Activation of Src family kinase ameliorates secretory trafficking in mutant prion protein cells
| Autor: | Elena Quaglio, Vanessa Capone, Vladimiro Artuso, Fabio Fiordaliso, Ignazio Roiter, Davide Ortolan, Michele Sallese, Alessandro Corbelli, Galina V. Beznoussenko, Elena Restelli, Manuela Pozzoli, Roberto Chiesa |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
Protein Folding HFs human fibroblasts PDI protein disulfide isomerase Mutant Golgi Apparatus Endoplasmic Reticulum medicine.disease_cause VSV-G vesicular stomatite virus glycoprotein Biochemistry Creutzfeldt-Jakob Syndrome DMEM Dulbecco’s modified Eagle’s medium Mice Gerstmann-Straussler-Scheinker Disease Src family kinase protein misfolding genetic prion disease Cells Cultured Mutation Secretory Pathway FFI fatal familial insomnia moPrP mouse PrP Chemistry MEFs mouse embryonic fibroblasts GPCR G-protein-coupled receptors VGCCs voltage-gated calcium channels GSS Gerstmann–Sträussler–Scheinker Cell biology src-Family Kinases CCD charge-coupled device PM plasma membrane symbols RT room temperature PC-I procollagen I protein trafficking Intracellular Research Article GPI glycosylphosphatidylinositol GFP-GPI GPI-anchored green fluorescent protein reporter Mice Transgenic gCJD genetic Creutzfeldt–Jakob disease Rab GDP dissociation inhibitor (GDI) CNS central nervous system Insomnia Fatal Familial Prion Proteins AMPARs α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors KDEL-R KDEL receptor MEM minimum essential medium ER endoplasmic reticulum 03 medical and health sciences symbols.namesake YAG yttrium aluminum garnet Tg transgenic medicine Animals Humans IF immunofluorescence Molecular Biology Secretory pathway Fatal familial insomnia EM electron microscopy PrP prion protein KO knockout 030102 biochemistry & molecular biology Endoplasmic reticulum Cell Biology Golgi apparatus medicine.disease LD labeling density WT wild-type PG14 moPrP with a nine-octapeptide repeat insertion Enzyme Activation Mice Inbred C57BL Disease Models Animal PenStrep penicillin/streptomycin 030104 developmental biology prion protein Mice Inbred CBA SFK Src family kinase BSA bovine serum albumin mo mouse |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 0021-9258 |
| DOI: | 10.1016/j.jbc.2021.100490 |
| Popis: | Fatal familial insomnia (FFI), genetic Creutzfeldt–Jakob disease (gCJD), and Gerstmann–Sträussler–Scheinker (GSS) syndrome are neurodegenerative disorders linked to prion protein (PrP) mutations. The pathogenic mechanisms are not known, but increasing evidence points to mutant PrP misfolding and retention in the secretory pathway. We previously found that the D178N/M129 mutation associated with FFI accumulates in the Golgi of neuronal cells, impairing post-Golgi trafficking. In this study we further characterized the trafficking defect induced by the FFI mutation and tested the 178N/V129 variant linked to gCJD and a nine-octapeptide repeat insertion associated with GSS. We used transfected HeLa cells, embryonic fibroblasts and primary neurons from transgenic mice, and fibroblasts from carriers of the FFI mutation. In all these cell types, the mutant PrPs showed abnormal intracellular localizations, accumulating in the endoplasmic reticulum (ER) and Golgi. To test the efficiency of the membrane trafficking system, we monitored the intracellular transport of the temperature-sensitive vesicular stomatite virus glycoprotein (VSV-G), a well-established cargo reporter, and of endogenous procollagen I (PC-I). We observed marked alterations in secretory trafficking, with VSV-G accumulating mainly in the Golgi complex and PC-I in the ER and Golgi. A redacted version of mutant PrP with reduced propensity to misfold did not impair VSV-G trafficking, nor did artificial ER or Golgi retention of wild-type PrP; this indicates that both misfolding and intracellular retention were required to induce the transport defect. Pharmacological activation of Src family kinase (SFK) improved intracellular transport, suggesting that mutant PrP impairs secretory trafficking through corruption of SFK-mediated signaling. |
| Databáze: | OpenAIRE |
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