The debated toxic role of aggregated TDP-43 in amyotrophic lateral sclerosis: a resolution in sight?

Hergesheimer, Rudolf; Chami, Anna A.; Assis, Dênis Reis de; Vourc’h, Patrick; Andres, Christian R.; Corcia, Philippe; Lanznaster, Débora; Blasco, Hélène

doi:10.1093/brain/awz078

Cited by 149 publications

(150 citation statements)

References 204 publications

(252 reference statements)

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“…Our screening system will also be useful for identifying compounds that prevent cytoplasmic TDP-43 aggregation, which may contribute to a novel therapeutic strategy for ALS/FTLD. Although our screening system generally well recapitulated cytoplasmic TDP-43 aggregation in ALS/FTLD, our results were partially inconsistent with some previous reports on the pathology of inherited ALS [17,18]. Notably, we did not nd any evidence of TDP-43 aggregation in the neuronal cells expressing the inherited ALS/FTLD causative genes related to protein degradation (e.g., VCP, UBQLN2, SQSTM1, and TBK1), although the previous studies reported TDP-43 pathology in inherited ALS patients with these mutations [17].…”

Section: Discussioncontrasting

confidence: 99%

“…Although our screening system generally well recapitulated cytoplasmic TDP-43 aggregation in ALS/FTLD, our results were partially inconsistent with some previous reports on the pathology of inherited ALS [17,18]. Notably, we did not nd any evidence of TDP-43 aggregation in the neuronal cells expressing the inherited ALS/FTLD causative genes related to protein degradation (e.g., VCP, UBQLN2, SQSTM1, and TBK1), although the previous studies reported TDP-43 pathology in inherited ALS patients with these mutations [17]. Moreover, the co-localization of TDP-43 and the inherited ALS/FTLD causative gene products are still controversial even though TDP-43 pathology is widely observed in the patients with ALS.…”

Section: Discussioncontrasting

confidence: 99%

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Aggresome Formation and Liquid-liquid Phase Separation Independently Induce Cytoplasmic Aggregation of TAR DNA-binding protein 43

Yamanaka

Watanabe

Inami

et al. 2020

Preprint

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Background Cytoplasmic inclusion of TAR DNA-binding protein 43 (TDP-43) is a pathological hallmark of amyotrophic lateral sclerosis (ALS) and a subtype of frontotemporal lobar degeneration (FTLD). Recent studies have suggested that the formation of cytoplasmic TDP-43 aggregates is dependent on a liquid-liquid phase separation (LLPS) mechanism. However, it is unclear whether TDP-43 pathology is induced through a single intracellular mechanism such as LLPS.Methods We established a TDP-43 aggregation screening system using a cultured neuronal cell line stably expressing EGFP-fused TDP-43 and a mammalian expression library of the inherited ALS/FTLD causative genes. We performed a screening to identify the intracellular mechanisms responsible for TDP-43 aggregation. An immunofluorescence study was conducted using the sporadic ALS spinal cord sections.Results We found that microtubule-related proteins (MRPs) and RNA-binding proteins (RBPs) co-aggregated with TDP-43. MRPs and RBPs sequestered TDP-43 into the cytoplasmic aggregates through distinct mechanisms such as microtubules and LLPS, respectively. The MRPs-induced TDP-43 aggregates were co-localized with aggresomal markers and dependent on histone deacetylase 6 (HDAC6), suggesting that aggresome formation induced the co-aggregation. However, the MRPs-induced aggregates were not affected by 1,6-hexanediol, an LLPS inhibitor. On the other hand, the RBPs-induced TDP-43 aggregates were sensitive to 1,6-hexanediol, but not dependent on microtubules or HDAC6. In sporadic ALS patients, approximately half of skein-like TDP-43 inclusions were co-localized with HDAC6, but round and granular type inclusion were not. Moreover, HDAC6-positive and HDAC6-negative inclusions were found in the same ALS patient, suggesting that the two distinct pathways are both involved in TDP-43 pathology.Conclusion Our findings suggest that at least two distinct pathways (i.e., aggresome formation and LLPS ) are involved in inducing the TDP-43 pathologies.

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Section: Discussioncontrasting

confidence: 99%

Section: Discussioncontrasting

confidence: 99%

Aggresome Formation and Liquid-liquid Phase Separation Independently Induce Cytoplasmic Aggregation of TAR DNA-binding protein 43

Yamanaka

Watanabe

Inami

et al. 2020

Preprint

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“…These results establish altFUS, rather than FUS, as the protein 199 responsible of the inhibition of autophagy. 200 Furthermore, altFUS interactome analysis suggested a role in the cellular stress response, which 201 is known to be altered in ALS with a TDP-43 cytoplasmic accumulation in 98 % of patients 41, 42 . In 202 FUS-linked ALS and some sALS cases, FUS cytoplasmic aggregates or mislocalization are also 203 observed [43][44][45] .…”

Section: As Cells Transfected With Fus or Fus-192mentioning

confidence: 99%

“…Our work shows that FUS synonymous 328 mutations found in patients cluster on altFUS genomic locus. We tested 4 of these mutations, 329 synonymous for the FUS protein and missense for the altFUS protein, and we found that each of 330 them potentiated TDP-43 cytoplasmic aggregation, a pathological hallmark of ALS 42 Throughout the neural conversion and patterning phase (D0-18) the neuroepithelial layer was 419 enzymatically dissociated twice (at D4-5 and D10-12) using dispase (GIBCO, 1 mg ml-1). 420…”

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confidence: 99%

FUSgene is dual-coding with both proteins united in FUS-mediated toxicity

Brunet

Jacques

Nassari

et al. 2019

Preprint

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MAIN 44Amyotrophic Lateral Sclerosis (ALS) is a fatal motor neuron degenerative disease 1-3 characterized 45 by a progressive loss of motor neurons in the brain, brainstem and spinal cord 1 . Most cases are 46 sporadic (sALS), although up to 10 % are familial (fALS) 2 . The first gene associated with ALS (SOD1) 47 was described in 1993 4 . Since then, over 25 genes have been associated with fALS, sALS or both, 48 with the four most common being C9ORF72, SOD1, FUS, and TARDBP 2,5 . Notably, mutations in 49FUS are linked to both fALS and sALS, and present some of the earliest and more rapidly 50 progressive forms of the disease [6][7][8] . FUS is also involved in other neurodegenerative diseases, such 51 as frontotemporal dementia (FUS-FTLD), all characterized by FUS cytoplasmic inclusions 9 . 52 FUS is a nuclear RNA-binding protein, with a C-terminal nuclear localization signal (NLS) 9,10 . The 53 protein is involved in RNA processing, DNA repair and cellular proliferation, although its functions 54 are not precisely elucidated 9 . Most of the mutations associated with neurodegenerative diseases 55 alter FUS NLS [11][12][13] . More recently, mutations in FUS 3'UTR were described in ALS patients and 56 linked to an increased level of FUS mRNA and protein [14][15][16] . Surprisingly, over-expression of wild-57 type FUS provokes an aggressive ALS phenotype in mice and fruit flies, in accordance with findings 58 in yeast and mammalian cells [17][18][19][20][21] . The mechanism of the wild-type or ALS-linked mutated FUS 59 toxicity remains unclear 3,9,22 . 60With currently non-annotated proteins being increasingly reported 23-26 , we hypothesized that the 61 toxicity resulting from wild-type FUS over-expression may come from another, unseen, actor 26 . 62These novel proteins are coded by alternative open reading frames (altORFs) that are located 63 within "non-coding" RNAs (ncRNA), within the 5' or 3' "untranslated" regions (UTR) of mRNAs, or 64 overlapping a known coding sequence (CDS) within a different frame of an mRNA 25-27 . 65 Serendipitous discoveries and ribosome profiling have recently highlighted the distribution of 66 4 altORFs throughout the human genome, and the consequences of their absence from current 67 databases 26 . For example, mass spectrometry-based proteomics has become the gold standard 68 for protein identification and has been extensively used in ALS studies 28,29 . However, if a protein 69 is not annotated, it is not included in the protein database (e.g. UniProtKB), and thus cannot be 70 detected by mass spectrometry. An estimated 50 % of mass spectra from a proteomics 71 experiment are unmatched at the end of the analysis 26,30 . 72 Genome annotations must avoid spurious ORF annotations. Thus unless functional 73 characterization has been published, they rely upon 2 arbitrary criteria: a minimum length of 100 74 codons and a single ORF per transcript. Several groups developed tools to challenge such criteria, 75 such as the sORFs repository 31 and the OpenProt database 27 , which ...

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“…It remains controversial whether TDP-43 aggregation is the principal cause of the motor neuronal degeneration observed in ALS patients, but recent studies shed a light into the key role of TDP-43 aggregates in causing the observed neurodegeneration. Further, TDP-43 aggregates are present in patients with and without any mutations in the TDP-43 encoding gene, TARDPB, and in patients with mutations in other genes involved in ALS pathology, like SOD-1 and C9orf72 (reviewed by [3]). Similar pathophysiological mechanisms are described for both genetic and sporadic ALS patients, and as 97% of ALS patients present TDP-43 aggregation, it is plausible to suggest a link between TDP-43 and some of the pathogenic mechanisms [4,5].…”

Section: Introductionmentioning

confidence: 99%

TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis

Lanznaster

Bourgeais

Bruno

et al. 2019

Cells

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Cytoplasmic TDP-43 aggregates are a hallmark of amyotrophic lateral sclerosis (ALS). Today, only two drugs are available for ALS treatment, and their modest effect prompts researchers to search for new therapeutic options. TDP-43 represents one of the most promising targets for therapeutic intervention, but reliable and reproducible in vitro protocols for TDP-43-mediated toxicity are lacking. Here, we used HEK293T cells transfected with increasing concentrations of TDP-43-expressing plasmid to evaluate different parameters of toxicity and alterations in cellular metabolism. Overexpression of TDP-43 induced aggregates occurrence followed by the detection of 25-and 35-kDa forms of TDP-43. TDP-43 overexpression decreased cell viability and increased cells arrested at G2/M phase and nuclear fragmentation. Analysis of the energetic metabolism showed a tendency to decrease oxidative phosphorylation and increase glycolysis, but no statistical differences were observed. Metabolomics revealed alterations in different metabolites (mainly sphingolipids and glycerophospholipids) in cells overexpressing TDP-43. Our data reveal the main role of TDP-43 aggregation in cellular death and highlight novel insight into the mechanism of cellular toxicity induced by TDP-43. Here, we provide a simple, sensitive, and reliable protocol in a human-derived cell line to be used in high-throughput screenings of potential therapeutic molecules for ALS treatment.

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The debated toxic role of aggregated TDP-43 in amyotrophic lateral sclerosis: a resolution in sight?

Cited by 149 publications

References 204 publications

Aggresome Formation and Liquid-liquid Phase Separation Independently Induce Cytoplasmic Aggregation of TAR DNA-binding protein 43

Aggresome Formation and Liquid-liquid Phase Separation Independently Induce Cytoplasmic Aggregation of TAR DNA-binding protein 43

FUSgene is dual-coding with both proteins united in FUS-mediated toxicity

TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis

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