Costello et al. identify ACBD5 and VAPB as key components of a peroxisome–ER tether in mammalian cells. Disruption of this tethering complex leads to reduced peroxisomal membrane expansion and increased peroxisomal movement.

Tail-anchored (TA) proteins contain a single transmembrane domain (TMD) at the C-terminus that anchors them to the membranes of organelles where they mediate critical cellular processes. Accordingly, mutations in genes encoding TA proteins have been identified in a number of severe inherited disorders. Despite the importance of correctly targeting a TA protein to its appropriate membrane, the mechanisms and signals involved are not fully understood. In this study, we identify additional peroxisomal TA proteins, discover more proteins that are present on multiple organelles, and reveal that a combination of TMD hydrophobicity and tail charge determines targeting to distinct organelle locations in mammals. Specifically, an increase in tail charge can override a hydrophobic TMD signal and re-direct a protein from the ER to peroxisomes or mitochondria and vice versa. We show that subtle changes in those parameters can shift TA proteins between organelles, explaining why peroxisomes and mitochondria have many of the same TA proteins. This enabled us to associate characteristic physicochemical parameters in TA proteins with particular organelle groups. Using this classification allowed successful prediction of the location of uncharacterized TA proteins for the first time.

Peroxisomes are dynamic organelles which fulfil essential roles in lipid and ROS metabolism. Peroxisome movement and positioning allows interaction with other organelles and is crucial for their cellular function. In mammalian cells, such movement is microtubule‐dependent and mediated by kinesin and dynein motors. The mechanisms of motor recruitment to peroxisomes are largely unknown, as well as the role this plays in peroxisome membrane dynamics and proliferation. Here, using a combination of microscopy, live‐cell imaging analysis and mathematical modelling, we identify a role for Mitochondrial Rho GTPase 1 (MIRO1) as an adaptor for microtubule‐dependent peroxisome motility in mammalian cells. We show that MIRO1 is targeted to peroxisomes and alters their distribution and motility. Using a peroxisome‐targeted MIRO1 fusion protein, we demonstrate that MIRO1‐mediated pulling forces contribute to peroxisome membrane elongation and proliferation in cellular models of peroxisome disease. Our findings reveal a molecular mechanism for establishing peroxisome‐motor protein associations in mammalian cells and provide new insights into peroxisome membrane dynamics in health and disease.

Assigning functions to the vast array of proteins present in eukaryotic cells remains challenging. To identify relationships between proteins, and thereby enable functional annotations of proteins, we determined changes of abundance of 10,323 human proteins in response to 294 biological perturbations using isotope-labelling mass spectrometry. We applied the machine learning algorithm treeClust to reveal functional associations between co-regulated human proteins from ProteomeHD, a compilation of our own data and datasets from the Proteomics Identifications (PRIDE) database. This produced a co-regulation map of the human proteome. Co-regulation was able to capture relationships between proteins that do not physically interact or co-localize. For example, co-regulation of the peroxisomal membrane protein PEX11β with mitochondrial respiration factors led us to discover an organelle interface between peroxisomes and mitochondria ✉

Cooperation between cellular organelles such as mitochondria, peroxisomes and the ER is essential for a variety of important and diverse metabolic processes. Effective communication and metabolite exchange requires physical linkages between the organelles, predominantly in the form of organelle contact sites. At such contact sites organelle membranes are brought into close proximity by the action of molecular tethers, which often consist of specific protein pairs anchored in the membrane of the opposing organelles. Currently numerous tethering components have been identified which link the ER with multiple other organelles but knowledge of the factors linking the ER with peroxisomes is limited. Peroxisome-ER interplay is important because it is required for the biosynthesis of unsaturated fatty acids, ether-phospholipids and sterols with defects in these functions leading to severe diseases. Here, we characterize acyl-CoA binding domain protein 4 (ACBD4) as a tail-anchored peroxisomal membrane protein which interacts with the ER protein, vesicle-associated membrane protein-associated protein–B (VAPB) to promote peroxisome-ER associations.