RNA-seq analysis revealed aberrant gene expression in motor neurons derived from ALS patient iPSCs bearing SOD1+/A272C mutation
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE95089 | GEO | 2017/04/17
SECONDARY ACCESSION(S): PRJNA376020
REPOSITORIES: GEO
Similar Datasets
Project description:Amyotrophic lateral sclerosis (ALS) is caused by the progressive degeneration of motor neurons. Mutations in the Cu/Zn superoxide dismutase (SOD1) are found in about 20% of patients with familial ALS. Mutant SOD1 causes motor neuron death through an acquired toxic property. Although, molecular mechanism underlying this toxic gain-of-function remains unknown, evidence support the role of mutant SOD1 expression in non-neuronal cells in shaping motor neuron degeneration. We have previously found that in contrast to non-transgenic, SOD1G93A-expressing astrocytes induced apoptosis of co-cultured motor neurons. This prompted us to investigate whether the effect on motor neuron survival was related to a change in the gene expression profile. Through high-density oligonucletide microarrays we found changes in the expression of genes involved in transcription, signaling, cell proliferation, extracellular matrix construction, response to stress and steroid and lipid metabolism. Decorin, a small multifunctional proteoglycan, was the most up-regulated gene. Down-regulated genes included the insulin-like growth factor-1 receptor and the RNA binding protein ROD1. We also analyzed the expression of selected genes in purified motor neurons expressing SOD1G93A and in spinal cord of asymptomatic and early symptomatic ALS-rodent model. The expression of mutated SOD1 in astrocytes cause gene expression changes with potential consequences for its interaction with motor neurons. The astrocyte-specific gene expression profile contributes to the identification of possible candidates for cell type-specific therapies in ALS Keywords: Cell type comparison
2008-12-18 | GSE7441 | GEO
Project description:Although many distinct mutations in a variety of genes are known to cause Amyotrophic Lateral Sclerosis (ALS), it remains poorly understood how they selectively impact motor neuron biology and whether they converge on common pathways to cause neural degeneration. Here, we have combined reprogramming and stem cell differentiation approaches with genome engineering and RNA sequencing to define the transcriptional changes that are induced in human motor neurons by mutant SOD1. Mutant SOD1 protein induced a transcriptional signature indicative of increased oxidative stress, reduced mitochondrial function, altered sub-cellular transport as well as activation of the ER stress and unfolded protein response pathways. Functional studies demonstrated that perturbations in these pathways were indeed the source of altered transcript levels. 5 samples, 2 patient-derived SOD1A4V and 3 isogenic control samples where the mutation has been corrected. All samples are motor neurons derived from induced pluripotent stem cells (iPSCs), and isolated after lentiviral infection with an Hb9:RFP construct and FACS purification. Each sample is a separate biological replicate.
2014-02-03 | E-GEOD-54409 | biostudies-arrayexpress
Project description:Amyotrophic lateral sclerosis (ALS) is caused by the progressive degeneration of motor neurons. Mutations in the Cu/Zn superoxide dismutase (SOD1) are found in about 20% of patients with familial ALS. Mutant SOD1 causes motor neuron death through an acquired toxic property. Although, molecular mechanism underlying this toxic gain-of-function remains unknown, evidence support the role of mutant SOD1 expression in non-neuronal cells in shaping motor neuron degeneration. We have previously found that in contrast to non-transgenic, SOD1G93A-expressing astrocytes induced apoptosis of co-cultured motor neurons. This prompted us to investigate whether the effect on motor neuron survival was related to a change in the gene expression profile. Through high-density oligonucletide microarrays we found changes in the expression of genes involved in transcription, signaling, cell proliferation, extracellular matrix construction, response to stress and steroid and lipid metabolism. Decorin, a small multifunctional proteoglycan, was the most up-regulated gene. Down-regulated genes included the insulin-like growth factor-1 receptor and the RNA binding protein ROD1. We also analyzed the expression of selected genes in purified motor neurons expressing SOD1G93A and in spinal cord of asymptomatic and early symptomatic ALS-rodent model. The expression of mutated SOD1 in astrocytes cause gene expression changes with potential consequences for its interaction with motor neurons. The astrocyte-specific gene expression profile contributes to the identification of possible candidates for cell type-specific therapies in ALS Keywords: Cell type comparison Astrocytes were plated at a density of 2x104 cells/cm2 and maintained as described in Cassina P, et al.J Neurosci Res. 2002;67(1):21-9. Confluent astrocytes monolayers were changed to supplemented L15 medium (Vargas et al., 2006) for 24h before RNA isolation. Total RNA was isolated with RNeasy kit/RNase-Free DNase Set (Qiagen, CA, USA). RNA quality was assessed with the A260/280 ratio and the 2100 Bioanalyzer (Agilent Technologies, CA, USA) to ensure integrity of the samples used for microarray analysis. Double-stranded cDNA was synthesized from 5 μg of total RNA and used for microarray analysis with the Rat Genome 230 2.0 array (Affymetrix, CA, USA). A total of six arrays divided in three control and three transgenic samples were used. Labeling was performed with one-cycle target labeling assay according to Affymetrix, including the eukaryotic poly-A RNA control and the eukaryotic hybridization control kit. Hybridization, washing and scanning were carried out as described in the Affymetrix GeneChip expression analysis technical manual at the Oregon State University’s Center for Genome Research and Biocomputing. Image processing was done using Affymetrix GCOS 1.4 software. The quality of hybridization and overall chip performance was determined by visual inspection of the raw scanned data and the GCOS-generated report file. Microarray data (.CEL files) was normalized using GC-RMA probe-level analysis in ArrayAssist 4.0 (Stratagene, CA, USA). Following variance stabilization and Log transformation, fold chance versus p-value was calculated using nontransgenic (NonTG) samples as base values.
2008-12-18 | E-GEOD-7441 | biostudies-arrayexpress
Project description:Microarray analysis has been applied to the study of ALS in order to investigate gene expression in whole spinal cord homogenates of SOD1 G93A mice and human ALS cases, although the massive presence of glial cells and inflammatory factors has made it difficult to define which gene expression changes were motor neuron specific. Recently, laser capture microdissection (LCM), combined with microarray analysis, has allowed the identification of motor neuron specific changes in gene expression in human ALS cases. The aim of the present study is to combine LCM and microarray analysis to study how motor neurons in the spinal cord of transgenic SOD1 G93A mice and transgenic SOD1 WT respond to stimuli determined by the presence of the human mutant protein throughout the evolution of the stages in motor neuron injury Experiment Overall Design: Motor neurons have been isolated from the spinal cord of G93A mice and non transgenic littermates at different time points and the transcription expression profile of the isolated motor neurons has been analysed
2008-06-11 | E-GEOD-10953 | biostudies-arrayexpress
Project description:Recent genetic studies of ALS patients have identified several forms of ALS that are associated with mutations in RNA binding proteins. In animals or cultured cells, such defects broadly affect RNA metabolism. This raises the question of whether all forms of ALS have general effects on RNA metabolism. We tested this hypothesis in a mouse model of ALS that is transgenic for a human disease-causing mutation in the enzyme superoxide dismutase 1 (SOD1). We analyzed RNA from laser-captured spinal cord motor neuron cell bodies of the mutant SOD1 strain, comparing the RNA profile with that from a corresponding wild-type SOD1 transgenic strain. We prepared the samples from animals that were presymptomatic, but which manifested abnormalities at the cellular level that are seen in ALS, including aggregation of the mutant protein in motor neuron cell bodies and defective morphology of neuromuscular junctions, the connections between neuron and muscle. We observed only minor changes in the level and splicing of RNA in the SOD1 mutant animals as compared with wild-type, suggesting that mutant SOD1 produces the toxic effects of ALS by a mechanism that does not involve global RNA disturbance. RNA-Seq of laser microdissection of motor neuron bodies from two biological replicates each of SOD1 YFP (wildtype 592) and SOD1 G85R YFP (737) transgenic mice.
2012-12-04 | E-GEOD-38820 | biostudies-arrayexpress
Project description:Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease caused by loss of motor neurons. SOD1 may have a toxic role in the pathogenesis of ALS when the protein aggregates in the cytoplasm; increased accumulation of soluble nuclear SOD1 (nSOD1) represents a protective cellular reaction.
2022-01-26 | GSE183204 | GEO
Project description:Ubiquitous expression of ALS-causing mutations in superoxide dismutase 1 (SOD1) provoke non-cell autonomous paralytic disease. By combining ribosome affinity purification and high-throughput sequencing, a cascade of mutant SOD1-dependent, cell type-specific changes are now identified. Initial mutant-dependent damage is restricted to motor neurons and includes synapse and metabolic abnormalities, endoplasmic reticulum (ER) stress, and selective activation of the PERK arm of the unfolded protein response. PERK activation correlates with what we identify to be a naturally low level of ER chaperones in motor neurons. Early changes in astrocytes are to genes involved in inflammation and metabolism and that are targets of the PPAR and LXR transcription factors. Dysregulation of myelination and lipid signaling pathways and activation of ETS transcription factors occur in oligodendrocytes only after disease initiation. Thus, pathogenesis involves a temporal cascade of cell type selective damage initiating in motor neurons, with subsequent damage within glia driving disease propagation. Cell type-specific mRNA was purified by ribosome affinity purification from the spinal cord of bacTRAP reporter mice that label selective cell types by EGFP-tagged ribosome RPL10A. Sequencing libraries were prepared from 3-6 biological replicates for each genotype to determine the mutant induced gene expression changes in specific cell types.
2015-12-09 | E-GEOD-74724 | biostudies-arrayexpress
Project description:Ubiquitous expression of ALS-causing mutations in superoxide dismutase 1 (SOD1) provoke non-cell autonomous paralytic disease. By combining ribosome affinity purification and high-throughput sequencing, a cascade of mutant SOD1-dependent, cell type-specific changes are now identified. Initial mutant-dependent damage is restricted to motor neurons and includes synapse and metabolic abnormalities, endoplasmic reticulum (ER) stress, and selective activation of the PERK arm of the unfolded protein response. PERK activation correlates with what we identify to be a naturally low level of ER chaperones in motor neurons. Early changes in astrocytes are to genes involved in inflammation and metabolism and that are targets of the PPAR and LXR transcription factors. Dysregulation of myelination and lipid signaling pathways and activation of ETS transcription factors occur in oligodendrocytes only after disease initiation. Thus, pathogenesis involves a temporal cascade of cell type selective damage initiating in motor neurons, with subsequent damage within glia driving disease propagation.
2015-12-09 | GSE74724 | GEO
Project description:Gene expression profiling has been performed previously on motor cortex and spinal cord homogenates and of sporadic ALS cases and controls, to identify genes and pathways differentially expressed in ALS. More recent studies have combined the use of laser capture microdissection (LCM) with gene expression profiling to isolate the motor neurons from the surrounding cells, such as microglia and astrocytes, in order to determine those genes differentially expressed in the vulnerable cell population – i.e. motor neuron. The aim of the present study is to combine LCM and microarray analysis to determine those genes and pathways differentially expressed in MNs from human SOD1-related MND and to establish potential pathways for therapeutic intervention. Keywords: Human motor neurons The aim of this study was to determine the gene expression profiles from a small subset of cases which all carry mutations in the SOD1 gene. Expression profiles from isolated motor neurons in SOD1-related ALS cases were compared to those from control motor neurons, in order to establish the pathways implicated in SOD1-related motor neuronal cell death. The 'control' samples were originally submitted to GEO as GSE19332.
2011-01-01 | E-GEOD-20589 | biostudies-arrayexpress
Project description:Missense mutations in the gene for the ubiquitously expressed superoxide dismutase-1 (SOD1) are one of the causes of familial amyotrophic lateral sclerosis (ALS), the most common adult onset motor neuron disease in humans killing selectively large motor neurons. Mice and rats overexpressing mutant SOD1 develop an adult onset neurodegenerative disease with hindlimb-paralysis and subsequent death similar to the human condition. In order to analyze the effects of mutant SOD1 expression onto the most affected cell-type in ALS, a small subpopulation of spinal cord cells, we propose to use laser microdissection to isolate mouse lumbar motor neurons and to assess the changes onto the mRNA expression profile using Affymetrix GeneChips compared to control animals. While two studies applying a genomic approach on the ALS mouse models used the entire spinal cord, contributions of changes to motor neurons were masked by the inflammatory effects of mutant SOD1 and the much larger population of non-motor neuronal cells. What is therefore needed is a cell-type specific expression profile that could reveal dysregulations in the transcriptome of the affected motor neurons. In order to find and analyze disease relevant gene expression profile changes in the mutant vs. the wildtype overexpressing SOD1 mice, we propose to use two different mutant SOD1 lines. Both lines develop the same ALS-like motor neuron disease, however, line 1 (with a G37R mutation) retains the full activity of the SOD1 enzyme while line 2 (with a G85R mutation) has almost no SOD1 enzymatic activity. We believe that by using these two most extreme variants of ALS-inducing mutant SOD1 forms, candidate genes that will be similarly dysregulated in both mutant lines will have great potential to be a disease relevant hit. We will collect from both mutant SOD1 mouse lines lumbar motor neurons from three presymptomatic timepoints, equally distributed during their adult life and before obvious hindlimb-weakness or paralysis signs and compare them to two control lines, negative littermates and wildtype SOD1 overexpressing mice. Furthermore, to determine if their are already mutant SOD1-induced changes since the formation of the affected cells, we will also isolate embryonic spinal cord motor neurons and compare them between mutant SOD1 and wildtype SOD1 overexpressing animals. Both, mice deleted in wildtype SOD1 and mice overexpressing mutant human SOD1 clearly established that the toxic property of mutant SOD1 is based upon a gain-of-toxic function phenomenon rather than a loss-of-function effect, since the SOD1 knock-out mice are overall normal. Furthermore, mice overexpressing wildtype SOD1 are healthy and serve as ideal control animals. Therefore, one of the most important question in ALS is to determine what slowly acting mechanism, or build-up of toxic products is responsible for ultimately killing more than 50-70% of the large lumbar spinal cord motor neurons. We hypothezise that the toxic property of mutant SOD1 must already induce changes onto the transcriptome of the most at-risk cell-type, the large lumbar spinal cord motor neuron, very early on in the presymptomatic phase of the disease course, when the animal is still without any paralysis signs and moving normally around the cage. We will choose 3 timepoints before phenotypic disease-onset that is at 22 weeks for the SOD1G37R line (L42) and at 11 months for the SOD1G85R line (L148). The timepoints are 8, 15 and 18 weeks for SOD1G37R line and 4, 7.5 and 9 months for the SOD1G85R line all compared to age-matched control animals (wildtype SOD1 overexpressing mice (L76) for the SOD1G37R line (high mutant SOD1 levels) and negative littermates for the SOD1G85R line (low mutant SOD1 levels)). Using the Leica lasermicrodissection system on fresh frozen spinal cord sections, we process a single spinal cord sample in 3 days resulting in approximately 4000 ventral horn motor neurons collected from 150 consecutive 20 um sections of the most affected lumbar spinal cord region (L4-L6). To visualize the motor neurons, we have established a fast Nissel-staining protocol that is optimized for speed to preserve the integrity of the RNA. Before lasermicrodissection slides containing 5 spinal cord sections are dessicated for 1 hour and samples are collected directly into RNA-preserving lysis-buffer. Using Stratagene's NanoPrep RNA-extraction columns a yield of around 150 ng of total RNA was obtained (Ribogreen-quantification), that is enough for both the 2-round linear RNA amplification and for future candidate confirmation studies using real-time PCR analysis. We are using the Affymetrix Two-Cycle 3'-Amplification Kit, starting with 100 ng of total RNA and will hybridize on Affymetrix Mouse 430 2.0 GeneChips. Embryonic motor neurons are isolated from e14 mutant SOD1G93A overexpressing rat embryos and compared to wildtype SOD1 overexpressing embryos using a metrazimide gradient followed by a p75-antibody purification in combination with magnetic beads. Cells from one litter of embryos were directly collected into lysis-buffer, yielding at least 200-500 ng of total RNA, enough for one Rat 230 2.0 GeneChip (2-round amplification using 100 ng total RNA). Keywords: time-course
2007-04-11 | GSE7493 | GEO