A RAB7A phosphoswitch coordinates Rubicon Homology protein regulation of Parkin-dependent mitophagy

J Cell Biol. 2024 Jul 1;223(7):e202309015. doi: 10.1083/jcb.202309015. Epub 2024 May 10.

Abstract

Activation of PINK1 and Parkin in response to mitochondrial damage initiates a response that includes phosphorylation of RAB7A at Ser72. Rubicon is a RAB7A binding negative regulator of autophagy. The structure of the Rubicon:RAB7A complex suggests that phosphorylation of RAB7A at Ser72 would block Rubicon binding. Indeed, in vitro phosphorylation of RAB7A by TBK1 abrogates Rubicon:RAB7A binding. Pacer, a positive regulator of autophagy, has an RH domain with a basic triad predicted to bind an introduced phosphate. Consistent with this, Pacer-RH binds to phosho-RAB7A but not to unphosphorylated RAB7A. In cells, mitochondrial depolarization reduces Rubicon:RAB7A colocalization whilst recruiting Pacer to phospho-RAB7A-positive puncta. Pacer knockout reduces Parkin mitophagy with little effect on bulk autophagy or Parkin-independent mitophagy. Rescue of Parkin-dependent mitophagy requires the intact pRAB7A phosphate-binding basic triad of Pacer. Together these structural and functional data support a model in which the TBK1-dependent phosphorylation of RAB7A serves as a switch, promoting mitophagy by relieving Rubicon inhibition and favoring Pacer activation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autophagy-Related Proteins* / genetics
  • Autophagy-Related Proteins* / metabolism
  • HEK293 Cells
  • HeLa Cells
  • Humans
  • Intracellular Signaling Peptides and Proteins / genetics
  • Intracellular Signaling Peptides and Proteins / metabolism
  • Mitochondria / genetics
  • Mitochondria / metabolism
  • Mitophagy*
  • Phosphorylation
  • Protein Binding
  • Protein Serine-Threonine Kinases* / genetics
  • Protein Serine-Threonine Kinases* / metabolism
  • Ubiquitin-Protein Ligases* / genetics
  • Ubiquitin-Protein Ligases* / metabolism
  • rab7 GTP-Binding Proteins*

Substances

  • Autophagy-Related Proteins
  • Intracellular Signaling Peptides and Proteins
  • parkin protein
  • Protein Serine-Threonine Kinases
  • rab7 GTP-Binding Proteins
  • rab7 GTP-binding proteins, human
  • RUBCN protein, human
  • TBK1 protein, human
  • Ubiquitin-Protein Ligases