Targeted DNA methylation of the ALS/FTD gene C9orf72 via endogenous DNA damage repair pathways (I)
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PROVIDER: PRJNA557108 | ENA |
REPOSITORIES: ENA
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Project description:DNA methyltransferases (DNMTs) are thought to be involved in the cellular response to DNA damage, thus linking DNA repair mechanisms with DNA methylation. This study presents a novel method of targeted DNA methylation that utilizes endogenous DNA double strand break repair pathways and applies it to the neurodegenerative disease gene C9orf72. A double strand break induced by CRISPR/cas9 in the promoter of C9orf72 is sufficient to induce DNA methylation, and methylation can be precisely targeted through the process of homology directed repair (HDR) via delivery of an in vitro methylated exogenous repair template. Long methylated double stranded DNA templates induce more methylation than shorter templates and with higher efficiency than a dCas9-DNMT3a fusion protein construct. Genome-wide methylation analysis reveals no significant off-target methylation changes when inducing methylation via HDR, whereas the dCas9-DNMT3a fusion construct causes significant off-target methylation at over 67,000 sites. This method is applied to generate a patient derived iPSC model of amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD) that exhibits stable DNA methylation patterns similar to those seen in patients. Using this model, it’s shown for the first time that DNA methylation of the 5’ regulatory region directly reduces C9orf72 expression and increases histone H3K9 tri-methylation levels.
2019-10-11 | GSE134995 | GEO
Project description:DNA methyltransferases (DNMTs) are thought to be involved in the cellular response to DNA damage, thus linking DNA repair mechanisms with DNA methylation. This study presents a novel method of targeted DNA methylation that utilizes endogenous DNA double strand break repair pathways and applies it to the neurodegenerative disease gene C9orf72. A double strand break induced by CRISPR/cas9 in the promoter of C9orf72 is sufficient to induce DNA methylation, and methylation can be precisely targeted through the process of homology directed repair (HDR) via delivery of an in vitro methylated exogenous repair template. Long methylated double stranded DNA templates induce more methylation than shorter templates and with higher efficiency than a dCas9-DNMT3a fusion protein construct. Genome-wide methylation analysis reveals no significant off-target methylation changes when inducing methylation via HDR, whereas the dCas9-DNMT3a fusion construct causes significant off-target methylation at over 67,000 sites. This method is applied to generate a patient derived iPSC model of amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD) that exhibits stable DNA methylation patterns similar to those seen in patients. Using this model, it’s shown for the first time that DNA methylation of the 5’ regulatory region directly reduces C9orf72 expression and increases histone H3K9 tri-methylation levels.
2019-10-11 | GSE134994 | GEO
Project description:Targeted DNA methylation of the ALS/FTD gene C9orf72 via endogenous DNA damage repair pathways
| PRJNA557106 | ENA
Project description:To differentiate, characterize and examine intrinsic phenotypes of C9orf72 ALS/FTD patient-derived induced pluripotent stem cells into microglia (iPSC-MG). Moderate molecular and functional differences were observed in C9orf72 iPSC-MG mono-cultures despite the presence of C9orf72 pathological features.
2023-05-23 | GSE233262 | GEO
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 | GSE203580 | GEO
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 | GSE203579 | GEO
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
Project description:Targeted DNA methylation of the ALS/FTD gene C9orf72 via endogenous DNA damage repair pathways (II)
| PRJNA557107 | ENA
Project description:A hexanucleotide repeat expansion (HRE) in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Human imaging and experimental studies hint at early changes in the brain in C9-ALS/FTD, which remain poorly understood. To define these changes, we used cerebral organoid models derived from C9-ALS/FTD patients and controls to create a single cell RNA sequencing dataset at day 90. Together, these dataset will help to shed light on initial pathologies crucial for understanding disease onset and the design of therapeutic strategies.
2024-09-06 | GSE264012 | GEO
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