Mutant TDP-43 in Astrocytes Kills Motor Neurons in Rats through Neurotoxic Gain and Neuroprotective Loss in Astrocytes
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ABSTRACT:
ORGANISM(S): Rattus norvegicus
PROVIDER: GSE42091 | GEO | 2013/05/20
SECONDARY ACCESSION(S): PRJNA179143
REPOSITORIES: GEO
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Project description:Mutation in TDP-43 is causative to amyotrophic lateral sclerosis (ALS). TDP-43 is a multifunctional ribonucleoprotein and is reproted to regulate thousands of genes in neurons, but how astrocytes contribute to TDP-43 pathogenesis is not known. This study examined how mutant TDP-43 in astrocytes kills motor neurons and causes ALS phenotypes. Primary astrocytes were isolated from transgenic rats expressing mutant TDP-43 or from control rats without mutant TDP-43 expression. Cultured astrocytes were induced to express mutant human TDP-43 and their gene expression profiles were determined by microarray assays. Microarray analysis revealed that hundreds of genes were altered in astrocytes in response to mutant TDP-43 expression. As mutant TDP-43 transgene is under the control of tetracycline-regulated pomoter elements (TRE), mutant TDP-43 expression is subjected to Doxycline regulation. Astrocytes isolated from GFAP-tTA/TRE-TDP43M337V rats were desiginated as M337V groups and astrocytes isolated from GFAP-tTA single transgenic rats were desiginated as tTA control groups. Total RNA was isolated from cultured astrocytes at varying times (3, 4, or 6 days after Dox withdrawal) after mutant TDP-43 was induced in astrocytes. Upon mutant TDP-43 induction in astroyctes, gene expression profiles in astroyctes were determined by Illumina Direct Hybridization Assay and compared between tTA and M337V groups at the varying time points of mutant TDP-43 induction.
2013-05-20 | E-GEOD-42091 | biostudies-arrayexpress
Project description:The majority of patients with amyotrophic lateral sclerosis (ALS) have abnormal TDP-43 aggregates in the nucleus and/or cytosol of their surviving neurons and glia. Although accumulating evidence indicates that astroglial dysfunctions contribute to motor neuron degeneration in ALS, the normal physiological functions of TDP-43 in astrocytes are largely unknown and whether the loss of astroglial TDP-43 contributes to ALS remains to be clarified. Here, we showed that TDP-43 deleted astrocytes showed cell-autonomously enhanced GFAP immunoreactivity without affecting astrocyte or microglia proliferation. At the transcriptomic level, TDP-43 deleted astrocytes resemble the A1-reactive astrocytes and induce microglia to increase C1q expression. These astrocytic changes do not cause the loss of motor neurons in spinal cords or denervation at the neuromuscular junctions. In contrast, there was a selective reduction of mature oligodendrocytes, but not oligodendrocyte precursor cells, suggesting a tri-glial dysfunction mediated by TDP-43-deleted astrocytes. Mice with astroglial TDP-43 deletion developed motor, but not sensory, deficits. Taken together, our results demonstrate that TDP-43 is required to maintain the protective functions of astrocytes relevant to the development of motor deficits in mice.
2020-11-01 | GSE156542 | GEO
Project description:Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts. Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS. Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS. Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism. RNA was extracted from lower motor neurons obtained by laser capture microdissection from autopsy material from neurologically healthy controls (n=6) and cases of sporadic ALS (n=3) and ALS due to C9ORF72 mutations (n=3).
2014-06-01 | E-GEOD-56500 | biostudies-arrayexpress
Project description:Mutations in the TDP-43 gene (also called TARDBP), which encodes transactive response DNA-binding protein 43 (TDP-43), have been found in 1–5% of sporadic and familial cases of the devastating adult-onset neurodegenerative disorder amyotrophic lateral sclerosis (ALS). TDP-43 is a predominantly nuclear RNA/DNA-binding protein that has diverse roles in RNA processing, but in diseased motor neurons it is depleted from the nucleus and sequestered into cytoplasmic protein aggregates that are a hallmark of the disease. How the loss of the RNA/DNA-binding activity of TDP-43 contributes to ALS pathogenesis is largely unknown. Using a large-scale RNAi screening approach, we recently identified TDP-43 as a gene required for maintaining genomic stability. We showed that induced pluripotent stem cells (iPSCs) and neurons derived from ALS patients harboring TDP-43 mutations are defective in two major pathways for repair of DNA double-strand breaks, non-homologous end joining and homologous recombination. Accordingly, TDP-43-mutant ALS iPSCs and neurons display increased DNA damage, which we also observed in brain tissues from ALS mice harboring a knock-in ALS-linked Tdp-43 mutation and from human ALS patients. To gain insight into the mechanistic basis by which ALS-associated TDP-43 mutants result in increased DNA damage, we performed transcriptome profiling (RNA-seq) in TDP-43-mutant ALS iPSCs, and found aberrant alternative pre-mRNA splicing of a number of genes involved in DNA repair. Our results suggest a role for TDP-43 in DNA repair and reveal a previously unknown mechanism that likely contributes to the pathogenesis of ALS harboring TDP-43 mutations.
2024-02-23 | GSE167557 | GEO
Project description:Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts. Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS. Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS. Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism. RNA was extracted from fibroblasts grown from neurologically healthy controls (n=6) and 3 groups of patients with ALS: 1) sporadic cases (n=6); 2) cases due to mutations of SOD1 (n=4); 3) cases due to mutations of TARDBP (n=3). The three ALS groups were compared to the controls.
2014-06-01 | E-GEOD-33855 | biostudies-arrayexpress
Project description:Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts. Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS. Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS. Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism. RNA was extracted from NSC34 motor neuronal cells depleted of TDP-43 by shRNA (n=4), treated with control shGFP (n=4), and treated with control shLuciferase (n=3).
2014-06-01 | E-GEOD-56503 | biostudies-arrayexpress
Project description:Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts. Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS. Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS. Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism.
2014-06-01 | GSE33855 | GEO
Project description:Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts. Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS. Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS. Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism.
2014-06-01 | GSE56503 | GEO
Project description:Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts. Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS. Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS. Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism.
2014-06-01 | GSE56500 | GEO
Project description:Motor neurons (MNs) and astrocytes (ACs) are implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS), but their interaction and the sequence of molecular events leading to MN death remain unresolved. Herewe optimized directed differentiation of induced pluripotent stem cells (iPSCs) into highly enriched (>85%) functional populations of spinal cord MNs and ACs. We identifysignificantlyincreased cytoplasmic TDP-43 and ER stress as primary pathogenic events in patient-specific valosin-containing protein (VCP)-mutant MNs, with secondary mitochondrial dysfunction and oxidative stress. Cumulatively these cellular stresses result in synaptic pathology and cell death in VCP-mutant MNs. We additionallyidentify a cell-autonomous VCP-mutant AC survival phenotype, which is not attributable to the same molecular pathology occurring in VCP-mutant MNs. Finally, through iterative co-culture experiments, we uncover non-cell-autonomous effects of VCP-mutant ACs on both control and mutant MNs. This work elucidates molecular events and cellular interplay thatcould guide future therapeutic strategies in ALS.
2017-06-10 | GSE99843 | GEO