abstract

The kinase PINK1 and the E3 ubiquitin (Ub) ligase Parkin participate in mitochondrial quality control. The phosphorylation of Ser65 in Parkin's ubiquitin-like (UBl) domain by PINK1 stimulates Parkin activation and translocation to damaged mitochondria, which induces mitophagy generating polyUb chain. However, Parkin Ser65 phosphorylation is insufficient for Parkin mitochondrial translocation. Here we report that Ser65 in polyUb chain is also phosphorylated by PINK1, and that phosphorylated polyUb chain on mitochondria tethers Parkin at mitochondria. The expression of Tom70MTS-4xUb SE, which mimics phospho-Ser65 polyUb chains on the mitochondria, activated Parkin E3 activity and its mitochondrial translocation. An E3-dead form of Parkin translocated to mitochondria with reduced membrane potential in the presence of Tom70(MTS)-4xUb SE, whereas non-phospho-polyUb mutant Tom70(MTS)-4xUb SA abrogated Parkin translocation. Parkin binds to the phospho-polyUb chain through its RING1-In-Between-RING (IBR) domains, but its RING0-linker is also required for mitochondrial translocation. Moreover, the expression of Tom70(MTS)-4xUb SE improved mitochondrial degeneration in PINK1-deficient, but not Parkin-deficient, Drosophila. Our study suggests that the phosphorylation of mitochondrial polyUb by PINK1 is implicated in both Parkin activation and mitochondrial translocation, predicting a chain reaction mechanism of mitochondrial phospho-polyUb production by which rapid translocation of Parkin is achieved.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Animals, Genetically Modified
  • Cells, Cultured
  • Drosophila
  • HEK293 Cells
  • HeLa Cells
  • Humans
  • Mice
  • Mitochondria / metabolism*
  • Phosphorylation
  • Polyubiquitin / metabolism*
  • Protein Binding
  • Protein Kinases / metabolism*
  • Protein Transport
  • Ubiquitin-Protein Ligases / metabolism*
  • Ubiquitination

Substances

  • Polyubiquitin
  • Ubiquitin-Protein Ligases
  • parkin protein
  • Protein Kinases
  • PTEN-induced putative kinase

Grants and funding

This study was supported by the Takeda Science Foundation (YIm), the Life Science Foundation of Japan (YIm), the Daiichi-Sankyo Foundation for Life Science (YIm), the Mochida Memorial Foundation for Medical and Pharmaceutical Research (YIm), and Otsuka Pharmaceutical (NH, YIm), the Grant-in-Aid for Scientific Research 26461280 (KSF) and 26293070 (YIm) from MEXT in Japan, and a Grant-in-Aid for Scientific Research on Innovative Areas 23111003 (NH) and 26117727 (YIm). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.