Neurogenesis: Regulation by Alternative Splicing and Related Posttranscriptional Processes

The complexity of the mammalian brain requires highly specialized protein function and diversity. As neurons differentiate and the neuronal circuitry is established, several mRNAs undergo alternative splicing and other posttranscriptional changes that expand the variety of protein isoforms produced. Recent advances are beginning to shed light on the molecular mechanisms that regulate isoform switching during neurogenesis and the role played by specific RNA binding proteins in this process. Neurogenesis and neuronal wiring were recently shown to also be regulated by RNA degradation through nonsense-mediated decay. An additional layer of regulatory complexity in these biological processes is the interplay between alternative splicing and long noncoding RNAs. Dysregulation of posttranscriptional regulation results in defective neuronal differentiation and/or synaptic connections that lead to neurodevelopmental and psychiatric disorders.

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Targeting Poison Exons to Treat Developmental and Epileptic Encephalopathy

Developmental Neuroscience ◽

10.1159/000516143 ◽

2021 ◽

pp. 1-6

Author(s):

Miriam C. Aziz ◽

Patricia N. Schneider ◽

Gemma L. Carvill

Keyword(s):

Alternative Splicing ◽

Rna Binding ◽

De Novo ◽

Rna Binding Proteins ◽

Autism Spectrum ◽

Epileptic Encephalopathy ◽

Nonsense Mediated Decay ◽

Subsequent Degradation ◽

Alternative Splicing Events ◽

Genetic Epilepsies

Developmental and epileptic encephalopathies (DEEs) describe a subset of neurodevelopmental disorders categorized by refractory epilepsy that is often associated with intellectual disability and autism spectrum disorder. The majority of DEEs are now known to have a genetic basis with de novo coding variants accounting for the majority of cases. More recently, a small number of individuals have been identified with intronic <i>SCN1A</i> variants that result in alternative splicing events that lead to ectopic inclusion of poison exons (PEs). PEs are short highly conserved exons that contain a premature truncation codon, and when spliced into the transcript, lead to premature truncation and subsequent degradation by nonsense-mediated decay. The reason for the inclusion/exclusion of these PEs is not entirely clear, but research suggests an autoregulatory role in gene expression and protein abundance. This is seen in proteins such as RNA-binding proteins and serine/arginine-rich proteins. Recent studies have focused on targeting these PEs as a method for therapeutic intervention. Targeting PEs using antisense oligonucleotides (ASOs) has shown to be effective in modulating alternative splicing events by decreasing the amount of transcripts harboring PEs, thus increasing the abundance of full-length transcripts and thereby the amount of protein in haploinsufficient genes implicated in DEE. In the age of personalized medicine, cellular and animal models of the genetic epilepsies have become essential in developing and testing novel precision therapeutics, including PE-targeting ASOs in a subset of DEEs.

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The RNA-binding protein ELAVL1/HuR is essential for mouse spermatogenesis, acting both at meiotic and postmeiotic stages

Molecular Biology of the Cell ◽

10.1091/mbc.e11-03-0212 ◽

2011 ◽

Vol 22 (16) ◽

pp. 2875-2885 ◽

Cited By ~ 39

Author(s):

Mai Nguyen Chi ◽

Jacques Auriol ◽

Bernard Jégou ◽

Dimitris L. Kontoyiannis ◽

James M.A. Turner ◽

...

Keyword(s):

Germ Cells ◽

Posttranscriptional Regulation ◽

Target Gene ◽

Molecular Mechanisms ◽

Rna Binding ◽

Rna Binding Proteins ◽

Binding Protein ◽

Rna Binding Protein ◽

Female Sterility ◽

Targeted Deletion

Posttranscriptional mechanisms are crucial to regulate spermatogenesis. Accurate protein synthesis during germ cell development relies on RNA binding proteins that control the storage, stability, and translation of mRNAs in a tightly and temporally regulated manner. Here, we focused on the RNA binding protein Embryonic Lethal Abnormal Vision (ELAV) L1/Human antigen R (HuR) known to be a key regulator of posttranscriptional regulation in somatic cells but the function of which during gametogenesis has never been investigated. In this study, we have used conditional loss- and gain-of-function approaches to address this issue in mice. We show that targeted deletion of HuR specifically in germ cells leads to male but not female sterility. Mutant males are azoospermic because of the extensive death of spermatocytes at meiotic divisions and failure of spermatid elongation. The latter defect is also observed upon HuR overexpression. To elucidate further the molecular mechanisms underlying spermatogenesis defects in HuR-deleted and -overexpressing testes, we undertook a target gene approach and discovered that heat shock protein (HSP)A2/HSP70-2, a crucial regulator of spermatogenesis, was down-regulated in both situations. HuR specifically binds hspa2 mRNA and controls its expression at the translational level in germ cells. Our study provides the first genetic evidence of HuR involvement during spermatogenesis and reveals Hspa2 as a target for HuR.

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The CELF Family of RNA Binding Proteins Is Implicated in Cell-Specific and Developmentally Regulated Alternative Splicing

Molecular and Cellular Biology ◽

10.1128/mcb.21.4.1285-1296.2001 ◽

2001 ◽

Vol 21 (4) ◽

pp. 1285-1296 ◽

Cited By ~ 254

Author(s):

Andrea N. Ladd ◽

Nicolas Charlet-B. ◽

Thomas A. Cooper

Keyword(s):

Alternative Splicing ◽

Heart Development ◽

Binding Proteins ◽

Rna Binding ◽

Striated Muscle ◽

Rna Binding Proteins ◽

Binding Protein ◽

Protein Isoforms ◽

Developmentally Regulated ◽

Exon Inclusion

ABSTRACT Alternative splicing of cardiac troponin T (cTNT) exon 5 undergoes a developmentally regulated switch such that exon inclusion predominates in embryonic, but not adult, striated muscle. We previously described four muscle-specific splicing enhancers (MSEs) within introns flanking exon 5 in chicken cTNT that are both necessary and sufficient for exon inclusion in embryonic muscle. We also demonstrated that CUG-binding protein (CUG-BP) binds a conserved CUG motif within a human cTNT MSE and positively regulates MSE-dependent exon inclusion. Here we report that CUG-BP is one of a novel family of developmentally regulated RNA binding proteins that includes embryonically lethal abnormal vision-type RNA binding protein 3 (ETR-3). This family, which we call CELF proteins for CUG-BP- and ETR-3-like factors, specifically bound MSE-containing RNAs in vitro and activated MSE-dependent exon inclusion of cTNT minigenes in vivo. The expression of two CELF proteins is highly restricted to brain. CUG-BP, ETR-3, and CELF4 are more broadly expressed, and expression is developmentally regulated in striated muscle and brain. Changes in the level of expression and isoforms of ETR-3 in two different developmental systems correlated with regulated changes in cTNT splicing. A switch from cTNT exon skipping to inclusion tightly correlated with induction of ETR-3 protein expression during differentiation of C2C12 myoblasts. During heart development, the switch in cTNT splicing correlated with a transition in ETR-3 protein isoforms. We propose that ETR-3 is a major regulator of cTNT alternative splicing and that the CELF family plays an important regulatory role in cell-specific alternative splicing during normal development and disease.

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Novel autoregulatory cases of alternative splicing coupled with nonsense-mediated mRNA decay

10.1101/464404 ◽

2018 ◽

Author(s):

Dmitri Pervouchine ◽

Yaroslav Popov ◽

Andy Berry ◽

Beatrice Borsari ◽

Adam Frankish ◽

...

Keyword(s):

Alternative Splicing ◽

Negative Feedback ◽

Feedback Loop ◽

Transcriptional Control ◽

Rna Binding ◽

Rna Binding Proteins ◽

Negative Feedback Loop ◽

Control Of Gene Expression ◽

Nonsense Mediated Decay ◽

Premature Termination Codons

AbstractNonsense-mediated decay (NMD) is a eukaryotic mRNA surveillance system that selectively degrades transcripts with premature termination codons (PTC). Many RNA-binding proteins (RBP) regulate their expression levels by a negative feedback loop, in which RBP binds its own pre-mRNA and causes alternative splicing to introduce a PTC. We present a bioinformatic framework to identify novel such autoregulatory feedback loops by combining eCLIP assays for a large panel of RBPs with the data on shRNA inactivation of NMD pathway, and shRNA-depletion of RBPs followed by RNA-seq. We show that RBPs frequently bind their own pre-mRNAs and respond prominently to NMD pathway disruption. Poison and essential exons, i.e., exons that trigger NMD when included in the mRNA or skipped, respectively, respond oppositely to the inactivation of NMD pathway and to the depletion of their host genes, which allows identification of novel autoregulatory mechanisms for a number of human RBPs. For example, SRSF7 binds its own pre-mRNA and facilitates the inclusion of two poison exons; SFPQ binding promotes switching to an alternative distal 3’-UTR that is targeted by NMD; RPS3 activates a poison 5’-splice site in its pre-mRNA that leads to a frame shift; U2AF1 binding activates one of its two mutually exclusive exons, leading to NMD; TBRG4 is regulated by cluster splicing of its two essential exons. Our results indicate that autoregulatory negative feedback loop of alternative splicing and NMD is a generic form of post-transcriptional control of gene expression.

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Conceptual Advances in Control of Inflammation by the RNA-Binding Protein Tristetraprolin

Frontiers in Immunology ◽

10.3389/fimmu.2021.751313 ◽

2021 ◽

Vol 12 ◽

Author(s):

Pavel Kovarik ◽

Annika Bestehorn ◽

Jeanne Fesselet

Keyword(s):

Immune Responses ◽

Mrna Stability ◽

Binding Sites ◽

Molecular Mechanisms ◽

Rna Binding ◽

Rna Binding Proteins ◽

Rna Degradation ◽

Decay Rates ◽

Mrna Destabilization

Regulated changes in mRNA stability are critical drivers of gene expression adaptations to immunological cues. mRNA stability is controlled mainly by RNA-binding proteins (RBPs) which can directly cleave mRNA but more often act as adaptors for the recruitment of the RNA-degradation machinery. One of the most prominent RBPs with regulatory roles in the immune system is tristetraprolin (TTP). TTP targets mainly inflammation-associated mRNAs for degradation and is indispensable for the resolution of inflammation as well as the maintenance of immune homeostasis. Recent advances in the transcriptome-wide knowledge of mRNA expression and decay rates together with TTP binding sites in the target mRNAs revealed important limitations in our understanding of molecular mechanisms of TTP action. Such orthogonal analyses lead to the discovery that TTP binding destabilizes some bound mRNAs but not others in the same cell. Moreover, comparisons of various immune cells indicated that an mRNA can be destabilized by TTP in one cell type while it remains stable in a different cell linage despite the presence of TTP. The action of TTP extends from mRNA destabilization to inhibition of translation in a subset of targets. This article will discuss these unexpected context-dependent functions and their implications for the regulation of immune responses. Attention will be also payed to new insights into the role of TTP in physiology and tissue homeostasis.

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Transformer 2β Regulates the Alternative Splicing of Cell Cycle Regulator Genes to Promote Ovarian Cancer Cell Progression

10.21203/rs.3.rs-853170/v1 ◽

2021 ◽

Author(s):

Peiying Fu ◽

Ting Zhou ◽

Dong Chen ◽

ShiXuan Wang ◽

Ronghua Liu

Keyword(s):

Cell Cycle ◽

Ovarian Cancer ◽

Alternative Splicing ◽

Cancer Progression ◽

Poor Prognosis ◽

Molecular Mechanisms ◽

Rna Binding ◽

Rna Binding Proteins ◽

Expression Level

Abstract Background: Late-stage ovarian cancer (OV) has a poor prognosis and a high metastasis rate, but the underlying molecular mechanism is ambiguous. RNA binding proteins (RBPs) play important roles in posttranscriptional regulation in the contexts of neoplasia and tumor metastasis. Results: In this study, we explored the molecular functions of a canonical RBP, TRA2B, in cancer cells. TRA2B knockdown in HeLa cells and whole-transcriptome sequencing (RNA-seq) experiments revealed that the TRA2B-regulated alternative splicing (AS) profile was tightly associated with the mitotic cell cycle, apoptosis, and several cancer pathways. Moreover, hundreds of genes were regulated by TRA2B at the expression level, and their functions were enriched in cell proliferation, cell adhesion and angiogenesis, which are related to cancer progression. We also observed that AS regulation and expression regulation occurred independently by integrating the alternatively spliced and differentially expressed genes. We then explored and validated the carcinogenic functions of TRA2B by knocking down its expression in OV cells. In vivo, a high expression level of TRA2B was associated with a poor prognosis in OV patients. Conclusions: We demonstrated the important roles of TRA2B in ovarian neoplasia and OV progression and identified the underlying molecular mechanisms, facilitating the targeted treatment of OV in the future.

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Zebrafish mbnl mutants model physical and molecular phenotypes of myotonic dystrophy

10.1101/665380 ◽

2019 ◽

Author(s):

Melissa N. Hinman ◽

Jared I. Richardson ◽

Rose A. Sockol ◽

Eliza D Aronson ◽

Sarah J. Stednitz ◽

...

Keyword(s):

Alternative Splicing ◽

Myotonic Dystrophy ◽

Protein Function ◽

Rna Binding ◽

Rna Binding Proteins ◽

Genetic Disorder ◽

Loss Of Function ◽

High Fecundity ◽

Molecular Phenotypes ◽

Human Genetic Disorder

AbstractThe muscleblind RNA binding proteins (MBNL1, MBNL2, and MBNL3) are highly conserved across vertebrates and are important regulators of RNA alternative splicing. Loss of MBNL protein function through sequestration by CUG or CCUG RNA repeats is largely responsible for the phenotypes of the human genetic disorder myotonic dystrophy (DM). We generated the first stable zebrafish (Danio rerio) models of DM-associated MBNL loss of function through mutation of the three zebrafish mbnl genes. In contrast to mouse models, zebrafish double and triple homozygous mbnl mutants were viable to adulthood. Zebrafish mbnl mutants displayed disease-relevant physical phenotypes including decreased body size and impaired movement. They also exhibited widespread alternative splicing changes, including the misregulation of many DM-relevant exons. Physical and molecular phenotypes were more severe in compound mbnl mutants than in single mbnl mutants, suggesting partially redundant functions of Mbnl proteins. The high fecundity and larval optical transparency of this complete series of zebrafish mbnl mutants will make them useful for studying DM-related phenotypes and how individual Mbnl proteins contribute to them, and for testing potential therapeutics.

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

Blood ◽

10.1182/blood.v112.11.531.531 ◽

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.

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Posttranscriptional Regulation of Splicing Factor SRSF1 and Its Role in Cancer Cell Biology

BioMed Research International ◽

10.1155/2015/287048 ◽

2015 ◽

Vol 2015 ◽

pp. 1-10 ◽

Cited By ~ 16

Author(s):

Vânia Gonçalves ◽

Peter Jordan

Keyword(s):

Alternative Splicing ◽

Cancer Cell ◽

Cell Biology ◽

Molecular Mechanisms ◽

Rna Binding ◽

Rna Binding Proteins ◽

Current Knowledge ◽

Expression Patterns ◽

Regulatory Protein ◽

Gene Transcript

Over the past decade, alternative splicing has been progressively recognized as a major mechanism regulating gene expression patterns in different tissues and disease states through the generation of multiple mRNAs from the same gene transcript. This process requires the joining of selected exons or usage of different pairs of splice sites and is regulated by gene-specific combinations of RNA-binding proteins. One archetypical splicing regulator is SRSF1, for which we review the molecular mechanisms and posttranscriptional modifications involved in its life cycle. These include alternative splicing of SRSF1 itself, regulatory protein phosphorylation events, and the role of nuclear versus cytoplasmic SRSF1 localization. In addition, we resume current knowledge on deregulated SRSF1 expression in tumors and describe SRSF1-regulated alternative transcripts with functional consequences for cancer cell biology at different stages of tumor development.

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Dietary restriction induces posttranscriptional regulation of longevity genes

Life Science Alliance ◽

10.26508/lsa.201800281 ◽

2019 ◽

Vol 2 (4) ◽

pp. e201800281 ◽

Cited By ~ 9

Author(s):

Jarod A Rollins ◽

Dan Shaffer ◽

Santina S Snow ◽

Pankaj Kapahi ◽

Aric N Rogers

Keyword(s):

Life Span ◽

Dietary Restriction ◽

Posttranscriptional Regulation ◽

Translational Regulation ◽

Rna Binding ◽

Rna Binding Proteins ◽

Intron Retention ◽

Translation Efficiency ◽

Nonsense Mediated Decay ◽

Longevity Genes

Dietary restriction (DR) increases life span through adaptive changes in gene expression. To understand more about these changes, we analyzed the transcriptome and translatome of Caenorhabditis elegans subjected to DR. Transcription of muscle regulatory and structural genes increased, whereas increased expression of amino acid metabolism and neuropeptide signaling genes was controlled at the level of translation. Evaluation of posttranscriptional regulation identified putative roles for RNA-binding proteins, RNA editing, miRNA, alternative splicing, and nonsense-mediated decay in response to nutrient limitation. Using RNA interference, we discovered several differentially expressed genes that regulate life span. We also found a compensatory role for translational regulation, which offsets dampened expression of a large subset of transcriptionally down-regulated genes. Furthermore, 3′ UTR editing and intron retention increase under DR and correlate with diminished translation, whereas trans-spliced genes are refractory to reduced translation efficiency compared with messages with the native 5′ UTR. Finally, we find that smg-6 and smg-7, which are genes governing selection and turnover of nonsense-mediated decay targets, are required for increased life span under DR.

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