Abstract 3161: Massively parallel transcriptome sequencing in neuroblastoma reveals specific mRNA splicing patterns controlled by RNA binding splicing factors.

2013 ◽  
Author(s):  
Shile Zhang ◽  
Tom Badgett ◽  
Jun Wei ◽  
Young K. Song ◽  
Dominik Bogen ◽  
...  
Keyword(s):  
Transcriptome Sequencing ◽  
Rna Binding ◽  
Mrna Splicing ◽  
Massively Parallel ◽  
Splicing Factors ◽  
Splicing Patterns ◽  
Specific Mrna

scholarly journals miRNA-Based Regulation of Alternative RNA Splicing in Metazoans

2021 ◽  
Vol 22 (21) ◽  
pp. 11618
Author(s):  
Anna L. Schorr ◽  
Marco Mangone
Keyword(s):  
Neurological Disorders ◽  
Rna Splicing ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Transcriptional Level ◽  
Dependent Manner ◽  
Non Coding Rnas ◽  
Specific Mrna

Alternative RNA splicing is an important regulatory process used by genes to increase their diversity. This process is mainly executed by specific classes of RNA binding proteins that act in a dosage-dependent manner to include or exclude selected exons in the final transcripts. While these processes are tightly regulated in cells and tissues, little is known on how the dosage of these factors is achieved and maintained. Several recent studies have suggested that alternative RNA splicing may be in part modulated by microRNAs (miRNAs), which are short, non-coding RNAs (~22 nt in length) that inhibit translation of specific mRNA transcripts. As evidenced in tissues and in diseases, such as cancer and neurological disorders, the dysregulation of miRNA pathways disrupts downstream alternative RNA splicing events by altering the dosage of splicing factors involved in RNA splicing. This attractive model suggests that miRNAs can not only influence the dosage of gene expression at the post-transcriptional level but also indirectly interfere in pre-mRNA splicing at the co-transcriptional level. The purpose of this review is to compile and analyze recent studies on miRNAs modulating alternative RNA splicing factors, and how these events contribute to transcript rearrangements in tissue development and disease.

scholarly journals RES complex is associated with intron definition and required for zebrafish early embryogenesis

2017 ◽  
Author(s):  
Juan P. Fernandez ◽  
Miguel A. Moreno-Mateos ◽  
Andre Gohr ◽  
Shun Hang Chan ◽  
Manuel Irimia ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mrna Splicing ◽  
Vertebrate Development ◽  
Loss Of Function ◽  
Recognition Elements ◽  
Splicing Patterns ◽  
The Brain ◽  
Intron Definition

AbstractPre-mRNA splicing is a critical step of gene expression in eukaryotes. Transcriptome-wide splicing patterns are complex and primarily regulated by a diverse set of recognition elements and associated RNA-binding proteins. The retention and splicing (RES) complex is formed by three different proteins (Bud13p, Pml1p and Snu17p) and is involved in splicing in yeast. However, the importance of the RES complex for vertebrate splicing, the intronic features associated with its activity, and its role in development are unknown. In this study, we have generated loss-of-function mutants for the three components of the RES complex in zebrafish and showed that they are required during early development. The mutants showed a marked neural phenotype with increased cell death in the brain and a decrease in differentiated neurons. Transcriptomic analysis of bud13, snip1 (pml1) and rbmx2 (snu17) mutants revealed a global defect in intron splicing, with strong mis-splicing of a subset of introns. We found these RES-dependent introns were short, rich in GC and flanked by GC depleted exons, all of which are features associated with intron definition. Using these features we developed a predictive model that classifies RES dependent introns. Altogether, our study uncovers the essential role of the RES complex during vertebrate development and provides new insights into its function during splicing.

scholarly journals Serine Phosphorylation of SR Proteins Is Required for Their Recruitment to Sites of Transcription In Vivo

1998 ◽  
Vol 143 (2) ◽  
pp. 297-307 ◽  
Author(s):  
Tom Misteli ◽  
Javier F. Cáceres ◽  
Jade Q. Clement ◽  
Adrian R. Krainer ◽  
Miles F. Wilkinson ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Active Sites ◽  
Rna Binding ◽  
Sr Proteins ◽  
Mrna Splicing ◽  
Serine Phosphorylation ◽  
Splicing Factors ◽  
Nuclear Speckles

Expression of most RNA polymerase II transcripts requires the coordinated execution of transcription, splicing, and 3′ processing. We have previously shown that upon transcriptional activation of a gene in vivo, pre-mRNA splicing factors are recruited from nuclear speckles, in which they are concentrated, to sites of transcription (Misteli, T., J.F. Cáceres, and D.L. Spector. 1997. Nature. 387:523–527). This recruitment process appears to spatially coordinate transcription and pre-mRNA splicing within the cell nucleus. Here we have investigated the molecular basis for recruitment by analyzing the recruitment properties of mutant splicing factors. We show that multiple protein domains are required for efficient recruitment of SR proteins from nuclear speckles to nascent RNA. The two types of modular domains found in the splicing factor SF2/ ASF exert distinct functions in this process. In living cells, the RS domain functions in the dissociation of the protein from speckles, and phosphorylation of serine residues in the RS domain is a prerequisite for this event. The RNA binding domains play a role in the association of splicing factors with the target RNA. These observations identify a novel in vivo role for the RS domain of SR proteins and suggest a model in which protein phosphorylation is instrumental for the recruitment of these proteins to active sites of transcription in vivo.

scholarly journals PsiCLIP reveals dynamic RNA binding by DEAH-box helicases before and after exon ligation

2020 ◽  
Author(s):  
Lisa M. Strittmatter ◽  
Charlotte Capitanchik ◽  
Andrew J. Newman ◽  
Martina Hallegger ◽  
Christine M. Norman ◽  
...  
Keyword(s):  
Rna Binding ◽  
Positional Information ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Imaging Studies ◽  
Rna Helicases ◽  
Binding Profile ◽  
Exon Junction ◽  
Exon Ligation ◽  
Before And After

AbstractEight RNA helicases remodel the spliceosome to effect pre-mRNA splicing but their mechanism of action remains poorly understood. We have developed “purified spliceosome iCLIP” (psiCLIP) to define helicase-RNA contacts in specific spliceosomal states. psiCLIP reveals previously unappreciated dynamics of spliceosomal helicases. The binding profile of the helicase Prp16 is influenced by the distance between the branch-point and 3’ splice site, while Prp22 binds diffusely on the intron before exon ligation but switches to more narrow binding downstream of the exon junction after exon ligation. Notably, depletion of the exon-ligation factor Prp18 destabilizes Prp22 binding to the pre-mRNA, demonstrating that psiCLIP can be used to study the relationships between helicases and auxiliary splicing factors. Thus, psiCLIP is sensitive to spliceosome dynamics and complements the insights from structural and imaging studies by providing crucial positional information on helicase-RNA contacts during spliceosomal remodeling.

scholarly journals Sip1, a Novel RS Domain-Containing Protein Essential for Pre-mRNA Splicing

1998 ◽  
Vol 18 (2) ◽  
pp. 676-684 ◽  
Author(s):  
Wan-Jiang Zhang ◽  
Jane Y. Wu
Keyword(s):  
Protein Interactions ◽  
Rna Binding ◽  
Sequence Similarity ◽  
Functional Characterization ◽  
Nuclear Extract ◽  
Mrna Splicing ◽  
Splicing Factor ◽  
Splicing Factors ◽  
Interacting Protein

ABSTRACT Previous studies have shown that protein-protein interactions among splicing factors may play an important role in pre-mRNA splicing. We report here identification and functional characterization of a new splicing factor, Sip1 (SC35-interacting protein 1). Sip1 was initially identified by virtue of its interaction with SC35, a splicing factor of the SR family. Sip1 interacts with not only several SR proteins but also with U1-70K and U2AF65, proteins associated with 5′ and 3′ splice sites, respectively. The predicted Sip1 sequence contains an arginine-serine-rich (RS) domain but does not have any known RNA-binding motifs, indicating that it is not a member of the SR family. Sip1 also contains a region with weak sequence similarity to the Drosophila splicing regulator suppressor of white apricot (SWAP). An essential role for Sip1 in pre-mRNA splicing was suggested by the observation that anti-Sip1 antibodies depleted splicing activity from HeLa nuclear extract. Purified recombinant Sip1 protein, but not other RS domain-containing proteins such as SC35, ASF/SF2, and U2AF65, restored the splicing activity of the Sip1-immunodepleted extract. Addition of U2AF65 protein further enhanced the splicing reconstitution by the Sip1 protein. Deficiency in the formation of both A and B splicing complexes in the Sip1-depleted nuclear extract indicates an important role of Sip1 in spliceosome assembly. Together, these results demonstrate that Sip1 is a novel RS domain-containing protein required for pre-mRNA splicing and that the functional role of Sip1 in splicing is distinct from those of known RS domain-containing splicing factors.

scholarly journals Mechanistic Insights of Aberrant Splicing with Splicing Factor Mutations Found in Myelodysplastic Syndromes

2021 ◽  
Vol 22 (15) ◽  
pp. 7789
Author(s):  
Naoyuki Kataoka ◽  
Eri Matsumoto ◽  
So Masaki
Keyword(s):  
Myelodysplastic Syndrome ◽  
Regulatory Elements ◽  
Mrna Splicing ◽  
Driver Mutations ◽  
Splicing Factors ◽  
Aberrant Splicing ◽  
Order Of Accuracy ◽  
Splicing Regulators ◽  
Risk Of Progression ◽  
Splicing Patterns

Pre-mRNA splicing is an essential process for gene expression in higher eukaryotes, which requires a high order of accuracy. Mutations in splicing factors or regulatory elements in pre-mRNAs often result in many human diseases. Myelodysplastic syndrome (MDS) is a heterogeneous group of chronic myeloid neoplasms characterized by many symptoms and a high risk of progression to acute myeloid leukemia. Recent findings indicate that mutations in splicing factors represent a novel class of driver mutations in human cancers and affect about 50% of Myelodysplastic syndrome (MDS) patients. Somatic mutations in MDS patients are frequently found in genes SF3B1, SRSF2, U2AF1, and ZRSR2. Interestingly, they are involved in the recognition of 3′ splice sites and exons. It has been reported that mutations in these splicing regulators result in aberrant splicing of many genes. In this review article, we first describe molecular mechanism of pre-mRNA splicing as an introduction and mainly focus on those four splicing factors to describe their mutations and their associated aberrant splicing patterns.

scholarly journals RBM20 phosphorylation on serine/arginine domain is crucial to regulate pre-mRNA splicing and protein shuttling in the heart

2020 ◽  
Author(s):  
Mingming Sun ◽  
Yutong Jin ◽  
Chaoqun Zhu ◽  
Yanghai Zhang ◽  
Martin Liss ◽  
...  
Keyword(s):  
Protein Trafficking ◽  
Rna Binding ◽  
Heart Function ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Binding Motif ◽  
Amino Acid Residues ◽  
Cellular Mechanisms ◽  
Sr Protein ◽  
Protein Protein Interaction

AbstractMolecular and cellular mechanisms of mutations of splicing factors in heart function are not well understood. The splicing of precursor mRNA is dependent on an essential group of splicing factors containing serine-arginine (SR) domain(s) that are critical for protein-RNA and protein-protein interaction in the spliceosome assembly. Phosphorylation of SR domains plays a key role in splicing control and the distribution of splicing factors in the cell. RNA binding motif 20 (RBM20) is a splicing factor predominantly expressed in muscle tissues with the highest expression level in the heart. However, its phosphorylation status is completely unknown up-to-date. In this study, we identified sixteen amino acid residues that are phosphorylated by middle-down mass spectrometry. Four of them are located in the SR domain, and two out of the four residues, S638 and S640, play an essential role in splicing control and facilitate RBM20 shuttling from the nucleus to the cytoplasm. Re-localization of RBM20 promotes protein aggregation in the cytoplasm. We have also verified that SR-protein kinases (SRPKs), cdc2-like kinases (CLKs) and protein kinase B (PKB or AKT) phosphorylate S638 and S640. Mutations of S638A and S640G reduce RBM20 phosphorylation and disrupt the splicing. Taken together, we determine the phosphorylation status of RBM20 and provide the first evidence that phosphorylation on SR domain is crucial for pre-mRNA splicing and protein trafficking. Our findings reveal a new role of RBM20 via protein shuttling in cardiac function.

scholarly journals Poly(A) Polymerase and the Nuclear Poly(A) Binding Protein, PABPN1, Coordinate the Splicing and Degradation of a Subset of Human Pre-mRNAs

2015 ◽  
Vol 35 (13) ◽  
pp. 2218-2230 ◽  
Author(s):  
Lisa Muniz ◽  
Lee Davidson ◽  
Steven West
Keyword(s):  
Rna Binding ◽  
Binding Protein ◽  
Mrna Splicing ◽  
Oculopharyngeal Muscular Dystrophy ◽  
Splicing Factors ◽  
Human Genes ◽  
Protein Encoding ◽  
Cleavage And Polyadenylation ◽  
Regulatory Potential ◽  
Terminal Poly

Most human protein-encoding transcripts contain multiple introns that are removed by splicing. Although splicing catalysis is frequently cotranscriptional, some introns are excised after polyadenylation. Accumulating evidence suggests that delayed splicing has regulatory potential, but the mechanisms are still not well understood. Here we identify a terminal poly(A) tail as being important for a subset of intron excision events that follow cleavage and polyadenylation. In these cases, splicing is promoted by the nuclear poly(A) binding protein, PABPN1, and poly(A) polymerase (PAP). PABPN1 promotes intron excision in the context of 3′-end polyadenylation but not when bound to internal A-tracts. Importantly, the ability of PABPN1 to promote splicing requires its RNA binding and, to a lesser extent, PAP-stimulatory functions. Interestingly, an N-terminal alanine expansion in PABPN1 that is thought to cause oculopharyngeal muscular dystrophy cannot completely rescue the effects of PABPN1 depletion, suggesting that this pathway may have relevance to disease. Finally, inefficient polyadenylation is associated with impaired recruitment of splicing factors to affected introns, which are consequently degraded by the exosome. Our studies uncover a new function for polyadenylation in controlling the expression of a subset of human genes via pre-mRNA splicing.

scholarly journals A Collection of Pre-mRNA Splicing Mutants in Arabidopsis thaliana

2020 ◽  
Vol 10 (6) ◽  
pp. 1983-1996 ◽  
Author(s):  
Tatsuo Kanno ◽  
Peter Venhuizen ◽  
Ming-Tsung Wu ◽  
Phebe Chiou ◽  
Chia-Liang Chang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Arabidopsis Thaliana ◽  
Alternative Splicing ◽  
Reporter Gene ◽  
Rna Binding ◽  
Binding Protein ◽  
Genetic Screen ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Sequencing Data

To investigate factors influencing pre-mRNA splicing in plants, we conducted a forward genetic screen using an alternatively-spliced GFP reporter gene in Arabidopsis thaliana. This effort generated a collection of sixteen mutants impaired in various splicing-related proteins, many of which had not been recovered in any prior genetic screen or implicated in splicing in plants. The factors are predicted to act at different steps of the spliceosomal cycle, snRNP biogenesis pathway, transcription, and mRNA transport. We have described eleven of the mutants in recent publications. Here we present the final five mutants, which are defective, respectively, in RNA-BINDING PROTEIN 45D (rbp45d), DIGEORGE SYNDROME CRITICAL REGION 14 (dgcr14), CYCLIN-DEPENDENT KINASE G2 (cdkg2), INTERACTS WITH SPT6 (iws1) and CAP BINDING PROTEIN 80 (cbp80). We provide RNA-sequencing data and analyses of differential gene expression and alternative splicing patterns for the cbp80 mutant and for several previously published mutants, including smfa and new alleles of cwc16a, for which such information was not yet available. Sequencing of small RNAs from the cbp80 mutant highlighted the necessity of wild-type CBP80 for processing of microRNA (miRNA) precursors into mature miRNAs. Redundancy tests of paralogs encoding several of the splicing factors revealed their functional non-equivalence in the GFP reporter gene system. We discuss the cumulative findings and their implications for the regulation of pre-mRNA splicing efficiency and alternative splicing in plants. The mutant collection provides a unique resource for further studies on a coherent set of splicing factors and their roles in gene expression, alternative splicing and plant development.

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