Defects In RNA Metabolism Research Articles

5-Fluorouracil treatment represses pseudouridine-containing miRNA export into extracellular vesicles.

5-Fluorouracil (5-FU) has been used for chemotherapy for colorectal and other cancers for over 50 years. The prevailing view of its mechanism of action is inhibition of thymidine synthase leading to defects in DNA replication and repair. However, 5-FU is also incorporated into RNA causing defects in RNA metabolism, inhibition of pseudouridine modification, and altered ribosome function. We examined the impact of 5-FU on post-transcriptional small RNA modifications (PTxMs) and the expression and export of RNA into small extracellular vesicles (sEVs). EVs are secreted by all cells and contain a variety of proteins and RNAs that can function in cell-cell communication. We found that treatment of colorectal cancer (CRC) cells with 5-FU represses sEV export of miRNA and snRNA-derived RNAs, but promotes export of snoRNA-derived RNAs. Strikingly, 5-FU treatment significantly decreased the levels of pseudouridine on both cellular and sEV small RNA profiles. In contrast, 5-FU exposure led to increased levels of cellular small RNAs containing a variety of methyl-modified bases. These unexpected findings show that 5-FU exposure leads to altered RNA expression, base modification, and aberrant trafficking and localization of small RNAs.

Pathological phase transitions in ALS-FTD impair dynamic RNA-protein granules.

The genetics of human disease serves as a robust and unbiased source of insight into human biology, both revealing fundamental cellular processes and exposing the vulnerabilities associated with their dysfunction. Over the last decade, the genetics of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) have epitomized this concept, as studies of ALS-FTD-causing mutations have yielded fundamental discoveries regarding the role of biomolecular condensation in organizing cellular contents while implicating disturbances in condensate dynamics as central drivers of neurodegeneration. Here we review this genetic evidence, highlight its intersection with patient pathology, and discuss how studies in model systems have revealed a role for aberrant condensation in neuronal dysfunction and death. We detail how multiple, distinct types of disease-causing mutations promote pathological phase transitions that disturb the dynamics and function of ribonucleoprotein (RNP) granules. Dysfunction of RNP granules causes pleiotropic defects in RNA metabolism and can drive the evolution of these structures to end-stage pathological inclusions characteristic of ALS-FTD. We propose that aberrant phase transitions of these complex condensates in cells provide a parsimonious explanation for the widespread cellular abnormalities observed in ALS as well as certain histopathological features that characterize late-stage disease.

RNA Is a Double-Edged Sword in ALS Pathogenesis.

Amyotrophic lateral sclerosis (ALS) is a progressive and fatal neurodegenerative disease that affects upper and lower motor neurons. Familial ALS accounts for a small subset of cases (<10–15%) and is caused by dominant mutations in one of more than 10 known genes. Multiple genes have been causally or pathologically linked to both ALS and frontotemporal dementia (FTD). Many of these genes encode RNA-binding proteins, so the role of dysregulated RNA metabolism in neurodegeneration is being actively investigated. In addition to defects in RNA metabolism, recent studies provide emerging evidence into how RNA itself can contribute to the degeneration of both motor and cortical neurons. In this review, we discuss the roles of altered RNA metabolism and RNA-mediated toxicity in the context of TARDBP, FUS, and C9ORF72 mutations. Specifically, we focus on recent studies that describe toxic RNA as the potential initiator of disease, disease-associated defects in specific RNA metabolism pathways, as well as how RNA-based approaches can be used as potential therapies. Altogether, we highlight the importance of RNA-based investigations into the molecular progression of ALS, as well as the need for RNA-dependent structural studies of disease-linked RNA-binding proteins to identify clear therapeutic targets.

Stress Granule Dysregulation in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease with no current cure. ALS causes degeneration of both upper and lower motor neurons leading to atrophy of the innervating muscles and progressive paralysis. The exact mechanism of the pathology of ALS is unknown. However, 147 genes have been identified that are causative, associated with, or modify disease progression. While the causative mechanism is unknown, a number of pathological processes have been associated with ALS. These include mitochondrial dysfunction, protein accumulation, and defects in RNA metabolism. RNA metabolism is a complicated process that is regulated by many different RNA-binding proteins (RBPs). A small defect in RNA metabolism can produce results as dramatic as determining cell survival. Stress granules (SGs) control RNA translation during stressed conditions. This is a protective reaction, but in conditions of chronic stress can become pathogenic. SGs are even hypothesized to act as a seeding mechanism for the pathological aggregation of proteins seen in many neurodegenerative diseases, including TAR DNA-binding protein 43 (TDP-43) in ALS. In this review, we will be summarizing the current findings of SG pathology in ALS. We also focus on the role of SG dysregulation in protein aggregate formation and mitochondrial dysfunction. In addition, we outline therapeutic strategies that target SG components in ALS.

USP15 Deubiquitinates TUT1 Associated with RNA Metabolism and Maintains Cerebellar Homeostasis.

Precise regulation of RNA metabolism is crucial for dynamic gene expression and controlling cellular functions. In the nervous system, defects in RNA metabolism are implicated in the disturbance of brain homeostasis and development. Here, we report that deubiquitinating enzyme, ubiquitin specific peptidase 15 (USP15), deubiquitinates terminal uridylyl transferase 1 (TUT1) and changes global RNA metabolism. We found that the expression of USP15 redistributes TUT1 from the nucleolus to nucleoplasm, resulting in the stabilization of U6 snRNA. We also found that lack of the Usp15 gene induces an impairment in motor ability with an unconventional cerebellar formation. Moreover, inhibition of the USP15-TUT1 cascade triggered mild and chronic endoplasmic reticulum (ER) stress. Therefore, our results suggest that USP15 is crucial for mRNA metabolism and maintains a healthy brain. These findings provide a possibility that disturbance of the USP15-TUT1 cascade induces chronic and mild ER stress, leading to an acceleration of the neurodegenerative phenotype.

New immunodeficiency syndromes that help us understand the IFN-mediated antiviral immune response.

Studying primary immunodeficiencies (PIDs) provides insights into human antiviral immunity in the natural infectious environment. This review describes new PIDs with genetic defects that impair innate antiviral responses. New genetic defects in the interferon (IFN) signaling pathway include IFNAR1 deficiency, which causes uncontrolled infections with measles-mumps-rubella or yellow fever vaccines, and possibly also cytomegalovirus (CMV); and IRF9 deficiency, which results in influenza virus susceptibility. Genetic defects in several pattern recognition receptors include MDA5 deficiency, which impairs viral RNA sensing and confers human rhinovirus susceptibility; RNA polymerase III haploinsufficiency, which impairs sensing of A:T-rich virus DNA and confers VZV susceptibility; and TLR3 deficiency, which causes HSV-1 encephalitis (HSE) or influenza virus pneumonitis. Defects in RNA metabolism, such as that caused by Debranching enzyme 1 deficiency, can cause virus meningoencephalitis. Finally, defects in host restriction factors for virus replication, such as in CIB1 deficiency, contribute to uncontrolled β-HPV infections. Several new PIDs highlight the role of type I/III IFN signaling pathway, virus sensors, and host virus restriction factors in human antiviral immunity.

F47. Screening of GLE1 mutations in Chinese amyotrophic lateral sclerosis patients

Introduction Amyotrophic lateral sclerosis (ALS) is a fatal disease involving both upper and lower motor neuron, resulting in a 3–5 year of survival from symptom onset caused by rapidly progressive paralysis and respiratory failure. It is etiologically heterogeneous. Approximately 90% of ALS cases are sporadic (SALS) with unknown causes. But The remaining 10% of patients have a family history (FALS) and harbor a causative gene mutation, reflecting genetic factor play an important role in the pathogenesis of ALS. To date, mutations in over 20 genes have been reported in ALS populations. Mutations in GLE1 are the causation of two fetal motoneuron diseases: lethal congenital contracture syndrome 1 (LCCS1) and lethal arthrogryposis with anterior horn cell disease (LAAHD). Recently, Kaneb et al. identifiedGLE1variants within sporadic and familial Caucasian ALS cases, supporting defects in RNA metabolism were associated with pathogenesis of ALS. However, GLE1 mutations have not been investigated in Chinese ALS patients so far. Accordingly, we evaluated whether GLE1 mutations harbored in ALS patients from mainland China. Methods The current study recruited probands of 20 ALS families (mean age of onset was 50.1 ± 10.9 years; 10 females, 10 males), 230 SALS patients (mean age of onset was 52.1 ± 11.9 years; 96 females, 134 males). All 16 exons (plus 50 bp of flanking introns) of GLE1 (NM_001003722.1) including the entire coding region were amplified using polymerase chain reaction (PCR). The reaction conditions of PCR were as follows: 5 min of denaturation at 95 °C, 8 cycles were used with 20 s at 94 °C, 20 s at 65 °C (decreasing by 1 °C with each cycle) and 30 s at 72 °C. 20 s at 94 °C, 20 s at 58 °C and 30 s at 72 °C for the subsequent 34 cycles, a final extension step at 72 °C for 5 min. PCR products were subjected to Sanger sequencing using ABI 3730 automated DNA-sequencing system (Applied Biosystems). The sequences were aligned with reference sequence using CodonCode Aligner. Results GLE1 mutation screening of 250 ALS cases revealed 4 synonymous genetic variants(c.444G > A, p. R148R; c.946C > A, p. R316R; c.1641T > C, p. Y547Y; c.2082C > T, P. S694S), which presented in dbSNP database. Bioinformatic analysis using ESE finder and Human Splicing Finder 3.0 showed that all the silent mutations did not affect transcription process. We did not identify any novel or reported rare variants in all 16 coding exons of GLE1from 250 Chinese ALS patients. Conclusion In summary, our results suggested that GLE1 mutations are not common in FALS and SALS of Chinese origin. Further study in larger and additional populations would be required to evaluate the role of GLE1 in ALS causation.

Failure to Deliver and Translate-New Insights into RNA Dysregulation in ALS.

Amyotrophic Lateral Sclerosis (ALS) is a progressive and fatal neurodegenerative disease affecting both upper and lower motor neurons. The molecular mechanisms underlying disease pathogenesis remain largely unknown. Multiple genetic loci including genes involved in proteostasis and ribostasis have been linked to ALS providing key insights into the molecular mechanisms underlying disease. In particular, the identification of the RNA binding proteins TDP-43 and fused in sarcoma (FUS) as causative factors of ALS resulted in a paradigm shift centered on the study of RNA dysregulation as a major mechanism of disease. With wild-type TDP-43 pathology being found in ~97% of ALS cases and the identification of disease causing mutations within its sequence, TDP-43 has emerged as a prominent player in ALS. More recently, studies of the newly discovered C9orf72 repeat expansion are lending further support to the notion of defects in RNA metabolism as a key factor underlying ALS. RNA binding proteins are involved in all aspects of RNA metabolism ranging from splicing, transcription, transport, storage into RNA/protein granules, and translation. How these processes are affected by disease-associated mutations is just beginning to be understood. Considerable work has gone into the identification of splicing and transcription defects resulting from mutations in RNA binding proteins associated with disease. More recently, defects in RNA transport and translation have been shown to be involved in the pathomechanism of ALS. A central hypothesis in the field is that disease causing mutations lead to the persistence of RNA/protein complexes known as stress granules. Under times of prolonged cellular stress these granules sequester specific mRNAs preventing them from translation, and are thought to evolve into pathological aggregates. Here we will review recent efforts directed at understanding how altered RNA metabolism contributes to ALS pathogenesis.

Defects in RNA metabolism in mitochondrial disease

The expression of mitochondrially-encoded genes requires the efficient processing of long precursor RNAs at the 5′ and 3′ ends of tRNAs, a process which, when disrupted, results in disease. Two such mutations reside within mt-tRNALeu(UUR); a m.3243A>G transition, which is the most common cause of MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes), and m.3302A>G which often causes mitochondrial myopathy (MM). We used parallel analysis of RNA ends (PARE) that captures the 5′ terminal end of 5′-monophosphorylated mitochondrial RNAs to compare the effects of the m.3243A>G and m.3302A>G mutations on mitochondrial tRNA processing and downstream RNA metabolism. We confirmed previously identified RNA processing defects, identified common internal cleavage sites and new sites unique to the m.3243A>G mutants that do not correspond to transcript ends. These sites occur in regions of predicted RNA secondary structure, or are in close proximity to such regions, and may identify regions of importance to the processing of mtRNAs.

An amyotrophic lateral sclerosis-linked mutation in GLE1 alters the cellular pool of human Gle1 functional isoforms.

Amyotrophic lateral sclerosis (ALS) is a lethal late onset motor neuron disease with underlying cellular defects in RNA metabolism. In prior studies, two deleterious heterozygous mutations in the gene encoding human (h)Gle1 were identified in ALS patients. hGle1 is an mRNA processing modulator that requires inositol hexakisphosphate (IP6) binding for function. Interestingly, one hGLE1 mutation (c.1965-2A>C) results in a novel 88 amino acid C-terminal insertion, generating an altered protein. Like hGle1A, at steady state, the altered protein termed hGle1-IVS14-2A>C is absent from the nuclear envelope rim and localizes to the cytoplasm. hGle1A performs essential cytoplasmic functions in translation and stress granule regulation. Therefore, we speculated that the ALS disease pathology results from altered cellular pools of hGle1 and increased cytoplasmic hGle1 activity. GFP-hGle1-IVS14-2A>C localized to stress granules comparably to GFP-hGle1A, and rescued stress granule defects following siRNA-mediated hGle1 depletion. As described for hGle1A, overexpression of the hGle1-IVS14-2A>C protein also induced formation of larger SGs. Interestingly, hGle1A and the disease associated hGle1-IVS14-2A>C overexpression induced the formation of distinct cytoplasmic protein aggregates that appear similar to those found in neurodegenerative diseases. Strikingly, the ALS-linked hGle1-IVS14-2A>C protein also rescued mRNA export defects upon depletion of endogenous hGle1, acting in a potentially novel bi-functional manner. We conclude that the ALS-linked hGle1-c.1965-2A>C mutation generates a protein isoform capable of both hGle1A- and hGle1B-ascribed functions, and thereby uncoupled from normal mechanisms of hGle1 regulation.

The Genetics of Spinal Muscular Atrophy: Progress and Challenges.

Spinal muscular atrophies (SMAs) are a group of inherited disorders characterized by motor neuron loss in the spinal cord and lower brainstem, muscle weakness, and atrophy. The clinical and genetic phenotypes incorporate a wide spectrum that is differentiated based on age of onset, pattern of muscle involvement, and inheritance pattern. Over the past several years, rapid advances in genetic technology have accelerated the identification of causative genes and provided important advances in understanding the molecular and biological basis of SMA and insights into the selective vulnerability of the motor neuron. Common pathophysiological themes include defects in RNA metabolism and splicing, axonal transport, and motor neuron development and connectivity. Together these have revealed potential novel treatment strategies, and extensive efforts are being undertaken towards expedited therapeutics. While a number of promising therapies for SMA are emerging, defining therapeutic windows and developing sensitive and relevant biomarkers are critical to facilitate potential success in clinical trials. This review incorporates an overview of the clinical manifestations and genetics of SMA, and describes recent advances in the understanding of mechanisms of disease pathogenesis and development of novel treatment strategies.

Deleterious mutations in the essential mRNA metabolism factor, hGle1, in amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the selective death of motor neurons. Causative mutations in the global RNA-processing proteins TDP-43 and FUS among others, as well as their aggregation in ALS patients, have identified defects in RNA metabolism as an important feature in this disease. Lethal congenital contracture syndrome 1 and lethal arthrogryposis with anterior horn cell disease are autosomal recessive fetal motor neuron diseases that are caused by mutations in another global RNA-processing protein, hGle1. In this study, we carried out the first screening of GLE1 in ALS patients (173 familial and 760 sporadic) and identified 2 deleterious mutations (1 splice site and 1 nonsense mutation) and 1 missense mutation. Functional analysis of the deleterious mutants revealed them to be unable to rescue motor neuron pathology in zebrafish morphants lacking Gle1. Furthermore, in HeLa cells, both mutations caused a depletion of hGle1 at the nuclear pore where it carries out an essential role in nuclear export of mRNA. These results suggest a haploinsufficiency mechanism and point to a causative role for GLE1 mutations in ALS patients. This further supports the involvement of global defects in RNA metabolism in ALS.

Defects in RNA Metabolism links FTD and ALS Pathogenesis: TDP-43, FUS, and C9orf72

Increasing evidence supports the idea that frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) share underlying causes and lie on opposite ends of a disease spectrum that leads to these disorders. The discovery of two different RNA binding proteins, TDP-43 and FUS/TLS, as common neuronal inclusions in FTD and ALS patients has strengthened this connection. Subsequent identification of mutations in TDP-43 and FUS that cause ALS, and FTD in rare cases, provides additional linkage between these diseases. Furthermore, autosomal dominant forms of FTD, ALS, or a combined phenotype can occur in the same family. Genetic studies have linked some of these cases to chromosome 9p21 and are known as c9FTD/ALS. Recently, an expanded GGGGCC hexanucleotide repeat in the C9ORF72 gene has been identified in c9FTD/ALS cases as well as sporadic forms of both diseases. Preliminary data suggest that the increased nucleotide repeats in C9ORF72 lead to RNA inclusions or foci in the nucleus of affected cells. Analogous to other repeat disorders such as myotonic dystrophy, expanded repeats in C9ORF72 may cause disease pathogenesis by sequestering RNA binding proteins and/or perturbing the splicing and regulation of key proteins regulating neuronal health and survival. Intriguingly, TDP-43 and FUS/TLS play fundamental roles in RNA regulation and splicing, and their mutation leads to dysfunctional regulation of their targets in model systems. Finally, defects in RNA splicing have been reported in sporadic ALS and a number of other genes involved in RNA metabolism (ANG, SETX, TAF15, ELP3, ATXN2) are associated with ALS. Based on these recent findings, we propose that defects in RNA metabolism are a common pathway linking FTD and ALS, and are responsible for disease pathogenesis in a significant portion of cases. Further research to test this hypothesis and determine if these proteins function within common biological pathways and share similar pathogenic mechanisms will ultimately open up new routes of therapy for these devastating disorders. Keywords: AD, ALS, FTD, FTLD, FUS, TLS, C9ORF72, PD, TDP-43, proteinopathies, neurodegeneration, inclusion.

Changes in intra-nuclear mobility of mature snRNPs provide a mechanism for splicing defects in Spinal Muscular Atrophy.

It is becoming increasingly clear that defects in RNA metabolism can lead to disease. Spinal muscular atrophy (SMA), a leading genetic cause of infant mortality, results from insufficient amounts of survival motor neuron (SMN) protein. SMN is required for the biogenesis of small nuclear ribonucleoproteins (snRNPs): essential components of the spliceosome. Splicing abnormalities have been detected in models of SMA but it is unclear how lowered SMN affects the fidelity of pre-mRNA splicing. We have examined the dynamics of mature snRNPs in cells depleted of SMN and demonstrated that SMN depletion increases the mobility of mature snRNPs within the nucleus. To dissect the molecular mechanism by which SMN deficiency affects intranuclear snRNP mobility, we employed a panel of inhibitors of different stages of pre-mRNA processing. This in vivo modelling demonstrates that snRNP mobility is altered directly as a result of impaired snRNP maturation. Current models of nuclear dynamics predict that subnuclear structures, including the spliceosome, form by self-organization mediated by stochastic interactions between their molecular components. Thus, alteration of the intranuclear mobility of snRNPs provides a molecular mechanism for splicing defects in SMA.

Expression profiling of FSHD-1 and FSHD-2 cells during myogenic differentiation evidences common and distinctive gene dysregulation patterns.

BackgroundDetermine global gene dysregulation affecting 4q-linked (FSHD-1) and non 4q-linked (FSHD-2) cells during early stages of myogenic differentiation. This approach has been never applied to FSHD pathogenesis.Methodology/Principal FindingsBy in vitro differentiation of FSHD-1 and FSHD-2 myoblasts and gene chip analysis we derived that gene expression profile is altered only in FSHD-1 myoblasts and FSHD-2 myotubes. The changes seen in FSHD-1 regarded a general defect in cell cycle progression, probably due to the upregulation of myogenic markers PAX3 and MYOD1, and a deficit of factors (SUV39H1 and HMGB2) involved in D4Z4 chromatin conformation. On the other hand, FSHD-2 mytubes were characterized by a general defect in RNA metabolism, protein synthesis and degradation and, to a lesser extent, in cell cycle. Common dysregulations regarded genes involved in response to oxidative stress and in sterol biosynthetic process. Interestingly, our results also suggest that miRNAs might be implied in both FSHD-1 and FSHD-2 gene dysregulation. Finally, in both cell differentiation systems, we did not observe a gradient of altered gene expression throughout the 4q35 chromosome.Conclusions/SignificanceFSHD-1 and FSHD-2 cells showed, in different steps of myogenic differentiation, a global deregulation of gene expression rather than an alteration of expression of 4q35 specific genes. In general, FSHD-1 and FSHD-2 global gene deregulation interested common and distinctive biological processes. In this regard, defects of cell cycle progression (FSHD-1 and to a lesser extent FSHD-2), protein synthesis and degradation (FSHD-2), response to oxidative stress (FSHD-1 and FSHD-2), and cholesterol homeostasis (FSHD-1 and FSHD-2) may in general impair a correct myogenesis. Taken together our results recapitulate previously reported defects of FSHD-1, and add new insights into the gene deregulation characterizing both FSHD-1 and FSHD-2, in which miRNAs may play a role.

Therapeutic Rna Manipulation in Liver Disease

Posttranscriptional regulation of gene expression is increasingly recognized as a model for inherited and acquired disease. Recent work has expanded understanding of the range of mechanisms that regulate several of these distinct steps, including messenger RNA (mRNA) splicing, trafficking, and/or stability. Each of these pathways is implicated in disease pathogenesis, and each represents important avenues for therapeutic intervention. This review summarizes important mechanisms controlling mRNA processing and the regulation of mRNA degradation, including the role of microRNAs and RNA binding proteins. These pathways provide important opportunities for therapeutic targeting directed at splicing and degradation in order to attenuate genetic defects in RNA metabolism. We will highlight developments in vector development and validation for therapeutic manipulation of mRNA expression with a focus on potential applications in metabolic and immunomediated liver disease.

Spinal muscular atrophies reveal motor neuron vulnerability to defects in ribonucleoprotein handling.

Many forms of spinal muscular atrophy, a pure lower motor neuron disease, have been characterized clinically but the molecular basis of most of these is unknown. In this article we review recent developments in our understanding of the spinal muscular atrophies and how this knowledge has revealed important new insights into the causes of motor neuron vulnerability which may ultimately lead to novel therapies for this untreatable group of disorders. The identification of the genetic basis of two forms of autosomal recessive spinal muscular atrophy has revealed that lower motor neurons appear to have a specific vulnerability to defects in RNA metabolism. Most notably, the survival motor neuron protein, deficient in the most common form of spinal muscular atrophy, plays a multifunctional role in ribonucleoprotein metabolism and pre-messenger RNA splicing. The role of this protein in motor neurons is currently under intense study. The identification of two other spinal muscular atrophy-causing genes has provided support for the central role of components of the RNA metabolic pathway in determining motor neuron survival. Understanding the molecular basis of lower motor neuron vulnerability in disorders such as spinal muscular atrophy will ultimately allow these disabling disorders to be treated. In addition we can expect to learn much about basic neuronal biology and about the pathways that are relevant to more common neurodegenerative disorders such as amyotrophic lateral sclerosis.

A Common Core RNP Structure Shared between the Small Nucleoar Box C/D RNPs and the Spliceosomal U4 snRNP

The box C/D snoRNAs function in directing 2′-O-methylation and/or as chaperones in the processing of ribosomal RNA. We show here that Snu13p (15.5kD in human), a component of the U4/U6.U5 tri-snRNP, is also associated with the box C/D snoRNAs. Indeed, genetic depletion of Snu13p in yeast leads to a major defect in RNA metabolism. The box C/D motif can be folded into a stem-internal loop-stem structure, almost identical to the 15.5kD binding site in the U4 snRNA. Consistent with this, the box C/D motif binds Snu13p/15.5kD in vitro. The similarities in structure and function observed between the U4 snRNP (chaperone for U6) and the box C/D snoRNPs raises the interesting possibility that these particles may have evolved from a common ancestral RNP.

The RNA-binding properties of SMN: deletion analysis of the zebrafish orthologue defines domains conserved in evolution.

Spinal muscular atrophy (SMA) is a common autosomal recessive disorder that results in the degeneration of spinal motor neurons. SMA is caused by alterations of the survival motor neuron ( SMN ) gene which encodes a novel protein of hitherto unclear function. The SMN protein associates with ribonucleoprotein particles involved in RNA processing and exhibits an RNA-binding capacity. We have isolated the zebrafish Danio rerio and nematode Caenorhabditis elegans orthologues and have found that the RNA-binding capacity is conserved across species. Purified recombinant SMN proteins from both species showed selectivity to poly(G) homopolymer RNA in vitro, similar to that of the human protein. Studying deletions of the zebrafish SMN protein, we defined an RNA-binding element in exon 2a, which is highly conserved across species, and revealed that its binding activity is modulated by protein domains encoded by exon 2b and exon 3. Finally, the deleted recombinant zebrafish protein mimicking an SMA frameshift mutation showed a dramatic change in vitro in the formation of the RNA-protein complexes. These observations indicate that the RNA-binding capacity of SMN is an evolutionarily conserved function and further support the view that defects in RNA metabolism most likely account for the pathogenesis of SMA.

KH domains within the FMR1 sequence suggest that fragile X syndrome stems from a defect in RNA metabolism

KH domains within the FMR1 sequence suggest that fragile X syndrome stems from a defect in RNA metabolism