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Cellular stress pathways that inhibit translation initiation lead to transient formation of cytoplasmic RNA/protein complexes known as stress granules. Many of the proteins found within stress granules and the dynamics of stress granule formation and dissolution are implicated in neurodegenerative disease. Whether stress granule formation is protective or harmful in neurodegenerative conditions is not known. To address this, we took advantage of the alphavirus protein nsP3, which selectively binds dimers of the central stress granule nucleator protein G3BP and markedly reduces stress granule formation without directly impacting the protein translational inhibitory pathways that trigger stress granule formation. In Drosophila and rodent neurons, reducing stress granule formation with nsP3 had modest impacts on lifespan even in the setting of serial stress pathway induction. In contrast, reducing stress granule formation in models of ataxia, amyotrophic lateral sclerosis and frontotemporal dementia largely exacerbated disease phenotypes. These data support a model whereby stress granules mitigate, rather than promote, neurodegenerative cascades.
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Tandem repeat sequences comprise approximately 8% of the human genome and are linked to more than 50 neurodegenerative disorders. Accurate characterization of disease-associated repeat loci remains resource intensive and often lacks high resolution genotype calls. We introduce a multiplexed, targeted nanopore sequencing panel and HMMSTR, a sequence-based tandem repeat copy number caller. HMMSTR outperforms current signal- and sequence-based callers relative to two assemblies and we show it performs with high accuracy in heterozygous regions and at low read coverage. The flexible panel allows us to capture disease associated regions at an average coverage of >150x. Using these tools, we successfully characterize known or suspected repeat expansions in patient derived samples. In these samples we also identify unexpected expanded alleles at tandem repeat loci not previously associated with the underlying diagnosis. This genotyping approach for tandem repeat expansions is scalable, simple, flexible, and accurate, offering significant potential for diagnostic applications and investigation of expansion co-occurrence in neurodegenerative disorders.
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A GGGGCC (G4C2) hexanucleotide repeat expansion in C9ORF72 causes amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD), while a CGG trinucleotide repeat expansion in FMR1 leads to the neurodegenerative disorder Fragile X-associated tremor/ataxia syndrome (FXTAS). These GC-rich repeats form RNA secondary structures that support repeat-associated non-AUG (RAN) translation of toxic proteins that contribute to disease pathogenesis. Here we assessed whether these same repeats might trigger stalling and interfere with translational elongation. We find that depletion of ribosome-associated quality control (RQC) factors NEMF, LTN1 and ANKZF1 markedly boost RAN translation product accumulation from both G4C2 and CGG repeats while overexpression of these factors reduces RAN production in both reporter assays and C9ALS/FTD patient iPSC-derived neurons. We also detected partially made products from both G4C2 and CGG repeats whose abundance increased with RQC factor depletion. Repeat RNA sequence, rather than amino acid content, is central to the impact of RQC factor depletion on RAN translation-suggesting a role for RNA secondary structure in these processes. Together, these findings suggest that ribosomal stalling and RQC pathway activation during RAN translation inhibits the generation of toxic RAN products. We propose augmenting RQC activity as a therapeutic strategy in GC-rich repeat expansion disorders.
Asunto(s)
Esclerosis Amiotrófica Lateral , Proteína C9orf72 , Demencia Frontotemporal , Biosíntesis de Proteínas , Proteínas Ribosómicas , Expansión de Repetición de Trinucleótido , Humanos , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Ataxia , Proteína C9orf72/genética , Proteína C9orf72/metabolismo , Expansión de las Repeticiones de ADN/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/metabolismo , Demencia Frontotemporal/genética , Demencia Frontotemporal/metabolismo , Secuencia Rica en GC , Células HEK293 , Células Madre Pluripotentes Inducidas/metabolismo , Neuronas/metabolismo , Ribosomas/metabolismo , Ribosomas/genética , Temblor , Expansión de Repetición de Trinucleótido/genética , Proteínas Ribosómicas/metabolismoRESUMEN
Cellular stress pathways that inhibit translation initiation lead to transient formation of cytoplasmic RNA/protein complexes known as stress granules. Many of the proteins found within stress granules and the dynamics of stress granule formation and dissolution are implicated in neurodegenerative disease. Whether stress granule formation is protective or harmful in neurodegenerative conditions is not known. To address this, we took advantage of the alphavirus protein nsP3, which selectively binds dimers of the central stress granule nucleator protein G3BP (rin in Drosophila) and markedly reduces stress granule formation without directly impacting the protein translational inhibitory pathways that trigger stress granule formation. In Drosophila and rodent neurons, reducing stress granule formation with nsP3 had modest impacts on lifespan even in the setting of serial stress pathway induction. In contrast, reducing stress granule formation in models of ataxia, amyotrophic lateral sclerosis and frontotemporal dementia largely exacerbated disease phenotypes. These data support a model whereby stress granules mitigate, rather than promote, neurodegenerative cascades.
RESUMEN
The premutation of the fragile X messenger ribonucleoprotein 1 (FMR1) gene is characterized by an expansion of the CGG trinucleotide repeats (55 to 200 CGGs) in the 5' untranslated region and increased levels of FMR1 mRNA. Molecular mechanisms leading to fragile X-premutation-associated conditions (FXPAC) include cotranscriptional R-loop formations, FMR1 mRNA toxicity through both RNA gelation into nuclear foci and sequestration of various CGG-repeat-binding proteins, and the repeat-associated non-AUG (RAN)-initiated translation of potentially toxic proteins. Such molecular mechanisms contribute to subsequent consequences, including mitochondrial dysfunction and neuronal death. Clinically, premutation carriers may exhibit a wide range of symptoms and phenotypes. Any of the problems associated with the premutation can appropriately be called FXPAC. Fragile X-associated tremor/ataxia syndrome (FXTAS), fragile X-associated primary ovarian insufficiency (FXPOI), and fragile X-associated neuropsychiatric disorders (FXAND) can fall under FXPAC. Understanding the molecular and clinical aspects of the premutation of the FMR1 gene is crucial for the accurate diagnosis, genetic counseling, and appropriate management of affected individuals and families. This paper summarizes all the known problems associated with the premutation and documents the presentations and discussions that occurred at the International Premutation Conference, which took place in New Zealand in 2023.
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Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Síndrome del Cromosoma X Frágil , Humanos , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Mutación/genética , ARN Mensajero/metabolismo , Expansión de Repetición de Trinucleótido/genética , Síndrome del Cromosoma X Frágil/diagnóstico , Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/terapiaRESUMEN
A GGGGCC (G4C2) hexanucleotide repeat expansion in C9ORF72 causes amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD), while a CGG trinucleotide repeat expansion in FMR1 leads to the neurodegenerative disorder Fragile X-associated tremor/ataxia syndrome (FXTAS). These GC-rich repeats form RNA secondary structures that support repeat-associated non-AUG (RAN) translation of toxic proteins that contribute to disease pathogenesis. Here we assessed whether these same repeats might trigger stalling and interfere with translational elongation. We find that depletion of ribosome-associated quality control (RQC) factors NEMF, LTN1, and ANKZF1 markedly boost RAN translation product accumulation from both G4C2 and CGG repeats while overexpression of these factors reduces RAN production in both reporter cell lines and C9ALS/FTD patient iPSC-derived neurons. We also detected partially made products from both G4C2 and CGG repeats whose abundance increased with RQC factor depletion. Repeat RNA sequence, rather than amino acid content, is central to the impact of RQC factor depletion on RAN translation - suggesting a role for RNA secondary structure in these processes. Together, these findings suggest that ribosomal stalling and RQC pathway activation during RAN translation elongation inhibits the generation of toxic RAN products. We propose augmenting RQC activity as a therapeutic strategy in GC-rich repeat expansion disorders.
RESUMEN
Neurodegeneration in Fragile X-associated tremor/ataxia syndrome (FXTAS) is caused by a CGG trinucleotide repeat expansion in the 5' UTR of FMR1. Expanded CGG repeat RNAs form stable secondary structures, which in turn support repeat-associated non-AUG (RAN) translation to produce toxic peptides. The parameters that impact RAN translation initiation efficiency are not well understood. Here we used a Drosophila melanogaster model of FXTAS to evaluate the role of the eIF4G family of eukaryotic translation initiation factors (EIF4G1, EIF4GII and EIF4G2/DAP5) in modulating RAN translation and CGG repeat-associated toxicity. DAP5 knockdown robustly suppressed CGG repeat-associated toxicity and inhibited RAN translation. Furthermore, knockdown of initiation factors that preferentially associate with DAP5 (such as EIF2ß, EIF3F and EIF3G) also selectively suppressed CGG repeat-induced eye degeneration. In mammalian cellular reporter assays, DAP5 knockdown exhibited modest and cell-type specific effects on RAN translation. Taken together, these data support a role for DAP5 in CGG repeat associated toxicity possibly through modulation of RAN translation.
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Proteínas de Drosophila , Síndrome del Cromosoma X Frágil , Animales , Drosophila/metabolismo , Temblor/genética , Drosophila melanogaster/metabolismo , Factor 4G Eucariótico de Iniciación/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/genética , Expansión de Repetición de Trinucleótido , Ataxia/genética , Mamíferos/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismoRESUMEN
Over a third of the human genome is comprised of repetitive sequences, including more than a million short tandem repeats (STRs). While studies of the pathologic consequences of repeat expansions that cause syndromic human diseases are extensive, the potential native functions of STRs are often ignored. Here, we summarize a growing body of research into the normal biological functions for repetitive elements across the genome, with a particular focus on the roles of STRs in regulating gene expression. We propose reconceptualizing the pathogenic consequences of repeat expansions as aberrancies in normal gene regulation. From this altered viewpoint, we predict that future work will reveal broader roles for STRs in neuronal function and as risk alleles for more common human neurological diseases.
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Genoma Humano , Repeticiones de Microsatélite , Humanos , Repeticiones de Microsatélite/genética , Regulación de la Expresión GénicaRESUMEN
Pathogenic repeat sequences underlie several human disorders, including amyotrophic lateral sclerosis, Huntington's disease, and myotonic dystrophy. Here, we speak to several researchers about how repeat sequences have been implicated in affecting all aspects of the Central Dogma of molecular biology through their effects on DNA, RNA, and protein.
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Esclerosis Amiotrófica Lateral , Enfermedad de Huntington , Distrofia Miotónica , Humanos , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , Proteínas/genética , Enfermedad de Huntington/genética , ARN/genética , Distrofia Miotónica/genética , Expansión de Repetición de Trinucleótido/genéticaRESUMEN
Cerebellar ataxia with neuropathy and vestibular areflexia syndrome (CANVAS) is a late onset, recessively inherited neurodegenerative disorder caused by biallelic, non-reference pentameric AAGGG(CCCTT) repeat expansions within the second intron of replication factor complex subunit 1 (RFC1). To investigate how these repeats cause disease, we generated CANVAS patient induced pluripotent stem cell (iPSC) derived neurons (iNeurons) and utilized calcium imaging and transcriptomic analysis to define repeat-elicited gain-of-function and loss-of-function contributions to neuronal toxicity. AAGGG repeat expansions do not alter neuronal RFC1 splicing, expression, or DNA repair pathway functions. In reporter assays, AAGGG repeats are translated into pentapeptide repeat proteins that selectively accumulate in CANVAS patient brains. However, neither these proteins nor repeat RNA foci were detected in iNeurons, and overexpression of these repeats in isolation did not induce neuronal toxicity. CANVAS iNeurons exhibit defects in neuronal development and diminished synaptic connectivity that is rescued by CRISPR deletion of a single expanded allele. These phenotypic deficits were not replicated by knockdown of RFC1 in control neurons and were not rescued by ectopic expression of RFC1. These findings support a repeat-dependent but RFC1-independent cause of neuronal dysfunction in CANVAS, with important implications for therapeutic development in this currently untreatable condition.
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Background: The original Fragile X-associated Tremor Ataxia Syndrome Rating Scale (FXTAS-RS) contained 61 items, some requiring modifications to better meet recommendations for patient-focused rating scale development. Purpose: Provide initial validation of a revised version of the FXTAS-RS for motor signs. Method: We conducted a two-phase mixed-method approach. In Phase 1, revision, we implemented a Delphi technique identifying pertinent domains/subdomains and developing items through expert consensus. In Phase 2, content validation, we conducted cognitive pretesting assessing comprehensibility, comprehensiveness, and relevance of items to FXTAS motor signs. Results: After five rounds of Delphi panel and two rounds of cognitive pretesting, the revised version of the FXTAS-RS was established with 18 items covering five domains and 13 subdomains of motor signs. Cognitive pretesting revealed adequate content validity for the assessment of FXTAS motor signs. Conclusion: The revised FXTAS-RS has been successfully validated for content and it is now ready for large-scale field validation.
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BACKGROUND: X-linked dystonia-parkinsonism (XDP) is a neurodegenerative disorder caused by the intronic insertion of a SINE-VNTR-Alu (SVA) retrotransposon carrying an (AGAGGG)n repeat expansion in the TAF1 gene. The molecular mechanisms by which this mutation causes neurodegeneration remain elusive. OBJECTIVES: We investigated whether (AGAGGG)n repeats undergo repeat-associated non-AUG (RAN) translation, a pathogenic mechanism common among repeat expansion diseases. METHODS: XDP-specific RAN translation reporter plasmids were generated, transfected in HEK293 cells, and putative dipeptide repeat proteins (DPRs) were detected by Western blotting. Immunocytochemistry was performed in COS-7 cells to determine the subcellular localization of one DPR. RESULTS: We detected putative DPRs from two reading frames, supporting the translation of poly-(Glu-Gly) and poly-(Arg-Glu) species. XDP RAN translation initiates within the (AGAGGG)n sequence and poly-(Glu-Gly) DPRs formed nuclear inclusions in transfected cells. CONCLUSIONS: In summary, our work provides the first in-vitro proof of principle that the XDP-linked (AGAGGG)n repeat expansions can undergo RAN translation. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.
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Trastornos Distónicos , Enfermedades Genéticas Ligadas al Cromosoma X , Humanos , Células HEK293 , Trastornos Distónicos/metabolismo , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Intrones , Proteína C9orf72/genéticaRESUMEN
CGG repeat expansions in the FMR1 5'UTR cause the neurodegenerative disease Fragile X-associated tremor/ataxia syndrome (FXTAS). These repeats form stable RNA secondary structures that support aberrant translation in the absence of an AUG start codon (RAN translation), producing aggregate-prone peptides that accumulate within intranuclear neuronal inclusions and contribute to neurotoxicity. Here, we show that the most abundant RAN translation product, FMRpolyG, is markedly less toxic when generated from a construct with a non-repetitive alternating codon sequence in place of the CGG repeat. While exploring the mechanism of this differential toxicity, we observed a +1 translational frameshift within the CGG repeat from the arginine to glycine reading frame. Frameshifts occurred within the first few translated repeats and were triggered predominantly by RNA sequence and structural features. Short chimeric R/G peptides form aggregates distinct from those formed by either pure arginine or glycine, and these chimeras induce toxicity in cultured rodent neurons. Together, this work suggests that CGG repeats support translational frameshifting and that chimeric RAN translated peptides may contribute to CGG repeat-associated toxicity in FXTAS and related disorders.
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Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Enfermedades Neurodegenerativas , Agregación Patológica de Proteínas , Repeticiones de Trinucleótidos , Arginina/genética , Ataxia , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil , Glicina/genética , Humanos , Enfermedades Neurodegenerativas/genética , Péptidos/genética , Péptidos/metabolismo , Proteínas Recombinantes de Fusión/metabolismoRESUMEN
FMR1 premutation cytosine-guanine-guanine repeat expansion alleles are relatively common mutations in the general population that are associated with a neurodegenerative disease (fragile X-associated tremor/ataxia syndrome), reproductive health problems and potentially a wide range of additional mental and general health conditions that are not yet well-characterised. The International Fragile X Premutation Registry (IFXPR) was developed to facilitate and encourage research to better understand the FMR1 premutation and its impact on human health, to facilitate clinical trial readiness by identifying and characterising diverse cohorts of individuals interested in study participation, and to build community and collaboration among carriers, family members, researchers and clinicians around the world. Here, we describe the development and content of the IFXPR, characterise its first 747 registrants from 32 countries and invite investigators to apply for recruitment support for their project(s). With larger numbers, increased diversity and potentially the future clinical characterisation of registrants, the IFXPR will contribute to a more comprehensive and accurate understanding of the fragile X premutation in human health and support treatment studies.
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Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Enfermedades Neurodegenerativas , Humanos , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Expansión de Repetición de Trinucleótido/genética , Enfermedades Neurodegenerativas/genética , Sistema de Registros , GuaninaRESUMEN
Fragile Xassociated tremor/ataxia syndrome (FXTAS) is a debilitating late-onset neurodegenerative disease in premutation carriers of the expanded CGG repeat in FMR1 that presents with a spectrum of neurological manifestations, such as gait ataxia, intention tremor, and parkinsonism [P. J. Hagerman, R. J. Hagerman, Ann. N. Y. Acad. Sci. 1338, 5870 (2015); S. Jacquemont et al., JAMA 291, 460469 (2004)]. Here, we performed whole-genome sequencing (WGS) on male premutation carriers (CGG55200) and prioritized candidate variants to screen for candidate genetic modifiers using a Drosophila model of FXTAS. We found 18 genes that genetically modulate CGG-associated neurotoxicity in Drosophila, such as Prosbeta5 (PSMB5), pAbp (PABPC1L), e(y)1 (TAF9), and CG14231 (OSGEPL1). Among them, knockdown of Prosbeta5 (PSMB5) suppressed CGG-associated neurodegeneration in the fly as well as in N2A cells. Interestingly, an expression quantitative trait locus variant in PSMB5, PSMB5rs11543947-A, was found to be associated with decreased expression of PSMB5 and delayed onset of FXTAS in human FMR1 premutation carriers. Finally, we demonstrate evidence that PSMB5 knockdown results in suppression of CGG neurotoxicity via both the RAN translation and RNA-mediated toxicity mechanisms, thereby presenting a therapeutic strategy for FXTAS.
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Ataxia , Síndrome del Cromosoma X Frágil , Complejo de la Endopetidasa Proteasomal , Temblor , Animales , Ataxia/genética , Modelos Animales de Enfermedad , Drosophila melanogaster , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/genética , Humanos , Masculino , Complejo de la Endopetidasa Proteasomal/genética , Temblor/genéticaRESUMEN
Repeat-associated non-AUG (RAN) translation of expanded repeat-mutation mRNA produces toxic peptides in neurons of patients suffering from neurodegenerative diseases. Recent findings indicate that RAN translation in diverse model systems is not inhibited by cellular stressors that impair global translation through phosphorylation of the alpha subunit of eIF2, the essential eukaryotic translation initiation factor that brings the initiator tRNA to the 40S ribosome. Using in vitro, cell-based and Drosophila models, we examined the role of alternative ternary complex factors that may function in place of eIF2, including eIF2A, eIF2D, DENR and MCTS1. Among these factors, DENR knockdown had the greatest inhibitory effect on RAN translation of expanded GGGGCC and CGG repeat reporters and its reduction improved the survival of Drosophila expressing expanded GGGGCC repeats. Taken together, these data support a role for alternative initiation factors in RAN translation and suggest these may serve as novel therapeutic targets in neurodegenerative disease.
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Proteínas de Drosophila , Enfermedades Neurodegenerativas , Animales , Drosophila/genética , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Factor 2 Eucariótico de Iniciación/genética , Factor 2 Eucariótico de Iniciación/metabolismo , Factores Eucarióticos de Iniciación/genética , Factores Eucarióticos de Iniciación/metabolismo , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/metabolismo , Biosíntesis de Proteínas/genética , ARN Mensajero/genética , Ribosomas/genética , Ribosomas/metabolismoRESUMEN
Repeat associated non-AUG (RAN) translation of CGG repeats in the 5'UTR of FMR1 produces toxic proteins that contribute to fragile X-associated tremor/ataxia syndrome (FXTAS) pathogenesis. The most abundant RAN product, FMRpolyG, initiates predominantly at an ACG upstream of the repeat. Accurate FMRpolyG measurements in FXTAS patients are lacking. We used data-dependent acquisition and parallel reaction monitoring (PRM) mass spectrometry coupled with stable isotope labeled standard peptides to identify signature FMRpolyG fragments in patient samples. Following immunoprecipitation, PRM detected FMRpolyG signature peptides in transfected cells, and FXTAS tissues and cells, but not in controls. We identified two amino-terminal peptides: an ACG-initiated Ac-MEAPLPGGVR and a GUG-initiated Ac-TEAPLPGGVR, as well as evidence for RAN translation initiation within the CGG repeat itself in two reading frames. Initiation at all sites increased following cellular stress, decreased following eIF1 overexpression and was eIF4A and M7G cap-dependent. These data demonstrate that FMRpolyG is quantifiable in human samples and FMR1 RAN translation initiates via similar mechanisms for near-cognate codons and within the repeat through processes dependent on available initiation factors and cellular environment.
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Ataxia , Síndrome del Cromosoma X Frágil , Temblor , Proteína de Unión al GTP ran , Ataxia/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/genética , Humanos , Péptidos/metabolismo , Temblor/genética , Expansión de Repetición de Trinucleótido , Proteína de Unión al GTP ran/genéticaRESUMEN
Microbial dysbiosis is a colorectal cancer (CRC) hallmark and contributes to inflammation, tumor growth, and therapy response. Gut microbes signal via metabolites, but how the metabolites impact CRC is largely unknown. We interrogated fecal metabolites associated with mouse models of colon tumorigenesis with varying mutational load. We find that microbial metabolites from healthy mice or humans are growth-repressive, and this response is attenuated in mice and patients with CRC. Microbial profiling reveals that Lactobacillus reuteri and its metabolite, reuterin, are downregulated in mouse and human CRC. Reuterin alters redox balance, and reduces proliferation and survival in colon cancer cells. Reuterin induces selective protein oxidation and inhibits ribosomal biogenesis and protein translation. Exogenous Lactobacillus reuteri restricts colon tumor growth, increases tumor reactive oxygen species, and decreases protein translation in vivo. Our findings indicate that a healthy microbiome and specifically, Lactobacillus reuteri, is protective against CRC through microbial metabolite exchange.
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Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Microbioma Gastrointestinal , Gliceraldehído/análogos & derivados , Oxidación-Reducción , Propano/metabolismo , Animales , Biomarcadores , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Metabolismo Energético , Glutatión/metabolismo , Gliceraldehído/metabolismo , Gliceraldehído/farmacología , Interacciones Microbiota-Huesped , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiología , Mucosa Intestinal/patología , Metabolómica/métodos , Metagenómica/métodos , Ratones , Modelos Biológicos , Oxidación-Reducción/efectos de los fármacos , Estrés Oxidativo , Propano/farmacología , Transducción de Señal , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Transcribed CGG repeat expansions cause neurodegeneration in Fragile X-associated tremor/ataxia syndrome (FXTAS). CGG repeat RNAs sequester RNA-binding proteins (RBPs) into nuclear foci and undergo repeat-associated non-AUG (RAN) translation into toxic peptides. To identify proteins involved in these processes, we employed a CGG repeat RNA-tagging system to capture repeat-associated RBPs by mass spectrometry in mammalian cells. We identified several SR (serine/arginine-rich) proteins that interact selectively with CGG repeats basally and under cellular stress. These proteins modify toxicity in a Drosophila model of FXTAS. Pharmacologic inhibition of serine/arginine protein kinases (SRPKs), which alter SRSF protein phosphorylation, localization, and activity, directly inhibits RAN translation of CGG and GGGGCC repeats (associated with C9orf72 ALS/FTD) and triggers repeat RNA retention in the nucleus. Lowering SRPK expression suppressed toxicity in both FXTAS and C9orf72 ALS/FTD model flies, and SRPK inhibitors suppressed CGG repeat toxicity in rodent neurons. Together, these findings demonstrate roles for CGG repeat RNA binding proteins in RAN translation and repeat toxicity and support further evaluation of SRPK inhibitors in modulating RAN translation associated with repeat expansion disorders.