Mutation of the ATL1 gene is one of the most common causes of hereditary spastic paraplegia (HSP), a group of genetic neurodegenerative conditions characterised by distal axonal degeneration of the corticospinal tract axons. Atlastin-1, the protein encoded by ATL1, is one of three mammalian atlastins, which are homologous dynamin-like GTPases that control endoplasmic reticulum (ER) morphology by fusing tubules to form the three-way junctions that characterise ER networks. However, it is not clear whether atlastin-1 is required for correct ER morphology in human neurons and if so what the functional consequences of lack of atlastin-1 are. Using CRISPR-inhibition we generated human cortical neurons lacking atlastin-1. We demonstrate that ER morphology was altered in these neurons, with a reduced number of three-way junctions. Neurons lacking atlastin-1 had longer endosomal tubules, suggestive of defective tubule fission. This was accompanied by reduced lysosomal proteolytic capacity. As well as demonstrating that atlastin-1 is required for correct ER morphology in human neurons, our results indicate that lack of a classical ER-shaping protein such as atlastin-1 may cause altered endosomal tubulation and lysosomal proteolytic dysfunction. Furthermore, they strengthen the idea that defective lysosome function contributes to the pathogenesis of a broad group of HSPs, including those where the primary localisation of the protein involved is not at the endolysosomal system.This research was supported by the NIHR Cambridge Biomedical Research Centre [Grant numbers BRC-1215-20014 and NIHR203312], Medical Research Council Project Grants [Grant numbers MR/R026440/1 and MR/V028677/1] and by a grant from the Tom Wahlig Stiftung. EZ was supported by Medical Research Council Ph.D. studentship [Grant number MR/K50127X/1] and by a Gates Cambridge Trust Scholarship. JK was supported by Medical Research Council Ph.D. studentship [Grant number MR/N013433/1] and the E.G. Fearnsides Trust Fund. ZK is supported by a Wellcome Trust Fellowship [Grant number220597/Z/20/Z]. We are grateful to Hazel and Keith Satchell for their kind charitable support for our work on HSP
