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TDP-43 regulation of stress granule dynamics in neurodegenerative disease-relevant cell types.

ABSTRACT: Stress granules (SGs) are cytoplasmic foci that form in response to various external stimuli and are essential to cell survival following stress. SGs are studied in several diseases, including ALS and FTD, which involve the degeneration of motor and cortical neurons, respectively, and are now realized to be linked pathogenically by TDP-43, originally discovered as a component of ubiquitin-positive aggregates within patients' neurons and some glial cells. So far, studies to undercover the role of TDP-43 in SGs have used primarily transformed cell lines, and thus rely on the extrapolation of the mechanisms to cell types affected in ALS/FTD, potentially masking cell specific effects. Here, we investigate SG dynamics in primary motor and cortical neurons as well as astrocytes. Our data suggest a cell and stress specificity and demonstrate a requirement for TDP-43 for efficient SG dynamics. In addition, based on our in vitro approach, our data suggest that aging may be an important modifier of SG dynamics which could have relevance to the initiation and/or progression of age-related neurodegenerative diseases.

SUBMITTER: Khalfallah Y

PROVIDER: S-EPMC5953947 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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TDP-43 regulation of stress granule dynamics in neurodegenerative disease-relevant cell types.

Khalfallah Yousra Y Kuta Rachel R Grasmuck Camille C Prat Alexandre A Durham Heather D HD Vande Velde Christine C

Scientific reports 20180515 1

Stress granules (SGs) are cytoplasmic foci that form in response to various external stimuli and are essential to cell survival following stress. SGs are studied in several diseases, including ALS and FTD, which involve the degeneration of motor and cortical neurons, respectively, and are now realized to be linked pathogenically by TDP-43, originally discovered as a component of ubiquitin-positive aggregates within patients' neurons and some glial cells. So far, studies to undercover the role of ...[more]

PMID: 29765078

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Project description:Amyotrophic Lateral Sclerosis (ALS) is characterized by degeneration of motor neurons in the brain and spinal cord. Cytoplasmic inclusions of TDP-43 are frequently reported in motor neurons of ALS patients. TDP-43 has also been shown to associate with stress granules (SGs), a complex of proteins and mRNAs formed in response to stress stimuli that temporarily sequester mRNA translation. The effect of pathogenic TDP-43 mutations within glycine-rich regions (where the majority of ALS-causing TDP-43 mutations occur) on SG dynamics in motor neurons is poorly understood. To address this issue, we generated murine NSC-34 cell lines that stably over-express wild type TDP-43 (TDP-43 W T ) or mutant forms (ALS-causing TDP-43 mutations TDP-43 A315T or TDP-43 M337V). We then differentiated these NSC-34 lines into motoneuron-like cells and evaluated SG formation and disassembly kinetics in response to oxidative or osmotic stress treatment. Wild type and mutant TDP-43 appeared to be largely retained in the nucleus following exposure to arsenite-induced oxidative stress. Upon arsenite removal, mutant TDP-43 clearly accumulated within HuR positive SGs in the cytoplasm, whereas TDP-43 W T remained mostly within the nucleus. 24 h following arsenite removal, all SGs were disassembled in both wild type and mutant TDP-43 expressing cells. By contrast, we observed significant differences in the dynamics of mutant TDP-43 association with SGs in response to hyperosmotic stress. Specifically, in response to sorbitol treatment, TDP-43 W T remained in the nucleus, whereas mutant TDP-43 relocalized to HuR positive SGs in the cytoplasm following exposure to sorbitol stress, resulting in a significant increase in TDP-43 SG numbers. These SGs remained assembled for 24 h following removal of sorbitol. Our data reveal that under certain stress conditions the rates of SG formation and disassembly is modulated by TDP-43 mutations associated with ALS, and suggest that this may be an early event in the seeding of insoluble cytoplasmic inclusions observed in ALS.

Project description:BackgroundA G4C2 hexanucleotide repeat expansion in the noncoding region of C9orf72 is the major genetic cause of frontotemporal dementia and amyotrophic lateral sclerosis (c9FTD/ALS). Putative disease mechanisms underlying c9FTD/ALS include toxicity from sense G4C2 and antisense G2C4 repeat-containing RNA, and from dipeptide repeat (DPR) proteins unconventionally translated from these RNA products.MethodsIntracerebroventricular injections with adeno-associated virus (AAV) encoding 2 or 149 G4C2 repeats were performed on postnatal day 0, followed by assessment of behavioral and neuropathological phenotypes.ResultsRelative to control mice, gliosis and neurodegeneration accompanied by cognitive and motor deficits were observed in (G4C2)149 mice by 6 months of age. Recapitulating key pathological hallmarks, we also demonstrate that sense and antisense RNA foci, inclusions of poly(GA), poly(GP), poly(GR), poly(PR), and poly(PA) DPR proteins, and inclusions of endogenous phosphorylated TDP-43 (pTDP-43) developed in (G4C2)149 mice but not control (G4C2)2 mice. Notably, proteins that play a role in the regulation of stress granules - RNA-protein assemblies that form in response to translational inhibition and that have been implicated in c9FTD/ALS pathogenesis - were mislocalized in (G4C2)149 mice as early as 3 months of age. Specifically, we observed the abnormal deposition of stress granule components within inclusions immunopositive for poly(GR) and pTDP-43, as well as evidence of nucleocytoplasmic transport defects.ConclusionsOur in vivo model of c9FTD/ALS is the first to robustly recapitulate hallmark features derived from both sense and antisense C9orf72 repeat-associated transcripts complete with neurodegeneration and behavioral impairments. More importantly, the early appearance of persistent pathological stress granules prior to significant pTDP-43 deposition implicates an aberrant stress granule response as a key disease mechanism driving TDP-43 proteinopathy in c9FTD/ALS.

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TDP-43 regulation of stress granule dynamics in neurodegenerative disease-relevant cell types.

Publications

TDP-43 regulation of stress granule dynamics in neurodegenerative disease-relevant cell types.

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