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Loss of function of the ALS protein SigR1 leads to ER pathology associated with defective autophagy and lipid raft disturbances.

ABSTRACT: Intracellular accumulations of altered, misfolded proteins in neuronal and other cells are pathological hallmarks shared by many neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). Mutations in several genes give rise to familial forms of ALS. Mutations in Sigma receptor 1 have been found to cause a juvenile form of ALS and frontotemporal lobar degeneration (FTLD). We recently described altered localization, abnormal modification and loss of function of SigR1 in sporadic ALS. In order to further elucidate the molecular mechanisms underlying SigR1-mediated alterations in sporadic and familial ALS, we extended our previous studies using neuronal SigR1 knockdown cell lines. We found that loss of SigR1 leads to abnormal ER morphology, mitochondrial abnormalities and impaired autophagic degradation. Consistent with these results, we found that endosomal trafficking of EGFR is impaired upon SigR1 knockdown. Furthermore, in SigR1-deficient cells the transport of vesicular stomatitis virus glycoprotein is inhibited, leading to the accumulation of this cargo protein in the Golgi apparatus. Moreover, depletion of SigR1 destabilized lipid rafts and associated calcium mobilization, confirming the crucial role of SigR1 in lipid raft and intracellular calcium homeostasis. Taken together, our results support the notion that loss of SigR1 function contributes to ALS pathology by causing abnormal ER morphology, lipid raft destabilization and defective endolysosomal pathways.

SUBMITTER: Vollrath JT

PROVIDER: S-EPMC4611717 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Loss of function of the ALS protein SigR1 leads to ER pathology associated with defective autophagy and lipid raft disturbances.

Vollrath J T JT Sechi A A Dreser A A Katona I I Wiemuth D D Vervoorts J J Dohmen M M Chandrasekar A A Prause J J Brauers E E Jesse C M CM Weis J J Goswami A A

Cell death & disease 20140612

Intracellular accumulations of altered, misfolded proteins in neuronal and other cells are pathological hallmarks shared by many neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). Mutations in several genes give rise to familial forms of ALS. Mutations in Sigma receptor 1 have been found to cause a juvenile form of ALS and frontotemporal lobar degeneration (FTLD). We recently described altered localization, abnormal modification and loss of function of SigR1 in sporadic AL ...[more]

PMID: 24922074

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Project description:How lipid asymmetry impacts ordered lipid domain (raft) formation may yield important clues to how ordered domain formation is regulated in vivo. Under some conditions, a sphingomyelin (SM) and cholesterol-rich ordered domain in one leaflet induces ordered domain formation in the corresponding region of an opposite leaflet composed of unsaturated phosphatidylcholine (PC) and cholesterol. In other conditions, the formation of ordered domains in a SM and cholesterol-rich leaflet can be suppressed by an opposite leaflet containing unsaturated PC and cholesterol. To explore how PC unsaturation influences the balance between these behaviors, domain formation was studied in asymmetric and symmetric lipid vesicles composed of egg SM, cholesterol, and either unsaturated dioleoyl PC (DOPC) or 1-palmitoyl 2-oleoyl PC (POPC). The temperature dependence of ordered domain formation was measured using Förster resonance energy transfer, which detects nanodomains as well as large domains. In cholesterol-containing asymmetric SM+PC outside/PC inside vesicles, the PC-containing inner leaflet tended to destabilize ordered domain formation in the SM+PC-containing outer leaflet relative to ordered domain stability in cholesterol-containing symmetric SM/PC vesicles. Residual ordered domain formation was detected in cholesterol-containing asymmetric SM+DOPC outside/DOPC inside vesicles, but ordered domain formation was completely or almost completely suppressed by asymmetry in cholesterol-containing SM+POPC outside/POPC inside vesicles over the entire temperature range measured. Suppression of ordered domain formation in the latter vesicles was confirmed by fluorescence anisotropy measurements. Because mixtures of SM, POPC, and cholesterol form domains in symmetric vesicles, and this lipid composition mimics plasma membranes to a significant degree, it is possible that under some conditions in vivo the loss of lipid asymmetry could trigger ordered domain formation.

Project description:Mitochondria-associated membranes (MAMs) are essential communication subdomains of the endoplasmic reticulum (ER) that interact with mitochondria. We previously demonstrated that, upon macroautophagy/autophagy induction, AMBRA1 is recruited to the BECN1 complex and relocalizes to MAMs, where it regulates autophagy by interacting with raft-like components. ERLIN1 is an endoplasmic reticulum lipid raft protein of the prohibitin family. However, little is known about its association with the MAM interface and its involvement in autophagic initiation. In this study, we investigated ERLIN1 association with MAM raft-like microdomains and its interaction with AMBRA1 in the regulation of the autophagic process. We show that ERLIN1 interacts with AMBRA1 at MAM raft-like microdomains, which represents an essential condition for autophagosome formation upon nutrient starvation, as demonstrated by knocking down ERLIN1 gene expression. Moreover, this interaction depends on the "integrity" of key molecules, such as ganglioside GD3 and MFN2. Indeed, knocking down ST8SIA1/GD3-synthase or MFN2 expression impairs AMBRA1-ERLIN1 interaction at the MAM level and hinders autophagy. In conclusion, AMBRA1-ERLIN1 interaction within MAM raft-like microdomains appears to be pivotal in promoting the formation of autophagosomes.Abbreviations: ACSL4/ACS4: acyl-CoA synthetase long chain family member 4; ACTB/β-actin: actin beta; AMBRA1: autophagy and beclin 1 regulator 1; ATG14: autophagy related 14; BECN1: beclin 1; CANX: calnexin; Cy5: cyanine 5; ECL: enhanced chemiluminescence; ER: endoplasmic reticulum; ERLIN1/KE04: ER lipid raft associated 1; FB1: fumonisin B1; FE: FRET efficiency; FRET: Förster/fluorescence resonance energy transfer; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GD3: aNeu5Ac(2-8)aNeu5Ac(2-3)bDGalp(1-4)bDGlcp(1-1)ceramide; HBSS: Hanks' balanced salt solution; HRP: horseradish peroxidase; LMNB1: lamin B1; mAb: monoclonal antibody; MAMs: mitochondria-associated membranes; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MFN2: mitofusin 2; MTOR: mechanistic target of rapamycin kinase; MYC/cMyc: proto-oncogene, bHLH transcription factor; P4HB: prolyl 4-hydroxylase subunit beta; pAb: polyclonal antibody; PE: phycoerythrin; SCAP/SREBP: SREBF chaperone; SD: standard deviation; ST8SIA1: ST8 alpha-N-acetyl-neuraminide alpha-2,8 sialyltransferase 1; SQSTM1/p62: sequestosome 1; TOMM20: translocase of outer mitochondrial membrane 20; TUBB/beta-tubulin: tubulin beta class I; ULK1: unc-51 like autophagy activating kinase 1; VDAC1/porin: voltage dependent anion channel 1.

Dataset Information

Loss of function of the ALS protein SigR1 leads to ER pathology associated with defective autophagy and lipid raft disturbances.

Publications

Loss of function of the ALS protein SigR1 leads to ER pathology associated with defective autophagy and lipid raft disturbances.

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