Genome-wide CRISPR screen reveals v-ATPase as a drug target to lower levels of ALS protein ataxin-2
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE189417 | GEO | 2022/10/18
REPOSITORIES: GEO
Similar Datasets
Project description:Gene-based therapeutic strategies to lower ataxin-2 levels are emerging for neurodegenerative diseases amyotrophic lateral sclerosis (ALS) and spinocerebellar ataxia type 2 (SCA2). To identify additional ways of reducing ataxin-2 levels, we performed a genome-wide screen in human cells for regulators of ataxin-2 and identified RTN4R, the gene encoding the RTN4/NoGo-Receptor, as a top hit. This dataset contains the RNA-sequencing data used to support the conclusions from this study.
2022-10-18 | GSE200530 | GEO
Project description:Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative disorder, which is caused by an unstable CAG-repeat expansion in the SCA2 gene, that encodes a polyglutamine tract (polyQ-tract) expansion in ataxin-2 protein (ATXN2). The RNA-binding protein ATXN2 interacts with the poly(A)-binding protein PABPC1, localizing to ribosomes at the rough endoplasmic reticulum or to polysomes. Under cell stress ATXN2 and PABPC1 show redistribution to stress granules where mRNAs are kept away from translation and from degradation. It is unknown whether ATXN2 associates preferentially with specific mRNAs or how it modulates their processing. Here, we investigated Atxn2 knock-out (Atxn2-/-) mouse liver, cerebellum and midbrain regarding their RNA profile, employing oligonucleotide microarrays for screening and RNA deep sequencing for validation. Modest ~1.4-fold upregulations were observed for the level of many mRNAs encoding ribosomal proteins and other translation pathway factors. Quantitative reverse transcriptase PCR and immunoblots in liver tissue confirmed these effects and demonstrated an inverse correlation also with PABPC1 mRNA and protein. ATXN2 deficiency also enhanced phosphorylation of the ribosomal protein S6, while impairing the global protein synthesis rate, suggesting a block between the enhanced translation drive and the impaired execution. Furthermore, ATXN2 overexpression and deficiency retarded cell cycle progression. ATXN2 mRNA levels showed a delayed phasic twofold increase under amino acid and serum starvation, similar to ATXN3, but different from motor neuron disease genes MAPT and SQSTM1. ATXN2 mRNA levels depended particularly on mTOR signalling. Altogether the data implicate ATXN2 in the adaptation of mRNA translation and cell growth to nutrient availability and stress. Factorial design comparing ataxin-2 knock-out mice with wild type littermates in three different tissues (midbrain, cerebellum, liver) and 3 different ages.
2014-10-31 | E-GEOD-55177 | biostudies-arrayexpress
Project description:Ataxin-2 (ATXN2) is a gene implicated in spinocerebellar ataxia type II (SCA2), amyotrophic lateral sclerosis (ALS) and Parkinsonism. The encoded protein is a therapeutic target for ALS and related conditions. ATXN2 (or Atx2 in insects) functions in translational regulation, mRNA stability, and in the assembly of mRNP-granules, a process mediated by intrinsically disordered regions (IDRs). Previous work has shown that the LSm (Like-Sm) domain of Atx2, which mediates translational activation of some target mRNAs, antagonizes mRNP-granule assembly. Here we advance these findings through a series of experiments on Drosophila and human Ataxin-2 proteins. Results of Targets of RNA-Binding Proteins Identified by Editing (TRIBE) experiments indicate that a polyA-binding protein (PABP) interacting, PAM2 motif of Ataxin-2 may be a major determinant of the mRNA content of Ataxin-2 mRNP granules. Co-localization and co-immunoprecipitation analyses show that structured interactions between Ataxin-2 and PABP additionally help determine protein components of Ataxin-2-associated mRNP granules and contribute to Ataxin-2’s association with stress granules. Finally, in vivo experiments in Drosophila indicate that while the Atx2-LSm domain protects against neurodegeneration, structured PAM2- and unstructured IDR- interactions both promote degeneration. Taken together the data: (a) lead to a proposal for how Ataxin-2 interactions are remodelled during different stages of translational control; (b) show how structured and non-structured interactions of Ataxin-2 contribute differently to the specificity and efficiency of RNP granule condensation; and (c) demonstrate that the Ataxin-2 protein contains multiple activities that may respectively prevent or promote neurodegeneration.
2024-04-22 | GSE196739 | GEO
Project description:Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative disorder, which is caused by an unstable CAG-repeat expansion in the SCA2 gene, that encodes a polyglutamine tract (polyQ-tract) expansion in ataxin-2 protein (ATXN2). The RNA-binding protein ATXN2 interacts with the poly(A)-binding protein PABPC1, localizing to ribosomes at the rough endoplasmic reticulum or to polysomes. Under cell stress ATXN2 and PABPC1 show redistribution to stress granules where mRNAs are kept away from translation and from degradation. It is unknown whether ATXN2 associates preferentially with specific mRNAs or how it modulates their processing. Here, we investigated Atxn2 knock-out (Atxn2-/-) mouse liver, cerebellum and midbrain regarding their RNA profile, employing oligonucleotide microarrays for screening and RNA deep sequencing for validation. Modest ~1.4-fold upregulations were observed for the level of many mRNAs encoding ribosomal proteins and other translation pathway factors. Quantitative reverse transcriptase PCR and immunoblots in liver tissue confirmed these effects and demonstrated an inverse correlation also with PABPC1 mRNA and protein. ATXN2 deficiency also enhanced phosphorylation of the ribosomal protein S6, while impairing the global protein synthesis rate, suggesting a block between the enhanced translation drive and the impaired execution. Furthermore, ATXN2 overexpression and deficiency retarded cell cycle progression. ATXN2 mRNA levels showed a delayed phasic twofold increase under amino acid and serum starvation, similar to ATXN3, but different from motor neuron disease genes MAPT and SQSTM1. ATXN2 mRNA levels depended particularly on mTOR signalling. Altogether the data implicate ATXN2 in the adaptation of mRNA translation and cell growth to nutrient availability and stress.
2014-10-31 | GSE55177 | GEO
Project description:Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative disorder, caused by an unstable CAG-repeat expansion in the SCA2 gene, which encodes a polyglutamine (polyQ) domain expansion in the protein ataxin-2 (ATXN2). The RNA-binding protein ATXN2 interacts with the poly(A)-binding protein PABPC1, localizing to ribosomes in the rough endoplasmic reticulum or in polysomes. Under cell stress ATXN2 and PABPC1 are relocated to stress granules where mRNAs are protected from translation and from degradation. It is unknown whether ATXN2 associates preferentially with specific mRNAs or how it modulates their processing. Here, we investigated the RNA profile of liver and cerebellum from adult Atxn2 knock-out (Atxn2-/-) mice, employing oligonucleotide microarrays for screening and RNA deep sequencing for validation. We consistently observed modest upregulations for the level of many mRNAs encoding proteins of the ribosomal large and small subunit as well as the translation initiation complex. Quantitative reverse transcriptase PCR in liver tissue confirmed these upregulations for the ribosomal components Rpl14, Rpl18, Rps10, Rps18, Gnb2l1, the translation initiation factors Eif2s2, Eif3s6, Pabpc1, and the ribosomal biogenesis modulator Nop10. Quantitative immunoblots substantiated the effects for PABPC1. Thus, the physiological role of ATXN2 modifies the abundance of cellular translation factors.
2017-05-27 | GSE57998 | GEO
Project description:Spinocerebellar ataxia type 2 (SCA2) is among the progressive neurodegenerative polyglutamine (polyQ) diseases. It is caused by a CAG repeat expansion in an encoded region of the ATXN2 gene giving rise to an expanded polyQ domain in the encoded ATXN2 protein. SCA2 is an autosomal dominant disorder characterized by symptoms resulting from neurodegeneration of cerebellar Purkinje cells. To molecularly characterize SCA2 disease progression, we analyzed RNA-sequencing data produced using the cerebella of ATXN2Q127 mice collected at three distinct time points. The ATXN2Q127 mouse model contains 127 CAG repeats in the full-length ATXN2 cDNA under the control of the PC targeted Pcp2 promoter.
2017-12-09 | E-MTAB-6293 | biostudies-arrayexpress
Project description:Ataxin-2 is a conserved translational control protein as well as an important target for ALS therapeutics under development. Despite its clinical and biological significance, Ataxin-2’s activities, mechanisms and functions are not well understood. Using Drosophila Ataxin-2 (Atx2) we investigate how Ataxin-2 IDRs work with structured (Lsm, Lsm-AD and PAM2) domains to enable positive and negative regulation of target mRNAs. Using TRIBE (Targets of RNA-Binding Proteins Identified by Editing) technology, we identified and analysed Atx-2 target mRNAs in the Drosophila brain.
2021-03-03 | GSE153985 | GEO
Project description:Transactive Response DNA-binding protein 43 (TDP-43) is a multifunctional nuclear protein ubiquitously expressed in all tissues. Although it is primarily nuclear, Amyotrophic Lateral Sclerosis (ALS) patients frequently present with cytoplasmic aggregates of TDP-43 in motor neurons on post-mortem examination. Although only about 5% of all ALS patients have a TARDBP mutation (Ghasemi and Brown 2018; Ingre et al. 2015), which is known to cause these insoluble cytoplasmic aggregates, 97% of ALS patients will develop cytoplasmic, insoluble TDP-43 aggregates regardless of whether they have a mutation in TARDBP or not (Ling et al. 2013). Overexpression of wild-type TDP-43 has previously been shown to be a valid model of inducing neurotoxicity as well as a limited amount of cytoplasmic re-localization and aggregation characteristic of ALS (Elden et al. 2010; Wils et al. 2010). In the search for modifiers of toxicity as conferred by TDP-43 overexpression in these models, knockout of Ataxin-2 (ATXN2) was proposed as a protective intervention in the context of TDP-43 perturbation in non-human model systems (Elden et al. 2010, Becker et al. 2017). This promising finding had not previously been validated in a human motor neuron system, nor had a thorough investigation been conducted to determine the mechanism of neuroprotection that ATXN2 knockout employs to drive the therapeutic effect. This sequencing study examines both ATXN2 intact and ATXN2 knockout motor neurons in the context of TDP-43 perturbation to explore global transcriptomic changes that arise from these changes in cellular state that are associated with progression of ALS (ATXN2 intact) and what pathways could be implicated in protection from ALS-associated neurodegeneration (ATXN2 Knockout).
2024-03-23 | GSE261875 | GEO
Project description:Spinocerebellar Ataxia Type 2 (SCA2) is caused by expansion of a polyglutamine encoding triplet repeat in the human ATXN2 gene beyond (CAG)31. This is thought to mediate toxic gain-of-function by protein aggregation, and affect RNA processing, resulting in degenerative processes affecting preferentially cerebellar neurons. As a faithful animal model, we generated a knock-in mouse replacing the single CAG of murine Atxn2 with CAG42, a frequent patient genotype. This expansion size was inherited stably. The mice showed phenotypes with reduced weight and later motor incoordination. Although brain Atxn2 mRNA became elevated, soluble ATXN2 protein levels diminished over time, which might explain partial loss-of-function effects. Deficits in soluble ATXN2 protein correlated with the appearance of insoluble ATXN2, a progressive feature in cerebellum possibly reflecting toxic gains-of-function. Since in vitro ATXN2 overexpression was known to reduce levels of its protein interactor PABPC1, we studied expansion effects on PABPC1. In cortex, PABPC1 transcript, soluble and insoluble protein levels were increased. In more vulnerable cerebellum the progressive insolubility of PABPC1 was accompanied by decreased soluble protein levels, with PABPC1 mRNA showing no compensatory increase. The sequestration of PABPC1 into insolubility by ATXN2 function gains was validated in human cell culture. To understand consequences on mRNA processing, transcriptome profiles at medium and old age in three different tissues were studied and demonstrated selective induction of Fbxw8 in old cerebellum. Fbxw8 is encoded next to the Atxn2 locus and was shown in vitro to decrease the level of expanded insoluble ATXN2 protein. In conclusion, our data support the concept that expanded ATXN2 undergoes progressive insolubility and affects PABPC1 by a toxic gain-of-function mechanism with tissue-specific effects, which may be partially alleviated by the induction of FBXW8. Factorial design comparing Atxn2 knock-in mice with wild type littermates in three different tissues (brainstem, cerebellum, liver)
2012-07-24 | E-GEOD-39640 | biostudies-arrayexpress
Project description:We recently identified the RNA targets of Ataxin-2 by using PAR-CLIP. To elucidate the functional role of Ataxin-2 on target-binding, we have knocked down or overexpressed Ataxin-2 in HEK293T cells and extracted RNAs 48 hrs after transfection. Then, these total RNAs were subjected to whole transcripts microarray analysis to examine how the perturbation of Ataxin-2 expression affects target gene expression. Ataxin-2 was either knocked down using siRNA or overexpressed using the expression construct of Halo-Ataxin-2.Control siRNA was transfected as a control of knockdown experiment. The expression construct of Halo-tag alone was used as a control of overexpression experiment. Forty-eight hrs after transfection, total RNAs were extracted and subjected to microarray analysis.
2014-06-20 | E-GEOD-53189 | biostudies-arrayexpress