The p97-UBXD8 complex modulates ER-Mitochondria contact sites by modulating membrane lipid saturation and composition

Abstract

The intimate association between the endoplasmic reticulum (ER) and mitochondrial membranes at ER-mitochondria contact sites serves as a platform for several critical cellular processes, in particular lipid synthesis. Enzymes involved in lipid biosynthesis are enriched at contacts and membrane lipid composition at contacts is distinct relative to surrounding membranes. How contacts are remodeled and the subsequent biological consequences of altered contacts such as perturbed lipid metabolism remains poorly understood. Here we show that the p97 AAA-ATPase and its ER-tethered ubiquitin-X domain adaptor 8 (UBXD8) regulate the prevalence of ER-mitochondria contacts. The p97-UBXD8 complex localizes to contacts and loss of this complex increases contacts in a manner that is dependent on p97 catalytic activity. Quantitative proteomics of purified contacts demonstrates alterations in proteins regulating lipid metabolism upon loss of UBXD8. Furthermore, lipidomics studies indicate significant changes in distinct lipid species in UBXD8 knockout cells. We show that loss of p97-UBXD8 results in perturbed contacts due to an increase in membrane lipid saturation via SREBP1 and the lipid desaturase SCD1. Aberrant contacts in p97-UBXD8 loss of function cells can be rescued by supplementation with unsaturated fatty acids or overexpression of SCD1. Perturbation of contacts and inherent lipid synthesis is emerging as a hallmark to a variety of human disorders such as neurodegeneration. Notably, we find that the SREBP1-SCD1 pathway is negatively impacted in the brains of mice with p97 mutations that cause neurodegeneration. Our results suggest that contacts are exquisitely sensitive to alterations to membrane lipid composition and saturation in a manner that is dependent on p97-UBXD8.

The functional importance of VCP to maintaining cellular protein homeostasis

“…However, the mechanism by which this occurs is not known. VCP and the ERAD adaptor, UBXD8 regulate ER-mitochondria contacts by regulating membrane lipid saturation and composition ( Figure 2C ) [ 71 ]. VCP–UBXD8 regulates contacts by modulating the expression of the fatty acid desaturase SCD1 through the SREBP1 pathway.…”

“…Loss of VCP–UBXD8 stabilizes INSIG1, inactivating SREBP1, and decreasing SCD1 transcript and protein levels. This, in turn, leads to an increase in membrane saturation thereby stabilizing contacts ( Figure 2C ) [ 71 ]. This new function of VCP appears to be relevant in the context of disease.…”

“…This new function of VCP appears to be relevant in the context of disease. Motor neurons derived from ALS patients carrying the VCP Arg155His mutation have an increase in ER-mitochondria contacts and mouse models of VCP disease display a decrease in SREBP1 processing and depleted SCD1 protein levels in the brain [ 71 , 72 ]. Whether VCP can regulate other organelle contacts is not known but the loss of SCD1 protein and altered membrane saturation has been shown to impact ER-plasma membrane contacts [ 71 ].…”

UBXN1 maintains ER proteostasis and represses UPR activation by modulating translation independently of the p97 ATPase

“…The TMT-based proteomics was performed exactly as previously described 85 . Briefly, 100 µg protein of each sample was obtained by cell lysis in lysis buffer (8 M Urea, 200 mM N-(2-Hydroxyethyl)piperazine-N′-(3-propanesulfonic acid) (EPPS) pH 8.5) followed by reduction using 5 mM tris(2-carboxyethyl)phosphine (TCEP), alkylation with 14 mM iodoacetamide and quenched 33 using 5 mM dithiothreitol.…”

ALS-related p97 R155H mutation disrupts lysophagy in iPSC-derived motor neurons

“…Over 40 adaptors have been identified that enable p97 recruitment to ubiquitylated substrates 5 . We and others have shown that p97-adaptor complexes are important for a large number of cellular processes including organelle contact sites 6 , cell cycle regulation 7 , DNA damage response 8 , and autophagy 9 (see 9,10 for review). As a result, loss of p97 impacts the degradation of a large cohort of the ubiquitin-modified proteome.…”