Doa1 targets ubiquitinated substrates for mitochondria-associated degradation

Abstract

Mitochondria-associated degradation (MAD) mediated by the Cdc48 complex and proteasome degrades ubiquitinated mitochondrial outer-membrane proteins. MAD is critical for mitochondrial proteostasis, but it remains poorly characterized. We identified several mitochondrial Cdc48 substrates and developed a genetic screen assay to uncover regulators of the Cdc48-dependent MAD pathway. Surprisingly, we identified Doa1, a substrate-processing factor of Cdc48 that inhibits the degradation of some Cdc48 substrates, as a critical mediator of the turnover of mitochondrial Cdc48 substrates. Deletion of DOA1 causes the accumulation and mislocalization of substrates on mitochondria. Profiling of Cdc48 cofactors shows that Doa1 and Cdc48-Ufd1-Npl4 form a functional complex mediating MAD. Biochemically, Doa1 interacts with ubiquitinated substrates and facilitates substrate recruitment to the Cdc48-Ufd1-Npl4 complex. Functionally, Doa1 is critical for cell survival under mitochondrial oxidative stress, but not ER stress, conditions. Collectively, our results demonstrate the essential role of the Doa1–Cdc48-Ufd1-Npl4 complex in mitochondrial proteostasis and suggest that Doa1 plays dual roles on the Cdc48 complex.

Ubiquitination of ERMES components by the E3 ligase Rsp5 is involved in mitophagy

“…While our paper was under revision, another group published results confirming our observations concerning the ubiquitination of Mdm34 by Rsp5 and the role of the PY motif in this post-translational modification 55 . However, in their experimental conditions, Wu et al.…”

“…However, in their experimental conditions, Wu et al. observe very high levels of Mdm34 polyubiquitination (polyubiquitinated chains >170 kDa), which are implicated in the rapid turnover of the protein 55 . Their results do not conflict with those presented here but they indicate that the length of ubiquitin chains may change the fate of Mdm34.…”

VCP /p97 cooperates with YOD 1, UBXD 1 and PLAA to drive clearance of ruptured lysosomes by autophagy

“…A similar model has also been proposed for the role of p97 in mitophagy, where p97‐mediated extraction of factors on the outer membrane, such as mitofusins, is thought to facilitate the clearance of damaged mitochondria or mitophagy (Tanaka et al , ; Chan et al , ). Of note, extraction of mitofusins involves the Ufd1‐Npl4 cofactor (Kim et al , ; Wu et al , ), whereas we identified a distinct set of cofactors in lysophagy including the DUB, YOD1. DUBs such as YOD1 were also shown to contribute to extraction of proteins from the ER membrane during ER‐associated degradation (Ernst et al , ; Liu & Ye, ; Stein et al , ).…”

“…To explore the molecular basis for p97 activity in lysophagy, we aimed to identify the relevant cofactors that cooperate with p97 in the process. We speculated that the lysosomal damage response involves the p97 cofactors UBXD1 and PLAA, because they had previously been linked to the endolysosomal system or autophagy in different systems and are affected by p97 disease mutations (Ren et al , ; Ossareh‐Nazari et al , ; Ritz et al , ; Wu et al , ). Moreover, we considered the DUB, YOD1, whose homologue OTU1 physically and genetically interacts with p97 and PLAA/Ufd3 in yeast (Rumpf & Jentsch, ).…”

Mitochondrial AAA‐ATPase Msp1 detects mislocalized tail‐anchored proteins through a dual‐recognition mechanism

“…Data information: In this figure, GFP-Pex15D30 was expressed from a centromeric plasmid under the control of TEF1 promoter, and WT and mutant forms of Msp1-FLAG were expressed from the endogenous chromosomal locus. Msp1 E193Q -FLAG ( Fig 1D, lanes [5][6][7][8][9][10][11][12][13][14][15]. Therefore, most of the residues are required for substrate binding.…”