Single-cell RNA sequencing of microglia derived from C9orf72 mutant and isogenic control human iPSCs
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE301532 | GEO | 2025/12/10
REPOSITORIES: GEO
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Project description:Noncoding expansions of a hexanucleotide repeat (GGGGCC) in the C9orf72 gene are the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia. Here we report transgenic mice carrying a bacterial artificial chromosome (BAC) containing the full human C9orf72 gene with either a normal allele (15 repeats) or disease-associated expansion (~100â??1,000 repeats; C9-BACexp). C9-BACexp mice displayed pathologic features seen in C9orf72 expansion patients, including widespread RNA foci and repeat-associated non-ATG (RAN) translated dipeptides, which were suppressed by antisense oligonucleotides targeting human C9orf72. Nucleolin distribution was altered, supporting that either C9orf72 transcripts or RAN dipeptides promote nucleolar dysfunction. Despite early and widespread production of RNA foci and RAN dipeptides in C9-BACexp mice, behavioral abnormalities and neurodegeneration were not observed even at advanced ages, supporting the hypothesis that RNA foci and RAN dipeptides occur presymptomatically and are not sufficient to drive neurodegeneration in mice at levels seen in patients. To compare the RNA Seq profiles from the cortex and spinal cord of transgenic mice expressing unexpanded human C9orf72 (F08, n=4), expanded human C9orf72 (F112, n=3/4), and nontransgenic controls (n=4).
2016-02-10 | E-GEOD-77720 | biostudies-arrayexpress
Project description:Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative condition characterized by loss of motor neurons in the brain and spinal cord. Expansions of a hexanucleotide repeat (GGGGCC) in the noncoding region of the C9ORF72 gene are the most common cause of the familial form of ALS (C9-ALS), as well as frontotemporal lobar degeneration and other neurological diseases. How the repeat expansion causes disease remains unclear, with both loss of function (haploinsufficiency) and gain of function (either toxic RNA or protein products) proposed. We report a cellular model of C9-ALS with motor neurons differentiated from induced pluripotent stem cells (iPSCs) derived from ALS patients carrying the C9ORF72 repeat expansion. No significant loss of C9ORF72 expression was observed, and knockdown of the transcript was not toxic to cultured human motor neurons. Transcription of the repeat was increased, leading to accumulation of GGGGCC repeat–containing RNA foci selectively in C9-ALS iPSC-derived motor neurons. Repeat-containing RNA foci colocalized with hnRNPA1 and Pur-?, suggesting that they may be able to alter RNA metabolism. C9-ALS motor neurons showed altered expression of genes involved in membrane excitability including DPP6, and demonstrated a diminished capacity to fire continuous spikes upon depolarization compared to control motor neurons. Antisense oligonucleotides targeting the C9ORF72 transcript suppressed RNA foci formation and reversed gene expression alterations in C9-ALS motor neurons. These data show that patient-derived motor neurons can be used to delineate pathogenic events in ALS. Transcriptome profiling from iPSC derived motor neurons compared to controls
2013-11-07 | E-GEOD-52202 | biostudies-arrayexpress
Project description:Noncoding expansions of a hexanucleotide repeat (GGGGCC) in the C9orf72 gene are the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia. Here we report transgenic mice carrying a bacterial artificial chromosome (BAC) containing the full human C9orf72 gene with either a normal allele (15 repeats) or disease-associated expansion (~100–1,000 repeats; C9-BACexp). C9-BACexp mice displayed pathologic features seen in C9orf72 expansion patients, including widespread RNA foci and repeat-associated non-ATG (RAN) translated dipeptides, which were suppressed by antisense oligonucleotides targeting human C9orf72. Nucleolin distribution was altered, supporting that either C9orf72 transcripts or RAN dipeptides promote nucleolar dysfunction. Despite early and widespread production of RNA foci and RAN dipeptides in C9-BACexp mice, behavioral abnormalities and neurodegeneration were not observed even at advanced ages, supporting the hypothesis that RNA foci and RAN dipeptides occur presymptomatically and are not sufficient to drive neurodegeneration in mice at levels seen in patients.
2016-02-10 | GSE77720 | GEO
Project description:Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative condition characterized by loss of motor neurons in the brain and spinal cord. Expansions of a hexanucleotide repeat (GGGGCC) in the noncoding region of the C9ORF72 gene are the most common cause of the familial form of ALS (C9-ALS), as well as frontotemporal lobar degeneration and other neurological diseases. How the repeat expansion causes disease remains unclear, with both loss of function (haploinsufficiency) and gain of function (either toxic RNA or protein products) proposed. We report a cellular model of C9-ALS with motor neurons differentiated from induced pluripotent stem cells (iPSCs) derived from ALS patients carrying the C9ORF72 repeat expansion. No significant loss of C9ORF72 expression was observed, and knockdown of the transcript was not toxic to cultured human motor neurons. Transcription of the repeat was increased, leading to accumulation of GGGGCC repeat–containing RNA foci selectively in C9-ALS iPSC-derived motor neurons. Repeat-containing RNA foci colocalized with hnRNPA1 and Pur-α, suggesting that they may be able to alter RNA metabolism. C9-ALS motor neurons showed altered expression of genes involved in membrane excitability including DPP6, and demonstrated a diminished capacity to fire continuous spikes upon depolarization compared to control motor neurons. Antisense oligonucleotides targeting the C9ORF72 transcript suppressed RNA foci formation and reversed gene expression alterations in C9-ALS motor neurons. These data show that patient-derived motor neurons can be used to delineate pathogenic events in ALS.
2013-11-07 | GSE52202 | GEO
Project description:Purpose: The purpose of this experiment is to identify a C9-ALS/FTD specific genomic profile in fibroblast lines that is distinct from sporadic ALS without C9orf72 expansion and non-neurologic control cells. The study will then evaluate the effect on this identified profile of ASO treatment targeting the sense strand RNA transcript of the C9orf72 gene. Methods: Expression profiling was performed on RNAs from fibroblasts of four C9orf72 patients, four control individuals and four sporadic ALS patients using Multiplex Analysis of PolyA-linked Sequences method. Results: Hierarchical clustering of expression values for all genes showed that the four C9orf72 patient lines had an expression profile distinct from control and sporadic ALS lines. Statistical comparison of expression values between the four C9orf72 lines and the four control lines revealed that 122 genes were upregulated (defined by a False Discovery Rate FDR<0.05) and 34 genes were downregulated (defined by a False Discovery Rate FDR <0.05) in C9orf72 patient fibroblasts. Conclusions: A genome wide RNA signature can be defined in fibroblasts with C9orf72 expansion. ASO-mediated reduction of C9orf72 RNA levels in fibroblasts with the hexanucleotide expansion efficiently reduced accumulation of GGGGCC RNA foci. This did not, however, generate a reversal of the C9orf72 RNA profile. Use of Multiplex Analysis of PolyA-linked Sequences to identify expression changes in fibroblasts from amyotrophic lateral sclerosis and frontotemporal dementia patients harboring an hexanucleotide expansion in the C9orf72 gene.
2013-11-12 | E-GEOD-51684 | biostudies-arrayexpress
Project description:Interactome of G3BP1 in C9ORF72 hexanucleotide repeat expansion carrying iPSC derived motor neurons
2025-06-25 | PXD065424 | Pride
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:Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by motor neurons (MN) loss. Mutations in C9orf72 are the leading genetic cause of ALS and involve hexanucleotide repeat expansions (HRE) in the noncoding region of the gene. Neuroinflammation is a key feature and candidate therapeutic target in ALS, particularly in individuals with C9orf72 mutations (C9-ALS), as the gene affects microglial cell function and neuroinflammation. Here, we established a novel protocol to model the ALS neuroinflammatory axis by combining C9-ALS human iPSC-derived spinal MNs, astrocytes, and microglia. The resulting three-dimensional (3D) Spinal Microtissue (SM) represents a scalable multicellular platform for disease modeling and drug discovery. We next performed a screen of 190 FDA-approved compounds in C9-ALS SMs and in microglia-only tissues. Sartans, a class of Angiotensin II Receptor I Blockers (ARB), were identified as top hits in reducing C9-ALS-related neuroinflammation, including IL-6 and IL-8 levels in the culture supernatant. We validated these findings in SMs from two additional C9-ALS hiPSC lines and in isogenic control cultures using telmisartan, a particularly potent and brain penetrant sartan. Further, telmisartan rescued C9-ALS related MN death in both coculture and tricultures conditions and restored MN cell proportions within C9-ALS SMs. Our study suggests that C9-ALS microglia are toxic to MNs, and that telmisartan can attenuate microglia-related neuroinflammation and MN death presenting a potential treatment for C9-ALS.
2025-07-28 | PXD057067 | Pride
Project description:The G4C2 hexanucleotide repeat expansion (HRE) in C9orf72 is the commonest cause of familial amyotrophic lateral sclerosis (ALS). A number of different methods have been used in an attempt to generate isogenic control lines using CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 and NHEJ by deleting the repeat region without replacing it with the normal repeat size, but typically this strategy carries the risk of creating indels and genomic instability. In this study we demonstrate complete correction of an induced pluripotent stem cell (iPSC) line derived from a C9orf72-HR positive ALS patient using CRISPR/Cas9 genome editing and homology directed repair (HDR), resulting in replacement of the excised region with a donor template carrying the wild-type repeat size to maintain the genetic architecture of the locus. The isogenic correction of the C9orf72 HRE restored normal expression and methylation at the C9orf72 locus, reduced intron retention in the edited lines, and abolished pathological phenotypes associated with the C9orf72 HRE expansion in iPSC derived motor neurons (iPSMNs). RNA sequencing of the mutant line identified 2220 differentially expressed genes compared to its isogenic control. Enrichment analysis demonstrated an over-representation of ALS relevant pathways in the presence of the C9orf72 HRE, including calcium ion dependent exocytosis, synaptic transport and the KEGG ALS pathway, as well as new targets of potential relevance to ALS pathophysiology.Complete correction of the C9orf72 HRE in iPS MNs by CRISPR/Cas9 mediated HDR provides an ideal model to study the earliest effects of the hexanucleotide expansion on cellular homeostasis and the key pathways implicated in ALS pathophysiology.
2020-05-19 | GSE139144 | GEO
Project description:Compelling evidence indicates that defects in nucleocytoplasmic transport contribute to the pathogenesis of amyotrophic lateral sclerosis (ALS). In particular, hexanucleotide (G4C2) repeat expansions in C9orf72, the most common cause of genetic ALS, have a widespread impact on the transport machinery that regulates the nucleocytoplasmic distribution of proteins and RNAs, thereby affecting their functions. We have previously reported that the expression of G4C2 hexanucleotide repeats in cultured human and mouse cells caused a marked accumulation of poly(A) mRNAs in cell nuclei, suggesting that mRNA trafficking is impaired by expanded C9orf72-ALS repeats. To further characterize the process, in this work we set out to systematically identify the specific mRNAs that are altered in their nucleocytoplasmic distribution in the presence of C9orf72-ALS repeats. Results from RNAseq profiling of nuclear and cytoplasmic RNA fractions obtained from cells expressing G4C231 repeats show a significant accumulation of mRNAs in the nuclei of G4C231 cells, further suggesting that nuclear retention of mRNAs is a major effect of C9orf72 expanded repeats expression. Interestingly, pathway analysis shows that mRNAs involved in ER-to-Golgi trafficking are particularly enriched among the identified mRNAs. Most importantly, functional studies in cultured cells and Drosophila indicate that the membrane trafficking route regulated by ARFGAP1, a GTPase-activating protein that associates with Golgi, is specifically affected by expanded C9orf72 repeats, as well as by C9orf72-related dipeptide repeat proteins (C9-DPRs), pointing to ER-to-Golgi vesicle-mediated transport as a target of C9orf72 toxicity.
2023-08-21 | GSE235448 | GEO