CLIP-Seq Analysis Enables the Design of Ribosomal RNA Bait Oligonucleotides That Protect Against C9ORF72 ALS/FTD-Associated Poly-GR Pathophysiology
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE182008 | GEO | 2023/11/13
REPOSITORIES: GEO
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Project description:Hexanucleotide repeat expansion in the C9ORF72 gene is the most frequent inherited cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Poly-GR is one of the most toxic dipeptide repeat (DPR) proteins translated from the RNA repeats. It has been shown to affect protein synthesis, but how this contributes to neurodegeneration is not clear. Here, we found that poly-GR inhibits global translation by perturbing translation elongation. We identified that the transcripts with relatively slow elongation rate tend to be further stalled by poly-GR in iPSC-differentiated neurons. This increases ribosome collision and ZAKα-mediated ribotoxic stress response (RSR), which elevates the phosphorylation of p38 and promotes cell death. Knockdown of ZAKα or pharmacological inhibition of p38 can ameliorate the GR toxicity, and improve the survival of C9ORF72-ALS/FTD patient-derived iPSC-neurons. Our study reveals molecular mechanism of poly-GR mediated toxicity on global translatome, and identifies RSR as a potential therapeutic target for C9ORF72-ALS/FTD.
2024-08-23 | GSE230127 | GEO
Project description:Frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) patients with the C9orf72 mutation show predominantly cytoplasmic aggregates of poly-GR and poly-PR proteins that are acutely toxic in various model systems. To identify the molecular mediators of neurotoxicity of poly-GR/PR, we analyzed their interactomes in primary neurons. GFP-(GR)149 and (PR)175-GFP preferentially interacted with RNA-binding proteins, including stress granule-associated and nucleolar proteins, as well as ribosomes. Overexpression of the poly-GR/PR interactors Staufen 1/2 (STAU1/2) and YBX1 led to cytoplasmic aggregation of poly-GR/PR into large stress granule-like inclusions, while the poly-GR/PR interactor nucleophosmin (NPM1) recruited poly-GR into the nucleolus. In addition, poly-PR expression reduced ribosome levels and translation, which is consistent with the widespread reduction of synaptic proteins detected by proteomics. Surprisingly, only GFP-(GR)53, but not GFP-(GR)149, localized to the nucleolus and reduced ribosome levels and translation in neurons, suggesting impaired ribosome biogenesis is driving the acute toxicity commonly observed in vitro. In C9orf72 patient brains, we detected co-aggregation of poly-GR/PR inclusions with ribosomes, but not stress granules. Partial sequestration of ribosomes may chronically impair protein synthesis and contribute to C9orf72 ALS/FTD pathogenesis.
2018-06-27 | PXD008691 | Pride
Project description:The GGGGCC repeat expansion in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). However, it is not known which dysregulated molecular pathways are primarily responsible for disease initiation or progression. We established an inducible mouse model of poly(GR) toxicity in which (GR)80 gradually accumulates in cortical excitatory neurons. Low-level poly(GR) expression induced FTD/ALS-associated synaptic dysfunction and behavioral abnormalities, as well as age-dependent neuronal cell loss, microgliosis and DNA damage, probably caused in part by early defects in mitochondrial function. Poly(GR) bound preferentially to the mitochondrial complex V component ATP5A1 and enhanced its ubiquitination and degradation, consistent with reduced ATP5A1 protein level in both (GR)80 mouse neurons and patient brains. Moreover, inducing ectopic Atp5a1 expression in poly(GR)-expressing neurons or reducing poly(GR) level in adult mice after disease onset rescued poly(GR)-induced neurotoxicity. Thus, poly(GR)-induced mitochondrial defects are a major driver of disease initiation in C9ORF72-related ALS/FTD.
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Project description:C9ORF72-derived dipeptide repeat proteins have emerged as the pathogenic cause of neurodegeneration in amyotrophic lateral sclerosis and frontotemporal dementia (C9-ALS/FTD). However, the mechanisms underlying their expression are not fully understood. Here, we demonstrate that ZNF598, the rate-limiting factor for ribosome-associated quality control (RQC), co-translationally titrates the expression of C9ORF72-derived poly(GR) protein. A Drosophila genetic screen identified key RQC factors as potent modifiers of poly(GR)-induced neurodegeneration. ZNF598 overexpression in human neuroblastoma cells inhibited the nuclear accumulation of poly(GR) protein and decreased its cytotoxicity, whereas ZNF598 deletion had opposing effects. Poly(GR)-encoding sequences in the reporter RNAs caused translational stalling and generated ribosome-associated translation products, sharing molecular signatures with canonical RQC substrates. Furthermore, ZNF598 and listerin 1, the RQC E3 ubiquitin-protein ligase, promoted poly(GR) degradation via the ubiquitin-proteasome pathway. An ALS-relevant ZNF598R69C mutant displayed loss-of-function effects on poly(GR) expression, as well as on general RQC. Moreover, RQC function was impaired in C9-ALS patient-derived neurons, whereas lentiviral overexpression of ZNF598 lowered their poly(GR) expression and suppressed proapoptotic caspase-3 activation. Taken together, we propose that an adaptive nature of the RQC-relevant ZNF598 activity allows the co-translational surveillance to cope with the atypical expression of pathogenic poly(GR) protein, thereby acquiring a neuroprotective function in C9-ALS/FTD.
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Project description:Dysregulation of RNA processing contributes to neurodegenerative diseases, especially amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Expansion of an intronic (GGGGCC)n repeat within the C9ORF72 gene is the most common cause of both FTD and ALS (C9-FTD/ALS), characterized with aberrant repeat RNA foci in the nucleus and noncanonical translation-produced dipeptide repeat (DPR) protein inclusions in the cytoplasm. Here we elucidate that the (GGGGCC)n repeat RNA co-localizes with nuclear speckles and alters their phase separation properties and granule dynamics. Moreover, the essential nuclear speckle scaffold protein SRRM2 is sequestered into the poly-GR cytoplasmic inclusions in C9-FTD/ALS mouse model and patient postmortem tissues, exacerbating the nuclear speckle dysfunction. Impaired nuclear speckle integrity induces global exon-skipping and intron retention in human iPSC-derived neurons. Similar alternative splicing changes can be found in patient postmortem tissues. This work identified novel molecular mechanism of global RNA splicing defects by impaired nuclear speckle function in C9-FTD/ALS and revealed novel potential biomarkers or therapeutic targets.
2024-08-23 | GSE248404 | GEO
Project description:A GGGGCC hexanucleotide repeat expansion in the first intron of C9orf72 is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. Compelling evidence suggests that gain of toxicity from the bidirectionally transcribed repeat expanded RNAs plays a central role in disease pathogenesis. Two potential mechanisms have been proposed including RNA-mediated toxicity and/or the production of toxic dipeptide repeat proteins. In this review, we focus on the role of RNA mediated toxicity in ALS/FTD caused by the C9orf72 mutation and discuss arguments for and against this mechanism. In addition, we summarize how G4C2 repeat RNAs can elicit toxicity and potential therapeutic strategies to mitigate RNA-mediated toxicity.
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Project description:We show that GFP-(GR)100-expressing mice developed age-dependent neurodegeneration, brain atrophy, as well as motor and memory impairments through the accumulation of diffuse, cytoplasmic poly(GR).
2019-02-04 | GSE112931 | GEO
Project description:Amyotrophic lateral sclerosis (ALS) is characterized by a progressive degeneration of upper and lower motor neurons, resulting in fatal paralysis due to denervation of the muscle. Due to genetic, pathological and symptomatic overlap, ALS is now considered a spectrum disease together with frontotemporal dementia (FTD), the second most common cause of dementia in individuals under the age of 65. Interestingly, in both diseases, there is a large prevalence of RNA binding proteins (RBPs) that are mutated and considered disease-causing, or whose dysfunction contribute to disease pathogenesis. The most common shared genetic mutation in ALS/FTD is a hexanucleuotide repeat expansion within intron 1 of C9ORF72 (C9). Three potentially overlapping, putative toxic mechanisms have been proposed: loss of function due to haploinsufficient expression of the C9ORF72 mRNA, gain of function of the repeat RNA aggregates, or RNA foci, and repeat-associated non-ATG-initiated translation (RAN) of the repeat RNA into toxic dipeptide repeats (DPRs). Regardless of the causative mechanism, disease symptoms are ultimately caused by a failure of neurotransmission in three regions: the brain, the spinal cord, and the neuromuscular junction. Here, we review C9 ALS/FTD-associated synaptic dysfunction and aberrant neuronal excitability in these three key regions, focusing on changes in morphology and synapse formation, excitability, and excitotoxicity in patients, animal models, and in vitro models. We compare these deficits to those seen in other forms of ALS and FTD in search of shared pathways, and discuss the potential targeting of synaptic dysfunctions for therapeutic intervention in ALS and FTD patients.
| S-EPMC5997509 | biostudies-literature
Project description:What are the most important and treatable pathogenic mechanisms in C9orf72-FTD/ALS? Model-based efforts to address this question are forging ahead at a blistering pace, often with conflicting results. But what does the human neuropathological literature reveal? Here, we provide a critical review of the human studies to date, seeking to highlight key gaps or uncertainties in our knowledge. First, we engage the C9orf72-specific mechanisms, including C9orf72 haploinsufficiency, repeat RNA foci, and dipeptide repeat protein inclusions. We then turn to some of the most prominent C9orf72-associated features, such as TDP-43 loss-of-function, TDP-43 aggregation, and nuclear transport defects. Finally, we review potential disease-modifying epigenetic and genetic factors and the natural history of the disease across the lifespan. Throughout, we emphasize the importance of anatomical precision when studying how candidate mechanisms relate to neuronal, regional, and behavioral findings. We further highlight methodological approaches that may help address lingering knowledge gaps and uncertainties, as well as other logical next steps for the field. We conclude that anatomically oriented human neuropathological studies have a critical role to play in guiding this fast-moving field toward effective new therapies.
| S-EPMC6546170 | biostudies-literature
Project description:N6-methyladenosine (m6A) is the most prevalent internal mRNA modification and regulates RNA metabolism. Repeat expansion in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD). Here, we showed that m6A is downregulated in C9ORF72-ALS/FTD patient-derived iPSC-differentiated neurons and postmortem brain tissues. The global m6A hypomethylation leads to transcriptome-wide mRNA stabilization and upregulated gene expression, especially ones involved in synaptic activity and neuronal functions. The m6A modification in the C9ORF72 intron sequence preceding the expanded repeats enhances RNA degradation via the nuclear reader YTHDC1. The m6A reduction leads to increased accumulation of repeat RNA and poly-dipeptides. Moreover, the antisense RNA can also be regulated by m6A. Elevating m6A level significantly reduces both sense and antisense repeat RNA and poly-dipeptides, rescues global mRNA homeostasis, and improves survival of C9ORF72-ALS/FTD patient neurons.
2023-06-09 | GSE203578 | GEO