scholarly journals Asian Zika Virus Isolate Significantly Changes the Transcriptional Profile and Alternative RNA Splicing Events in a Neuroblastoma Cell Line

Viruses ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 510 ◽  
Author(s):  
Gaston Bonenfant ◽  
Ryan Meng ◽  
Carl Shotwell ◽  
Pheonah Badu ◽  
Anne F. Payne ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Zika Virus ◽  
Rna Splicing ◽  
Virus Isolate ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Mrna Splicing ◽  
Transcriptional Profile ◽  
Protein Isoforms ◽  
Functionally Diverse

The alternative splicing of pre-mRNAs expands a single genetic blueprint to encode multiple, functionally diverse protein isoforms. Viruses have previously been shown to interact with, depend on, and alter host splicing machinery. The consequences, however, incited by viral infection on the global alternative slicing (AS) landscape are under-appreciated. Here, we investigated the transcriptional and alternative splicing profile of neuronal cells infected with a contemporary Puerto Rican Zika virus (ZIKVPR) isolate, an isolate of the prototypical Ugandan ZIKV (ZIKVMR), and dengue virus 2 (DENV2). Our analyses revealed that ZIKVPR induced significantly more differential changes in expressed genes compared to ZIKVMR or DENV2, despite all three viruses showing equivalent infectivity and viral RNA levels. Consistent with the transcriptional profile, ZIKVPR induced a higher number of alternative splicing events compared to ZIKVMR or DENV2, and gene ontology analyses highlighted alternative splicing changes in genes associated with mRNA splicing. In summary, we show that ZIKV affects cellular RNA homeostasis not only at the transcriptional levels but also through the alternative splicing of cellular transcripts. These findings could provide new molecular insights into the neuropathologies associated with this virus.

scholarly journals Asian Zika virus isolate significantly changes the transcriptional profile and alternative RNA splicing events in a neuroblastoma cell line

2019 ◽  
Author(s):  
Gaston Bonenfant ◽  
Ryan Meng ◽  
Carl Shotwell ◽  
J. Andrew Berglund ◽  
Cara T. Pager
Keyword(s):  
Alternative Splicing ◽  
Zika Virus ◽  
Rna Splicing ◽  
Virus Isolate ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Transcriptional Profile ◽  
Protein Isoforms ◽  
And Function ◽  
Functionally Diverse

AbstractAlternative splicing of pre-mRNAs expands a single genetic blueprint to encode multiple functionally diverse protein isoforms. Viruses have previously been shown to interact with, depend on, and alter host splicing machinery. The consequences however incited by viral infection on the global alternative slicing (AS) landscape are under appreciated. Here we investigated the transcriptional and alternative splicing profile of neuronal cells infected with a contemporary Puerto Rican Zika virus (ZIKVPR) isolate, the prototypical Ugandan ZIKV (ZIKVMR) isolate and dengue virus 2 (DENV2). Our analyses revealed that ZIKVPRinduced significantly more differential changes in expressed genes compared to ZIKVMRor DENV2, despite all three viruses showing equivalent infectivity and viral RNA levels. Consistent with the transcriptional profile, ZIKVPRinduced a higher number of alternative splicing events compared to ZIKVMRor DENV2, and gene ontology analyses highlighted alternative splicing changes in genes associated with mRNA splicing. All three viruses modulated alternative splicing with ZIKVPRhaving the largest impact on splicing. ZIKV alteration of the transcriptomic landscape during infection caused changes in cellular RNA homeostasis, which might dysregulate neurodevelopment and function leading to neuropathologies such as microcephaly and Guillain-Barré syndrome associated with the ZIKV infection.

scholarly journals DoChaP: The Domain Change Presenter

2020 ◽  
Author(s):  
Shani T. Gal-Oz ◽  
Nimrod Haiat ◽  
Dana Eliyahu ◽  
Guy Shani ◽  
Tal Shay
Keyword(s):  
Alternative Splicing ◽  
Rna Splicing ◽  
Protein Domains ◽  
Visual Presentation ◽  
Structural Effect ◽  
Protein Isoforms ◽  
Multiple Transcripts ◽  
Genome Wide ◽  
Health And Disease ◽  
Specific Alternative

AbstractAlternative RNA splicing results in multiple transcripts of the same gene, possibly encoding for different protein isoforms with different protein domains and functionalities. Whereas it is possible to manually determine the effect of a specific alternative splicing event on the domain composition of a particular encoded protein, the process requires the tedious integration of several data sources; it is therefore error prone and its implementation is not feasible for genome-wide characterization of domains affected by differential splicing. To fulfill the need for an automated solution, we developed the Domain Change Presenter (DoChaP), a web server for the visualization of the exon–domain association. DoChaP visualizes all transcripts of a given gene, the domains of the proteins that they encode, and the exons encoding each domain. The visualization enables a comparison between the transcripts and between the protein isoforms they encode for. The organization and visual presentation of the information makes the structural effect of each alternative splicing event on the protein structure easily identified. To enable a study of the conservation of the exon structure, alternative splicing, and the effect of alternative splicing on protein domains, DoChaP also facilitates an inter-species comparison of domain–exon associations. DoChaP thus provides a unique and easy-to-use visualization of the exon–domain association and its conservation between transcripts and orthologous genes and will facilitate the study of the functional effects of alternative splicing in health and disease.

Abstract 124: Fetal-Like RNA Splicing Remodeling in Failing Heart Revealed by Total Transcriptome Analysis

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Chen Gao ◽  
Vincent Ren ◽  
Grace (Xinshu) Xiao ◽  
Jaunian Chen ◽  
Yibin Wang
Keyword(s):  
Alternative Splicing ◽  
Rna Splicing ◽  
Cardiac Development ◽  
Heart Diseases ◽  
Expression Profiles ◽  
Mrna Splicing ◽  
Mouse Heart ◽  
Failing Heart ◽  
Splicing Regulators ◽  
A Genome

The complexity of transcriptome and proteome is contributed by alternative splicing of mRNA. Altered mRNA splicing is also implicated in many human diseases including cancer. However, little knowledge is available about the scope of alternative splicing at whole genome level in heart diseases and even less about the mechanisms underlying the regulation of mRNA splicing in response to pathological injury in heart. Using a genome-wide RNA-Seq analysis, we have identified global alternative splicing changes associated with both development and pathological remodeling in mouse heart. Most significantly, the alternative RNA splicing events observed in failing heart mimicked the splicing profile in fetal hearts, suggesting a fetal like RNA splicing remodeling in failing hearts. After examining the expression profiles of splicing regulators in neonatal, normal adult, and failing adult hearts, Fox-1 was identified as one to be significantly down regulated in the failing and fetal hearts. Morpholino mediated Fox-1 knock-down in zebrafish embryos led to lethal phenotype associated with impaired cardiac development and function. This phenotype could be rescued by re-expressing both zebrafish and mouse Fox1 gene. Therefore, our established functional significance of Fox1 mediated RNA alternative splicing serves as a key molecular player in transcriptome remodeling during cardiac development and pathology.

Pre-mRNA Splicing Mechanisms, Misregulation in Disease, and Therapeutic Strategies

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. SCI-13-SCI-13
Author(s):  
Adrian Krainer
Keyword(s):  
Alternative Splicing ◽  
Research Funding ◽  
Therapeutic Strategies ◽  
Mrna Splicing ◽  
Protein Isoforms ◽  
Current Status ◽  
Targeted Therapeutics ◽  
Protein Coding ◽  
Protein Coding Genes ◽  
Normal Gene

Abstract Abstract SCI-13 Most eukaryotic protein-coding genes have one or more introns, and their transcripts can undergo alternative splicing, giving rise to multiple isoforms. Accurate splicing is essential for normal gene expression, and alternative splicing is a key mechanism for expanding the proteome and regulating the expression of diverse protein isoforms. This session will review the general mechanisms of pre-mRNA splicing and the regulation of alternative splicing. In addition, the process of how abnormal splicing arises as a result of intronic or exonic mutations in particular genes, or more globally as a result of splicing-factor misregulation, as well as the contribution of splicing misregulation to cancer, will be described. Lastly the current status of targeted therapeutics development, focusing on antisense approaches to correct abnormal splicing of specific genes or to modulate alternative splicing, will be discussed. Disclosures: Krainer: ISIS Pharmaceuticals: Consultancy, Patents & Royalties, Research Funding.

scholarly journals Long read sequencing reveals novel isoforms and insights into splicing regulation during cell state changes

BMC Genomics ◽  
2022 ◽  
Vol 23 (1) ◽  
Author(s):  
David J. Wright ◽  
Nicola A. L. Hall ◽  
Naomi Irish ◽  
Angela L. Man ◽  
Will Glynn ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Cellular Differentiation ◽  
Neuroblastoma Cell ◽  
Transcript Level ◽  
Neuroblastoma Cell Line ◽  
Future Research ◽  
Human Neuroblastoma Cell ◽  
Human Neuroblastoma ◽  
Cell State ◽  
Long Read

Abstract Background Alternative splicing is a key mechanism underlying cellular differentiation and a driver of complexity in mammalian neuronal tissues. However, understanding of which isoforms are differentially used or expressed and how this affects cellular differentiation remains unclear. Long read sequencing allows full-length transcript recovery and quantification, enabling transcript-level analysis of alternative splicing processes and how these change with cell state. Here, we utilise Oxford Nanopore Technologies sequencing to produce a custom annotation of a well-studied human neuroblastoma cell line SH-SY5Y, and to characterise isoform expression and usage across differentiation. Results We identify many previously unannotated features, including a novel transcript of the voltage-gated calcium channel subunit gene, CACNA2D2. We show differential expression and usage of transcripts during differentiation identifying candidates for future research into state change regulation. Conclusions Our work highlights the potential of long read sequencing to uncover previously unknown transcript diversity and mechanisms influencing alternative splicing.

Stage-Specific Switches in Alternative Pre-mRNA Splicing during Late Erythropoiesis Are Conserved from Mouse to Human

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 531-531
Author(s):  
Sherry Gee ◽  
Jonathan Villalobos ◽  
Miki Yamamoto ◽  
Tyson A. Clark ◽  
Jeong-Ah Kang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mrna Splicing ◽  
Protein Isoforms ◽  
Comparative Genomic ◽  
Erythroid Progenitors ◽  
Mrna Isoforms ◽  
Stop Codons ◽  
Primary Mouse

Abstract Spatial and temporal regulation of alternative pre-mRNA splicing determines which exons are incorporated into mature mRNA, modulating mRNA coding capacity to ensure synthesis of appropriate protein isoforms throughout normal differentiation and development. During erythropoiesis, a stage-specific switch in pre-mRNA splicing activates incorporation of protein 4.1R exon 16, thereby increasing 4.1R affinity for spectrin and actin and mechanically strengthening red blood cell membranes. We are exploring the hypothesis that stage-specific changes in pre-mRNA splicing regulate expression of other critical genes during terminal erythropoiesis. Last year we described exon microarray and RT-PCR studies that revealed several novel pre-mRNA splicing switches in terminally differentiating human erythroid progenitors. These alternative splicing events involved well-annotated exons with consensus exon-intron boundaries, supporting a model in which these events represent a regulated alternative splicing program rather than a breakdown of splicing integrity in late erythropoiesis. Here we report additional evidence for this model by showing that several erythroid stage-specific switches in alternative pre-mRNA splicing are conserved between human and mouse. Primary mouse splenic erythroblasts from FVA-infected mice were cultured in vitro under differentiation conditions and used as the source of RNA for analysis of murine erythroid splicing events. From a total of seven internal cassette exons whose splicing was activated in late human erythroblasts, five exhibited an analogous splicing switch in murine erythroblasts. Comparative genomic analysis showed that these alternative exons are embedded in regions of unusually high sequence conservation among vertebrate species, suggesting that important regulatory signals are contained within the adjacent introns. Indeed, the flanking introns for several of these exons contain binding motifs for Fox2, an RNA binding protein and known splicing regulator for many tissue-specific splicing events. Further analysis of the conserved erythroid splicing events revealed the following: three splicing switches occur in transcripts encoding RNA binding proteins (MBNL2, HNRPLL, and SNRP70), suggesting significant changes in the RNA processing machinery of late erythroblasts; and three of these alternative exons encode premature stop codons that could induce nonsense mediated decay (NMD) and contribute to down-regulation of these genes during terminal erythropoiesis. Consistent with the latter hypothesis, inhibition of NMD in murine erythroblast cultures led to increased accumulation of mRNA isoforms containing the premature stop codons. Together these results suggest the existence of a highly regulated alternative splicing program that is critical for late erythroid differentiation.

Abstract 235: Global RNA Splicing and Regulation in Cardiac Maturation and Diseases

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
CHEN GAO ◽  
Vincent Ren ◽  
Jae-Hyung Lee ◽  
Xinshu (Grace) Xiao ◽  
Jau-nian Chen ◽  
...  
Keyword(s):  
Heart Failure ◽  
Alternative Splicing ◽  
Cardiac Function ◽  
Rna Splicing ◽  
Cardiac Development ◽  
Mrna Splicing ◽  
Pcr Analysis ◽  
Mouse Heart ◽  
Global Pattern ◽  
Alternative Mrna Splicing

Background: The complexity of transcriptome and proteome is contributed by alternative splicing of mRNA. Altered mRNA splicing is also implicated in many human diseases including cancer. However, the global pattern of alternative mRNA splicing during cardiac development and diseases is unknown, and the regulatory mechanisms remain unexplored. Methods and Results: Using deep RNA-Sequencing, we have identified global alternative splicing changes associated with both cardiac development and pathological remodeling in mouse heart following pressure-overload induced heart failure. The alternative RNA splicing events observed in failing hearts mimics the profile in fetal hearts, suggesting a fetal-like RNA splicing program induced in diseased hearts. Using RNA-Seq database and real-time PCR analysis, we examined the expression profile of a large number of known alternative splicing regulators. Among them, we identified Fox1 as a significantly induced regulator during cardiac development in zebrafish, mouse and human, and down-regulated in both mouse and human failing hearts. Morpholino mediated Fox1 knockdown in zebrafish embryos led to lethal phenotype associated with reduced cardiac function and defects in chamber specificity. This phenotype could be rescued by re-expressing both zebrafish and mouse Fox1 gene, suggesting a highly conserved cardiac function of Fox1 for normal cardiac development and function in vertebrates. Conclusion: Our study provided the first comprehensive analysis of mRNA splicing regulation in heart during post-natal development and heart failure, and identified Fox1 as a key regulator for alternative RNA splicing in heart. This study expands our current understanding to the complexity of cardiac transcriptome, and reveals the functional importance of RNA-splicing in cardiac development and diseases.

Ribosomal Protein Rpl22 Regulates Development and Transformation Via Altering RNA Processing

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3878-3878 ◽  
Author(s):  
Minshi Wang ◽  
Zheng Ser ◽  
Shuyun Rao ◽  
Shawn Fahl ◽  
Yong Zhang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Rna Processing ◽  
Rna Splicing ◽  
Ribosome Biogenesis ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Read Depth ◽  
Mrna Splicing ◽  
Transformation Potential ◽  
Morpholino Oligonucleotides

Abstract Although it has long been reported that mutations in ribosome proteins (RP) are associated with increased cancer risk in humans, the molecular basis why which RP mutations do so remains unclear. Nevertheless, the prevailing view is that RP mutations, such as Rps19, are thought to alter transformation potential through general impairment of ribosome biogenesis or function. Importantly, recent observations are beginning to challenge this notion as too simplistic. We have determined that the RP, Rpl22, is not essential for ribosome biogenesis or global protein synthesis; however, its inactivation impairs the development of normal T lymphocytes and increases their transformation potential. Indeed, RPL22 is inactivated in human T acute lymphoblastic leukemia (T-ALL) and this is associated with reduced survival. Moreover, Rpl22-deficiency accelerates development of leukemia in a myristylated Akt2 transgenic (MyrAkt2 Tg) mouse model of T-ALL. To gain insight into how Rpl22 inactivation facilitates development of leukemia, we are performing unbiased transcriptomic and proteomic analysis on Rpl22+/+ and Rpl22-/- thymic lymphomas arising in the MyrAkt2 Tg model, and in an Rpl22-/- lymphoma reconstituted with Rpl22. Interestingly, relatively few changes in mRNA transcript read depth were observed; however, substantial differences in the proteome were observed. Pathway analysis revealed that the loss of Rpl22 altered the expression of proteins regulating RNA-processing, in particular RNA-splicing. Interestingly, interrogation of the transrciptome data for alternative splicing revealed that alterations in exon usage. The ability of Rpl22 to influence splicing appears to be conserved across species as alternative splicing was also observed in zebrafish embryos in which Rpl22 was knocked down using morpholino oligonucleotides. Consequently, we hypothesize that Rpl22 regulates biological events through its ability of binding to RNA targets, and controlling the expression of their protein products at least in part through altering mRNA splicing. How Rpl22 changes mRNA splicing pattern is currently under investigation. Disclosures No relevant conflicts of interest to declare.

scholarly journals MicroRNAs 145 and 148a Are Upregulated During Congenital Zika Virus Infection

ASN NEURO ◽  
2019 ◽  
Vol 11 ◽  
pp. 175909141985098 ◽  
Author(s):  
Fernanda L. Castro ◽  
Victor E. V. Geddes ◽  
Fábio L. L. Monteiro ◽  
Raphael M. D. T. Gonçalves ◽  
Loraine Campanati ◽  
...  
Keyword(s):  
Zika Virus ◽  
Target Genes ◽  
Neuroblastoma Cell ◽  
In Silico Analysis ◽  
Neuroblastoma Cell Line ◽  
Cellular Migration ◽  
Postmortem Brain ◽  
Zikv Infection ◽  
Cellular Mirnas ◽  
Congenital Zika Syndrome

Zika virus (ZIKV) is an arthropod-borne virus (arbovirus) member of the Flaviviridae family, which has been associated with the development of the congenital Zika syndrome (CZS). RNA viruses, such as flaviviruses, have been reported to exert a profound impact on host microRNAs (miRNAs). Cellular miRNAs modulated by ZIKV may help identify cellular pathways of relevance to pathogenesis. Here, we screened 754 human cellular miRNAs modulated by ZIKV infection (Brazilian PE strain) in a neuroblastoma cell line. Seven miRNAs (miR-99a*, miR-126*, miR-190b, miR-361-3p, miR-522-3p, miR-299-5p, and miR-1267) were downregulated during ZIKV infection, while miR-145 was upregulated. Furthermore, 11 miRNAs were exclusively expressed in ZIKV-infected (miR-148a, miR-342-5p, miR-598, and miR-708-3p) or mock cells (miR-208, miR-329, miR-432-5p, miR-488, miR-518b, miR-520g, and miR-767-5p). Furthermore, in silico analysis indicated that some central nervous system, cellular migration, and adhesion function-related biological processes were overrepresented in the list of target genes of the miRNAs regulated in ZIKV-infected cells, especially for miR-145 and miR-148a. The induction of miR-145 and miR-148a was confirmed in postmortem brain samples from stillborn with severe CZS. Finally, we determined the expression regulation of microcephaly related genes through RNA interference pathway caused by ZIKV directly on neuron cells.

scholarly journals Long read sequencing reveals novel isoforms and insights into splicing regulation during cell state changes

2021 ◽  
Author(s):  
David J Wright ◽  
Nicola Hall ◽  
Naomi Irish ◽  
Angela L Man ◽  
Will Glynn ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Cellular Differentiation ◽  
Neuroblastoma Cell ◽  
Transcript Level ◽  
Neuroblastoma Cell Line ◽  
Human Neuroblastoma Cell ◽  
Human Neuroblastoma ◽  
Cell State ◽  
Novel Transcript ◽  
Long Read

ABSTRACTAlternative splicing (AS) is a key mechanism underlying cellular differentiation and a driver of complexity in mammalian neuronal tissues. However, understanding of which isoforms are differentially used or expressed and how this affects cellular differentiation remains unclear. Long read sequencing allows full-length transcript recovery and quantification, enabling transcript-level analysis of AS processes and how these change with cell state. Here, we utilise Oxford Nanopore Technologies sequencing to produce a custom annotation of a well-studied human neuroblastoma cell line and to characterise isoform expression and usage across differentiation. We identify many previously unannotated features, including a novel transcript of the voltage-gated calcium channel subunit gene, CACNA2D2. We show differential expression and usage of transcripts during differentiation, and identify a putative molecular regulator underlying this state change. Our work highlights the potential of long read sequencing to uncover previously unknown transcript diversity and mechanisms influencing alternative splicing.

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