UBXD1 and YOD1: p97 cofactors involved in autophagic mitochondrial quality control

Diminished mitochondrial function impacts on cellular metabolism but also critically influences life and also health span. Mitochondrial dysfunction due to accumulating
mitochondrial damage is considered one of the main factors of aging and aging-related disease such as Alzheimer's disease and other neurodegenerative disorders. Mitochondrial
quality control is essential to prevent dysfunction and associated deleterious outcomes. Multi-tiered molecular machinery is in place to remove and degrade superfuluous or damaged proteins to maintain mitochondrial proteostasis, cull mitochondrial subunits beyond repair or remove entire mitochondrial networks through apoptosis. Besides proteolytic pathways, autophagic removal is an important part of mitochondrial quality control.
Severe damage to mitochondria exceeding the repair capacity of proteolytic systems,but below the apoptotic threshold, leads to the removal of mitochondrial units through
mitochondria-specific autophagy or mitophagy under control of the kinase PINK1 and the ubiquitin ligase Parkin. Following recognition of damaged mitochondrial subunits by
PINK1, Parkin is recruited causing the ubiquitination of mitochondrial proteins. This results in the recruitment of autophagy cargo adaptors leading to the engulfment of
the damaged mitochondria and its subsequent degradation in the lysosome. Among the proteins recruited during mitophagy is the AAA-ATPase VCP/p97. As ubiquitously expressed protein, p97 acts in a plethora of cellular functions involving ubiquitination,including cell cycle control, transcriptional regulation as well as proteostasis. In addition,p97 was recently connected to ubiquitin-mediated degradation of mitochondrial proteins,Parkin-dependent mitophagy and deubiquitinating enzymes. These multiple diverse functions of p97 suggest tight spatial and temporal control of its activity, which is brought upon by the interaction with various cofactors promoting substrate recognition and processing by p97.
In this in vitro study the connection of p97 to mitochondrial quality control with focus on mitophagy was studied. In a first step, p97 cofactors were screened using subcellular localization analysis for their ability to translocate to mitochondria under mitophagic conditions. From this screen, UBXD1, SAKS1, and Erasin were found to alter their locations following mitophagic induction implicating these proteins in the autophagic clearance of mitochondria. Further analysis suggested a role for UBXD1 as recruitment factor for p97 to damaged mitochondria. It was shown that UBXD1 recognizes depolarized mitochondria via its C-terminal UBX domain and translocates to mitochondria in
a Parkin-dependent manner. Once translocated, UBXD1 recruits p97 to mitochondria via a bipartite binding motif consisting of its N-terminal VIM and PUB domains. Recruitment
of p97 by UBXD1 only depends on the presence of UBXD1 on mitochondria without the need for further mitochondrial signals. Following translocation of UBXD1 to CCCP-depolarized mitochondria and p97 recruitment, formation of autolysosomes is strongly enhanced and autophagic degradation of mitochondria is significantly accelerated.
Diminished levels of UBXD1 result in decreased mitophagic flux.
In a next step, a potential role in mitophagy for the deubiquitinating enzyme YOD1 was studied. YOD1 was previously reported to facilitate together with p97 and UBXD1 lyosphagy and ERAD. Alternative transcript analysis revealed a diferential role for YOD1. Under mitophagic conditions, the shorter YOD1.2 was translocating to mitochondrial while the longer YOD1.1 remained in the cytosol. In addition, ecotopic expression of UBXD1 greatly enhanced mitochondrial translocation of YOD1.2. Under these
conditions, UBXD1 and YOD1.2 translocate to depolarized mitochondria in a mutually exclusive manner with YOD1.2 seemingly to displace UBXD1. In summary, the work presented here suggests a novel role for UBXD1 as sensor for mitochondria undergoing mitophagy and mitochondrial recruitment factor for p97 during mitophagy. Furthermore, UBXD1 potentially initiates a multi-step cascade involving
YOD1.2 ultimately aiding the mitophagic quality control of damaged mitochondria.