Genome-wide analysis of alternative pre-mRNA splicing of the Drosophila hnRNP A/B family members
Ontology highlight
ABSTRACT:
OTHER RELATED OMICS DATASETS IN: PRJNA114613PRJNA114611PRJNA114617PRJNA110279PRJNA114615
ORGANISM(S): Drosophila melanogaster
PROVIDER: GSE13940 | GEO | 2008/12/30
SECONDARY ACCESSION(S): PRJNA110279
REPOSITORIES: GEO
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Project description:Heterogeneous nuclear ribonucleoproteins (hnRNPs) have been traditionally seen as proteins packaging RNA nonspecifically into ribonucleoprotein particles (RNPs), but evidence suggests specific cellular functions on discrete target pre-mRNAs. Here we report genome-wide analysis of alternative splicing patterns regulated by four Drosophila homologs of the mammalian hnRNP A/B family (hrp36, hrp38, hrp40, and hrp48). Analysis of the global RNA-binding distributions of each protein revealed both small and extensively bound regions on target transcripts. A significant subset of RNAs were bound and regulated by more than one hnRNP protein, revealing a combinatorial network of interactions. In vitro RNA-binding site selection experiments (SELEX) identified distinct binding motif specificities for each protein, which were overrepresented in their respective regulated and bound transcripts. These results indicate that individual heterogeneous ribonucleoproteins have specific affinities for overlapping, but distinct, populations of target pre-mRNAs controlling their patterns of RNA processing.
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Project description:Genome-wide analysis of alternative pre-mRNA splicing of the Drosophila hnRNP A/B family members
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Project description:BACKGROUND: Alternative splicing (AS) of precursor mRNA (pre-mRNA) is an important gene regulation process that potentially regulates many physiological processes in plants, including the response to abiotic stresses such as salt stress. RESULTS: To analyze global changes in AS under salt stress, we obtained high-coverage (~200 times) RNA sequencing data from Arabidopsis thaliana seedlings that were treated with different concentrations of NaCl. We detected that ~49% of all intron-containing genes were alternatively spliced under salt stress, 10% of which experienced significant differential alternative splicing (DAS). Furthermore, AS increased significantly under salt stress compared with under unstressed conditions. We demonstrated that most DAS genes were not differentially regulated by salt stress, suggesting that AS may represent an independent layer of gene regulation in response to stress. Our analysis of functional categories suggested that DAS genes were associated with specific functional pathways, such as the pathways for the responses to stresses and RNA splicing. We revealed that serine/arginine-rich (SR) splicing factors were frequently and specifically regulated in AS under salt stresses, suggesting a complex loop in AS regulation for stress adaptation. We also showed that alternative splicing site selection (SS) occurred most frequently at 4 nucleotides upstream or downstream of the dominant sites and that exon skipping tended to link with alternative SS. CONCLUSIONS: Our study provided a comprehensive view of AS under salt stress and revealed novel insights into the potential roles of AS in plant response to salt stress.
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Project description:Alternative splicing plays key roles for cell type-specific regulation of protein function. It is controlled by cis-regulatory RNA elements that are recognized by RNA binding proteins (RBPs). The MALT1 paracaspase is a key factor of signaling pathways that mediate innate and adaptive immune responses. Alternative splicing of MALT1 is critical for controlling optimal T cell activation. We demonstrate that MALT1 splicing depends on RNA structural elements that sequester the splice sites of the alternatively spliced exon7. The RBPs hnRNP U and hnRNP L bind competitively to stem-loop RNA structures that involve the 5' and 3' splice sites flanking exon7. While hnRNP U stabilizes RNA stem-loop conformations that maintain exon7 skipping, hnRNP L disrupts these RNA elements to facilitate recruitment of the essential splicing factor U2AF2, thereby promoting exon7 inclusion. Our data represent a paradigm for the control of splice site selection by differential RBP binding and modulation of pre-mRNA structure.
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Project description:Understanding how RNA binding proteins control the splicing code is fundamental to human biology and disease. Here, we present a comprehensive study to elucidate how heterogeneous nuclear ribonucleoparticle (hnRNP) proteins, among the most abundant RNA binding proteins, coordinate to regulate alternative pre-mRNA splicing (AS) in human cells. Using splicing-sensitive microarrays, crosslinking and immunoprecipitation coupled with high-throughput sequencing (CLIP-seq), and cDNA sequencing, we find that more than half of all AS events are regulated by multiple hnRNP proteins and that some combinations of hnRNP proteins exhibit significant synergy, whereas others act antagonistically. Our analyses reveal position-dependent RNA splicing maps, in vivo consensus binding sites, a surprising level of cross- and autoregulation among hnRNP proteins, and the coordinated regulation by hnRNP proteins of dozens of other RNA binding proteins and genes associated with cancer. Our findings define an unprecedented degree of complexity and compensatory relationships among hnRNP proteins and their splicing targets that likely confer robustness to cells.
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Project description:The Yes-associated protein (YAP) is a transcriptional co-activator that plays critical roles in organ development and tumorigenesis, and is verified to be inhibited by the Hippo signaling pathway. In the present study, we show that the YAP 3'UTR is alternatively spliced to generate a novel 950 bp 3'UTR mRNA from the full length 3'UTR region (3483 bp) in human cancer cells. The ratio of full length 3'UTR YAP mRNA to alternatively spliced 3'UTR YAP mRNA is up-regulated by exposure of the cells to PKC inhibitor chelerythrine chloride. Further study using luciferase reporter assay showed that the expression of the alternatively spliced 3'UTR mRNA is much lower compared with the full length 3'UTR mRNA, suggesting that alternatively spliced 3'UTR YAP mRNA may have a shorter half-life than full length 3'UTR mRNA. Interestingly, PKC represses YAP 3'UTR-mediated mRNA stability is dependent on a splicing factor, hnRNP F. Activation of PKC induces nuclear translocation of cytosolic hnRNP F. Ectopic expression of hnRNP F enhances YAP 3'UTR splicing. Our results suggest that hnRNP F regulates YAP 3'UTR-mediated mRNA stability in an alternative splicing-dependent manner, and PKC regulated YAP expression is dependent on nuclear translocation of hnRNP F in human cancer cell lines.
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Project description:UnlabelledMetastatic colonization is an ominous feature of cancer progression. Recent studies have established the importance of pre-mRNA alternative splicing (AS) in cancer biology. However, little is known about the transcriptome-wide landscape of AS associated with metastatic colonization. Both in vitro and in vivo models of metastatic colonization were utilized to study AS regulation associated with cancer metastasis. Transcriptome profiling of prostate cancer cells and derivatives crossing in vitro or in vivo barriers of metastasis revealed splicing factors with significant gene expression changes associated with metastatic colonization. These include splicing factors known to be differentially regulated in epithelial-mesenchymal transition (ESRP1, ESRP2, and RBFOX2), a cellular process critical for cancer metastasis, as well as novel findings (NOVA1 and MBNL3). Finally, RNA-seq indicated a large network of AS events regulated by multiple splicing factors with altered gene expression or protein activity. These AS events are enriched for pathways important for cell motility and signaling, and affect key regulators of the invasive phenotype such as CD44 and GRHL1.ImplicationsTranscriptome-wide remodeling of AS is an integral regulatory process underlying metastatic colonization, and AS events affect the metastatic behavior of cancer cells.
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Project description:Pre-mRNA splicing is an essential step for the regulation of gene expression. In order to specifically capture splicing variants in plants for genome-wide association studies (GWAS), we developed a software tool to quantify and visualise Variations of Splicing in Population (VaSP). VaSP can quantify splicing variants from short-read RNA-seq datasets and discover genotype-specific splicing (GSS) events, which can be used to prioritise causal pre-mRNA splicing events in GWAS. We applied our method to an RNA-seq dataset with 328 samples from 82 genotypes from a rice diversity panel exposed to optimal and saline growing conditions. In total, 764 significant GSS events were identified in salt stress conditions. GSS events were used as markers for a GWAS with the shoot Na+ accumulation, which identified six GSS events in five genes significantly associated with the shoot Na+ content. Two of these genes, OsNUC1 and OsRAD23 emerged as top candidate genes with splice variants that exhibited significant divergence between the variants for shoot growth under salt stress conditions. VaSP is a versatile tool for alternative splicing analysis in plants and a powerful tool for prioritising candidate causal pre-mRNA splicing and corresponding genomic variations in GWAS.
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Project description:CHRNA1 gene, encoding the muscle nicotinic acetylcholine receptor alpha subunit, harbors an inframe exon P3A. Inclusion of exon P3A disables assembly of the acetylcholine receptor subunits. A single nucleotide mutation in exon P3A identified in congenital myasthenic syndrome causes exclusive inclusion of exon P3A. The mutation gains a de novo binding affinity for a splicing enhancing RNA-binding protein, hnRNP LL, and displaces binding of a splicing suppressing RNA-binding protein, hnRNP L. The hnRNP L binds to another splicing repressor PTB through the proline-rich region and promotes PTB binding to the polypyrimidine tract upstream of exon P3A, whereas hnRNP LL lacking the proline-rich region cannot bind to PTB. Interaction of hnRNP L with PTB inhibits association of U2AF(65) and U1 snRNP with the upstream and downstream of P3A, respectively, which causes a defect in exon P3A definition. HnRNP L and hnRNP LL thus antagonistically modulate PTB-mediated splicing suppression of exon P3A.
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