The Role of Ultrastructural Examination in Storage Diseases

Papa, Valentina; Tarantino, Lucia; Preda, Paola; Giorgi, L. Badiali De; Fanin, Marina; Pegoraro, Elena; Angelini, C.; Cenacchi, Giovanna

doi:10.3109/01913121003780593

Cited by 7 publications

(5 citation statements)

References 23 publications

(21 reference statements)

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“…Electron microscopy (EM) showed a high variation in appearance of endolysosomal compartments (Fig EV2A -D), but no inclusion bodies. This indicates that mutations in VPS41 do not cause a classical lysosomal storage phenotype (Papa et al, 2010) , which is in agreement with the metabolic studies performed on patient fibroblasts, leucocytes, and urine (see patient descriptions). To study lysosomal acidification and hydrolase activity, we performed fluorescence microscopy using MagicRed cathepsin B.…”

Section: Resultssupporting

confidence: 88%

Neurodegenerative VPS41 variants inhibit HOPS function and mTORC1‐dependent TFEB/TFE3 regulation

et al. 2021

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Vacuolar protein sorting 41 (VPS41) is as part of the Homotypic fusion and Protein Sorting (HOPS) complex required for lysosomal fusion events and, independent of HOPS, for regulated secretion. Here, we report three patients with compound heterozygous mutations in VPS41 (VPS41S285P and VPS41R662*; VPS41c.1423‐2A>G and VPS41R662*) displaying neurodegeneration with ataxia and dystonia. Cellular consequences were investigated in patient fibroblasts and VPS41‐depleted HeLa cells. All mutants prevented formation of a functional HOPS complex, causing delayed lysosomal delivery of endocytic and autophagic cargo. By contrast, VPS41S285P enabled regulated secretion. Strikingly, loss of VPS41 function caused a cytosolic redistribution of mTORC1, continuous nuclear localization of Transcription Factor E3 (TFE3), enhanced levels of LC3II, and a reduced autophagic response to nutrient starvation. Phosphorylation of mTORC1 substrates S6K1 and 4EBP1 was not affected. In a C. elegans model of Parkinson’s disease, co‐expression of VPS41S285P/VPS41R662* abolished the neuroprotective function of VPS41 against α‐synuclein aggregates. We conclude that the VPS41 variants specifically abrogate HOPS function, which interferes with the TFEB/TFE3 axis of mTORC1 signaling, and cause a neurodegenerative disease.

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Section: Resultssupporting

confidence: 88%

Neurodegenerative VPS41 variants inhibit HOPS function and mTORC1‐dependent TFEB/TFE3 regulation

et al. 2021

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“…Therefore, we suggest that this culture method represents a model that can reproduce the pathogenesis of NPC disease without concern for the reduction of stemness potentially caused by FBS. FL and electron microscopy revealed characteristic lysosomal accumulation, such as lamellar structures, in NSCs derived from patients with NPC; nevertheless, the frequency at which such accumulation occurred seemed to be lower than that of terminally differentiated cells, such as fibroblasts, macrophages, and hepatocytes [ [37] , [38] , [39] ]. A new finding in the current study was that intracellularly accumulated lipid droplets and mitochondria made contact in NPC patient-derived NSCs.…”

Section: Discussionmentioning

confidence: 99%

A neuropathological cell model derived from Niemann−Pick disease type C patient-specific iPSCs shows disruption of the p62/SQSTM1−KEAP1−NRF2 Axis and impaired formation of neuronal networks

Saito

Miyajima

Iwamoto

et al. 2021

Molecular Genetics and Metabolism Reports

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Niemann−Pick disease type C (NPC) is a rare neurodegenerative disorder caused by a recessive mutation in the NPC1 or NPC2 gene, in which patients exhibit lysosomal accumulation of unesterified cholesterol and glycolipids. Most of the research on NPC has been done in patient-derived skin fibroblasts or mouse models. Therefore, we developed NPC patient neurons derived from induced pluripotent stem cells (iPSCs) to investigate the neuropathological cause of the disease. Although an accumulation of cholesterol and glycolipids, which is characteristic of NPC, was observed in both undifferentiated iPSCs and derived neural stem cells (NSCs), we could not observed the abnormalities in differentiation potential and autophagic activity in such immature cells. However, definite neuropathological features were detected in mature neuronal cells generated from NPC patient-derived iPSCs. Abnormal accumulation of cholesterol and other lipids identified by lipid droplets and number of enlarged lysosomes was more prominent in mature neuronal cells rather than in iPSCs and/or NSCs. Thin-sectioning electron microscopic analysis also demonstrated numerous typical membranous cytoplasmic bodies in mature neuronal cells. Furthermore, TUJ1-positive neurite density was significantly reduced in NPC patient-derived neuronal cells. In addition, disruption of the p62/SQSTM1−KEAP1−NRF2 axis occurred in neurons differentiated from NPC patient-derived iPSCs. These data indicate the impairment of neuronal network formation associated with neurodegeneration in mature neuronal cells derived from patients with NPC.

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“…Electron microscopy (EM) showed a high variation in appearance of endo-lysosomal compartments in primary fibroblasts ( Figure S2), but no swollen lysosomes in the patient-derived cells. This indicates that the patient-specific mutations in VPS41 do not cause a classical lysosomal storage phenotype (37). To study lysosomal acidification and hydrolase activity we performed fluorescence microscopy using lysotracker-Green and MagicRed-cathepsin B.…”

Section: Patient Fibroblasts and Vps41 Ko Cells Contain Small-sized mentioning

confidence: 99%

VPS41recessive mutation causes ataxia and dystonia with retinal dystrophy and mental retardation by inhibiting HOPS function and mTORC1 signaling

Jobling

Burns

et al. 2019

Preprint

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The vacuolar protein sorting protein 41 (VPS41) is a neuroprotective protein in models of Parkinson's disease (PD). As part of the HOPS (Homotypic fusion and Protein Sorting) complex, VPS41 regulates fusion of lysosomes with late endosomes and autophagosomes. Independent of HOPS, VPS41 regulates transport of newly synthesized lysosomal membrane proteins and secretory proteins. Here we report two brothers with compound heterozygous mutations in VPS41 (VPS41 R662* and VPS41 S285P ), born to healthy and non-consanguineous parents. Both patients displayed transient retinal dystrophy, ataxia and dystonia, with brain MRI findings of cerebellar atrophy and a thin saber-shape corpus callosum. Patient-derived fibroblasts contained enzymatically active lysosomes that were poorly reached by endocytic cargo and failed to attract the mTORC1 complex. Consequently, transcription factor TFE3, a driver of autophagy and lysosomal genes, showed continuous nuclear localization which resulted in elevated LC3-II levels and an impaired response to nutrient starvation. CRISPR/CAS VPS41 HeLa knockout cells showed a similar phenotype that could be rescued by wildtype VPS41 but not by VPS41 S285P or VPS41 R662* . mTORC1 inhibition was also seen after knockout of HOPS subunits VPS11 or VPS18. Regulated neuropeptide secretion in PC12 VPS41 knockout cells was rescued by VPS41 S285P expression, indicating that this HOPS-independent function was preserved. Co-expression of the VPS41 S285P and VPS41 R662* variants in a C. elegans model of PD abolished the protective effect of VPS41 against α-synuclein-induced neurodegeneration. We conclude that both disease-associated VPS41 variants specifically abrogate HOPS function, which leads to a delay in endocytic cargo delivery to lysosomes, mTORC1 inhibition and irresponsiveness to autophagic clues. Our studies signify a link between HOPS function and mTORC1 signaling and imply that HOPS function is required for the neuroprotective effect of VPS41 in PD.

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The Role of Ultrastructural Examination in Storage Diseases

Cited by 7 publications

References 23 publications

Neurodegenerative VPS41 variants inhibit HOPS function and mTORC1‐dependent TFEB/TFE3 regulation

Neurodegenerative VPS41 variants inhibit HOPS function and mTORC1‐dependent TFEB/TFE3 regulation

A neuropathological cell model derived from Niemann−Pick disease type C patient-specific iPSCs shows disruption of the p62/SQSTM1−KEAP1−NRF2 Axis and impaired formation of neuronal networks

VPS41recessive mutation causes ataxia and dystonia with retinal dystrophy and mental retardation by inhibiting HOPS function and mTORC1 signaling

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