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

Dan A Tudorica; Bishal Basak; Alexia S Puerta Cordova; Grace Khuu; Kevin Rose; Michael Lazarou; Erika L F Holzbaur; James H Hurley

doi:10.1083/jcb.202309015

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.

Authors

Dan A Tudorica^{1

2

3}, Bishal Basak^{1

4}, Alexia S Puerta Cordova^{3

5}, Grace Khuu^{1

6}, Kevin Rose^{3

5}, Michael Lazarou^{1

6}, Erika L F Holzbaur^{1

4}, James H Hurley^{1

2

3

7}

Affiliations

¹ Aligning Science Across Parkinson's (ASAP) Collaborative Research Network , Chevy Chase, MD, USA.
² Graduate Group in Biophysics, University of California, Berkeley, Berkeley, CA, USA.
³ California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA, USA.
⁴ Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA.
⁵ Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, USA.
⁶ Walter and Eliza Hall Institute of Medical Research , Melbourne, Australia.
⁷ Helen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, CA, USA.

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

Grants and funding

ASAP-000350/Aligning Science Across Parkinson's