SNUPN deficiency causes splicing and mRNA expression dysregulation in primary dermal fibroblasts.
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE232712 | GEO | 2023/12/26
REPOSITORIES: GEO
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Project description:Alternative splicing (AS) can produce multiple transcripts with different exon-intron structures from a single pre-mRNA. Pre-mRNA splicing is catalyzed by a dynamic macromolecular ribonucleoprotein (RNP) complex termed the spliceosome. The spliceosome consists of several small nuclear ribonucleoproteins (snRNPs) that bind uridine-rich small nuclear RNA (snRNA). In U1, U2, U4 and U5 snRNPs, snRNA interacts with the conserved Smith antigen (Sm) proteins via a bipartite Sm sequence motif. In eukaryotes, seven Sm proteins (B, D1/2/3, E, F and G) form a heptameric ring-shaped complex surrounding the snRNA. Here, we performed a tandem affinity purification using SMEB as a bait in Arabidopsis cell suspension cultures. At least 45 known/hypothesized and potential novel spliceosome components were identified in Arabidopsis.
2023-03-03 | PXD016607 | Pride
Project description:Alterations in RNA-splicing are a molecular hallmark of several neurological diseases, including muscular dystrophies where mutations in genes involved in RNA metabolism or characterised by alterations in RNA splicing have been described. Here, we present five patients from two unrelated families with a limb-girdle muscular dystrophy (LGMD) phenotype carrying a biallelic variant in SNUPN gene. Snurportin-1, the protein encoded by SNUPN, plays an important role in the nuclear transport of small nuclear ribonucleoproteins (snRNPs), essential components of the spliceosome. We combine deep phenotyping, including clinical features, histopathology and muscle magnetic resonance image (MRI), with functional studies in patient-derived cells and muscle biopsies to demonstrate that variants in SNUPN are the cause of a new type of LGMD according to current definition. Moreover, an in vivo model in Drosophila melanogaster further supports the relevance of Snurportin-1 in muscle. SNUPN patients show a similar phenotype characterised by proximal weakness starting in childhood, restrictive respiratory dysfunction and prominent contractures, although interindividual variability in terms of severity even in individuals from the same family was found. Muscle biopsy showed myofibrillar-like features consisting of myotilin deposits and Z-disc disorganisation. MRI showed predominant impairment of paravertebral, vasti, sartorius, gracilis, peroneal and medial gastrocnemius muscles. Conservation and structural analyses of Snurportin-1 p.Ile309Ser variant suggest an effect in nuclear-cytosol snRNP trafficking. In patient-derived fibroblasts and muscle, cytoplasmic accumulation of snRNP components is observed, while total expression of Snurportin-1 and snRNPs remains unchanged, which demonstrates a functional impact of SNUPN variant in snRNP metabolism. Furthermore, RNA-splicing analysis in patients’ muscle showed widespread splicing deregulation, in particular in genes relevant for muscle development and splicing factors that participate in the early steps of spliceosome assembly. In conclusion, we report that SNUPN variants are a new cause of limb girdle muscular dystrophy with specific clinical, histopathological and imaging features, supporting SNUPN as a new gene to be included in genetic testing of myopathies. These results further support the relevance of splicing-related proteins in muscle disorders.
2024-02-29 | GSE253519 | GEO
Project description:Spliceosomal small nuclear RNAs (snRNAs) are modified by small Cajal body (CB) specific ribonucleoproteins (scaRNPs) to ensure snRNP biogenesis and pre-mRNA splicing. However, the function and subcellular site of snRNA modification are largely unknown. We show that CB localization of the protein Nopp140 is essential for concentration of scaRNPs in that nuclear condensate; and that phosphorylation by casein kinase 2 (CK2) at some 80 serines targets Nopp140 to CBs. Transiting through CBs, snRNAs are apparently modified by scaRNPs. Indeed, Nopp140 knockdown-mediated release of scaRNPs from CBs severely compromises 2’-O-methylation of spliceosomal snRNAs, identifying CBs as the site of scaRNP catalysis. Additionally, alternative splicing patterns change indicating that these modifications in U1, U2, U5, and U12 snRNAs safeguard splicing fidelity. Given the importance of CK2 in this pathway, compromised splicing could underlie the mode of action of small molecule CK2 inhibitors currently considered for therapy in cholangiocarcinoma, hematological malignancies, and COVID-19.
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Project description:Gene expression requires tight coordination of the molecular machineries that mediate transcription and splicing. While the interplay between transcription kinetics and spliceosome fidelity has been investigated before, less is known about mechanisms regulating the assembly of the spliceosomal machinery in response to transcription changes. Here we report an association of the Smn complex, which mediates spliceosomal snRNP biogenesis, with the 7SK complex involved in transcriptional regulation. We found that Smn interacts with the 7SK core components Larp7 and Mepce and specifically associates with 7SK subcomplexes containing hnRNP R. The association between Smn and 7SK complexes is enhanced upon transcriptional inhibition leading to reduced production of snRNPs. Taken together, our findings reveal a functional association of Smn and 7SK complexes that is governed by global changes in transcription. Thus, in addition to its canonical nuclear role in transcriptional regulation, 7SK has cytosolic functions in fine-tuning spliceosome production according to transcriptional demand.
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Project description:The SMA disease results from deletions or mutations in the “survival of motor neuron” (SMN1) gene. The SMN protein plays an essential role in the biogenesis of spliceosomal snRNPs but the link between a defect in this process and specific splicing inhibition is not known. Here, we report the construction of a temperature degron (td) allele of the S. pombe SMN protein and show that its depletion at 37°C affects snRNPs stability and splicing. In addition, the function of the tdSMN allele in snRNPs assembly is affected already at 25°C suggesting a deleterious effect of the tag at this temperature. Remarkably, using S. pombe tiling arrays and RT-PCR validation tests, we report that splicing is altered differentially in the tdSMN cells at permissive temperature. Our results suggest that the defects observed in tdSMN fission yeast cells mimic splicing deficits observed in SMN-deficient metazoan cells.
2010-04-22 | GSE17010 | GEO
Project description:Spn1/Iws1 is a conserved protein involved in transcription and chromatin dynamics, yet its general in vivo requirement for these functions is unknown. Using a Spn1 depletion system in S. cerevisiae, we demonstrate that Spn1 broadly influences several aspects of gene expression on a genome-wide scale. We show that Spn1 is globally required for normal mRNA levels and for normal splicing of ribosomal protein transcripts. Furthermore, Spn1 maintains the localization of H3K36 and H3K4 methylation across the genome and is required for normal histone levels at highly expressed genes. Finally, we show that the association of Spn1 with the transcription machinery is strongly dependent on its binding partner, Spt6, while the association of Spt6 and Set2 with transcribed regions is partially dependent on Spn1. Taken together, our results show that Spn1 affects multiple aspects of gene expression in vivo and provide additional evidence that it functions as a histone chaperone.
2020-07-01 | GSE153567 | GEO
Project description:The studies of spliceosomal interactions are challenging due to their dynamic nature. Here we developed spliceosome iCLIP, which immunoprecipitates SmB along with snRNPs and auxiliary RNA binding proteins (RBPs) to simultaneously map the spliceosomal binding to human snRNAs and pre-mRNAs. This identified 9 distinct regions on pre-mRNAs, which overlap with position-dependent binding patterns of 15 RBPs. Using spliceosome iCLIP, we additionally identified >50,000 branchpoints (BPs) that have canonical features, unlike those identified by RNA-seq. The iCLIP BPs generally overlap with the computationally predicted BPs, and alternative BPs are associated with extended regions of structurally accessible RNA. We find that the position and strength of BPs defines the binding patterns of SF3 and U2AF complexes, whereas the RNA structure around BPs affects the sensitivity of exons to perturbation of these complexes. Our findings introduce spliceosome iCLIP as a new method for transcriptomic studies of BPs and splicing mechanisms.
2019-08-30 | E-MTAB-6950 | biostudies-arrayexpress
Project description:The studies of spliceosomal interactions are challenging due to their dynamic nature. Here we developed spliceosome iCLIP, which immunoprecipitates SmB along with snRNPs and auxiliary RNA binding proteins (RBPs) to simultaneously map the spliceosomal binding to human snRNAs and pre-mRNAs. This identified 9 distinct regions on pre-mRNAs, which overlap with position-dependent binding patterns of 15 RBPs. Using spliceosome iCLIP, we additionally identified >50,000 branchpoints (BPs) that have canonical features, unlike those identified by RNA-seq. The iCLIP BPs generally overlap with the computationally predicted BPs, and alternative BPs are associated with extended regions of structurally accessible RNA. We find that the position and strength of BPs defines the binding patterns of SF3 and U2AF complexes, whereas the RNA structure around BPs affects the sensitivity of exons to perturbation of these complexes. Our findings introduce spliceosome iCLIP as a new method for transcriptomic studies of BPs and splicing mechanisms.
2019-08-03 | E-MTAB-8182 | biostudies-arrayexpress
Project description:Spn1/Iws1 is an essential eukaryotic transcription elongation factor that is conserved from yeast to humans. Several studies have shown that Spn1 functions as a histone chaperone to control transcription, RNA splicing, genome stability, and histone modifications as an integral member of the RNA polymerase II elongation complex. However, the precise role of Spn1 is not understood, and there is little understanding of why it is essential for viability. To address these issues, we have isolated eight suppressor mutations that bypass the essential requirement for Spn1 in Saccharomyces cerevisiae. Unexpectedly, the suppressors identify several functionally distinct complexes and activities, including the histone chaperone FACT, the histone methyltransferase Set2, the Rpd3S histone deacetylase complex, the histone acetyltransferase Rtt109, the nucleosome remodeler Chd1, and a member of the SAGA co-activator complex, Sgf73. The identification of these distinct groups and their analysis suggests that there are multiple mechanisms by which Spn1 bypass can occur, including changes in histone acetylation and alterations of other histone chaperones. Thus, Spn1 may participate in multiple functions during transcription. Our results suggest that bypass of a subset of these functions allows viability in the absence of Spn1.
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Project description:The neurodegenerative disease spinal muscular atrophy (SMA) is a leading genetic cause of infant death caused by deficiency in the survival motor neuron (SMN) protein. Currently approved SMA treatments aim to restore SMN, but the potential for expression of SMN beyond physiological levels is a unique feature of AAV9-SMN gene therapy. Here, we show that long-term AAV9-mediated SMN overexpression in mouse models induces dose-dependent, late-onset motor dysfunction associated with loss of proprioceptive synapses and neurodegeneration. Mechanistically, aggregation of overexpressed SMN in the cytoplasm of motor circuit neurons sequesters components of small nuclear ribonucleoproteins (snRNPs), leading to splicing dysregulation and widespread transcriptome abnormalities with prominent signatures of neuroinflammation and innate immune response. Thus, long-term SMN overexpression can interfere with its normal activity in RNA regulation and trigger SMA-like pathogenic events through toxic gain of function mechanisms. These unanticipated, SMN-dependent and neuron-specific liabilities of AAV9-SMN warrant further evaluation of the long-term safety of gene therapy in SMA.
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