Hereditary spastic paraplegia (HSP) is a genetically heterogeneous disease caused by mutations in many genes, including those encoding spastin, strumpellin, or REEP1. Allison et al. show that similar lysosomal phenotypes are associated with mutations in different classes of HSP proteins and suggest that defective ER–endosome contacts and endosome tubule fission may be a common cause of axon degeneration in the disease.

Exosomes are extracellular vesicles first described as such 30 years ago and since implicated in cell–cell communication and the transmission of disease states, and explored as a means of drug discovery. Yet fundamental questions about their biology remain unanswered. Here I explore what exosomes are, highlight the difficulties in studying them and explain the current definition and some of the outstanding issues in exosome biology.

Multivesicular endosomes/bodies (MVBs) contain intraluminal vesicles (ILVs) that bud away from the cytoplasm. Multiple mechanisms of ILV formation have been identified, but the relationship between different populations of ILVs and MVBs remains unclear. Here, we show in HeLa cells that different ILV subpopulations can be distinguished by size. EGF stimulation promotes the formation of large ESCRT-dependent ILVs, whereas depletion of the ESCRT-0 component, Hrs, promotes the formation of a uniformly sized population of small ILVs, the formation of which requires CD63. CD63 has previously been implicated in ESCRT-independent sorting of PMEL in MVBs and transfected PMEL is present on the small ILVs that form on Hrs depletion. Upregulation of CD63-dependent ILV formation by Hrs depletion indicates that Hrs and CD63 regulate competing machineries required for the generation of distinct ILV subpopulations. Taken together our results indicate that ILV size is influenced by their cargo and mechanism of formation and suggest a competitive relationship between ESCRT-dependent and -independent mechanisms of ILV formation within single MVBs.

The adaptor protein complex AP-5 is stably associated with the hereditary spastic paraplegia proteins SPG11 and SPG15 in an apparently equimolar ratio to form a new type of coat. Results suggest that SPG15 facilitates the docking of the coat onto late endosomal/lysosomal membranes and that SPG11 (possibly together with SPG15) acts as a scaffold.

Adaptor protein 4 (AP-4) is an ancient membrane trafficking complex, whose function has largely remained elusive. In humans, AP-4 deficiency causes a severe neurological disorder of unknown aetiology. We apply unbiased proteomic methods, including ‘Dynamic Organellar Maps’, to find proteins whose subcellular localisation depends on AP-4. We identify three transmembrane cargo proteins, ATG9A, SERINC1 and SERINC3, and two AP-4 accessory proteins, RUSC1 and RUSC2. We demonstrate that AP-4 deficiency causes missorting of ATG9A in diverse cell types, including patient-derived cells, as well as dysregulation of autophagy. RUSC2 facilitates the transport of AP-4-derived, ATG9A-positive vesicles from the trans-Golgi network to the cell periphery. These vesicles cluster in close association with autophagosomes, suggesting they are the “ATG9A reservoir” required for autophagosome biogenesis. Our study uncovers ATG9A trafficking as a ubiquitous function of the AP-4 pathway. Furthermore, it provides a potential molecular pathomechanism of AP-4 deficiency, through dysregulated spatial control of autophagy.