scholarly journals Alternative REST Splicing Underappreciated

2017 ◽  
Author(s):  
Guo-Lin Chen ◽  
Gregory M. Miller
Keyword(s):  
Alternative Splicing ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Protein Isoforms ◽  
Multiple Protein ◽  
Mrna Variants ◽  
Cellular Context ◽  
Active Transcription ◽  
Context Dependent ◽  
Ageing Brain

As a major orchestrator of the cellular epigenome, the repressor element-1 silencing transcription factor (REST) can either repress or activate thousands of genes depending on cellular context, suggesting a highly context-dependent REST function tuned by environmental cues. While REST shows cell-type non-selective active transcription1, an N-terminal REST4 isoform caused by alternative splicing – inclusion of an extra exon (N3c) which introduces a premature stop codon – has been implicated in neurogenesis and tumorigenesis2-5. Recently, in line with established epigenetic regulation of pre-mRNA splicing6,7, we demonstrated that REST undergoes extensive, context-dependent alternative splicing which results in the formation of a large number of mRNA variants predictive of multiple protein isoforms8. Supported by that immunoblotting/-staining with different anti-REST antibodies yield inconsistent results, alternative splicing allows production of various structurally and functionally different REST protein isoforms in response to shifting physiological requirements, providing a reasonable explanation for the diverse, highly context-dependent REST function. However, REST isoforms might be differentially assayed or manipulated, leading to data misinterpretation and controversial findings. For example, in contrast to the proposed neurotoxicity of elevated nuclear REST in ischemia9 and Huntington’s disease10,11, Lu et al. recently reported decreased nuclear REST in Alzheimer’s disease and neuroprotection of REST in ageing brain12. Unfortunately, alternative REST splicing was largely neglected by Lu et al., making it necessary for a reevaluation of their findings.

Abstract 293: Tcf7l2 Expression And Isoform Switching In Mouse Hearts

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
LU XIAO ◽  
Haiqing Bai ◽  
James Boyer ◽  
Bo Ye ◽  
Ning Hou ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Wnt Signaling ◽  
Stop Codon ◽  
Medical Center ◽  
Premature Stop Codon ◽  
Beta Catenin ◽  
Canonical Wnt Signaling ◽  
Cardiovascular Research ◽  
Reading Frame ◽  
Canonical Wnt

Lu Xiao, Haiqing Bai, James Boyer, Bo Ye, Ning Hou, Haodong Xu, and Faqian Li Department of Pathology and Laboratory Medicine and Cardiovascular Research Institute, University of Rochester Medical Center, Rochester, NY, USA Backgrounds: Canonical Wnt signaling appears to have multiphasic and often antagonistic roles in cardiac development. The molecular mechanism for these opposing actions is not clear. We hypothesized that alternative splicing of TCF7L2, a nuclear interaction partner of beta-catenin is involved in the specificity of canonical Wnt signaling. Methods: RT-PCR were performed on embryonic (E16.5) and neonatal (day 8) hearts with primers spanning the end of first exon and the beginning of last exon and the products were cloned and sequenced. Result: There are totally 18 exons identified so far in TCF7L2. We sequenced 56 clones and 53 clones (29 from day 8) and (24 from E16.5) contained TCF7L2 sequences. No exon 6 or exon 17 was found in TCF7L2 transcripts of mouse hearts. Most clones (more than 80%) from E16.5 and day 8 hearts excluded exon 4. Both E16.5 and day 8 hearts had one clone with exon 9 deletion which does not change reading frame and another with alterations in exon 3 that lead to reading frame shift and premature stop codon. As reported in other organs, there were extensive alternative splicing in the C-terminal exons 14, 15 and 16. The inclusion of exon 14 was more frequently in day 8 (18 of 29, 62%) than in E16.5 (8 of 24, 33%) hearts. The peptide encoded by exon 14 has conserved functional motif. Additionally, this alternative exon usage can change the C-terminus of TCF7L2 to include or exclude the so-called E tail with two binding motifs for C-terminal binding protein. Conclusion: The isoform switch of TCF7L2 occurs in neonatal mouse hearts and may have a role in the terminal differentiation of cardiac myocytes during this period.

scholarly journals Alternative splicing in endothelial cells: novel therapeutic opportunities in cancer angiogenesis

2020 ◽  
Vol 39 (1) ◽  
Author(s):  
Anna Di Matteo ◽  
Elisa Belloni ◽  
Davide Pradella ◽  
Ambra Cappelletto ◽  
Nina Volf ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Human Cancer ◽  
Single Gene ◽  
Predictive Biomarkers ◽  
Protein Isoforms ◽  
Therapeutic Interventions ◽  
Cancer Therapies ◽  
Functional Changes ◽  
Splicing Regulators ◽  
Multiple Protein

AbstractAlternative splicing (AS) is a pervasive molecular process generating multiple protein isoforms, from a single gene. It plays fundamental roles during development, differentiation and maintenance of tissue homeostasis, while aberrant AS is considered a hallmark of multiple diseases, including cancer. Cancer-restricted AS isoforms represent either predictive biomarkers for diagnosis/prognosis or targets for anti-cancer therapies. Here, we discuss the contribution of AS regulation in cancer angiogenesis, a complex process supporting disease development and progression. We consider AS programs acting in a specific and non-redundant manner to influence morphological and functional changes involved in cancer angiogenesis. In particular, we describe relevant AS variants or splicing regulators controlling either secreted or membrane-bound angiogenic factors, which may represent attractive targets for therapeutic interventions in human cancer.

An Autosomal-Recessive GFI1B Mutation Defines the Splice Isoform p37 As Essential for Biogenesis of Functional Human Platelets, but Dispensable for Erythropoiesis

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2644-2644
Author(s):  
Harald Schulze ◽  
Axel Schlagenhauf ◽  
Georgi Manukjan ◽  
Christine Beham-Schmid ◽  
Oliver Andres ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Growth Factor ◽  
Autosomal Recessive ◽  
Stop Codon ◽  
Zinc Fingers ◽  
Premature Stop Codon ◽  
Index Patient ◽  
Homozygous Mutation ◽  
Recessive Trait ◽  
Erythroid Progenitor

Abstract Growth factor independent 1 (GFI1) and Growth Factor Independent 1B (GFI1B) are zinc finger transcriptional repressors that share about 90% homology on amino acid sequence and are expressed during hematopoiesis. While GFI1 is most important for granulocyte-monocyte lineage commitment, GFI1B is an essential master regulator of erythroid and megakaryocytic lineages. Mice lacking Gfi1b are embryonic lethal due to anemia and thrombocytopenia. In humans, alternative splicing leads to a shorter p32 isoform that lacks the first 2 of 6 zinc fingers. GFI1B germline mutations have been reported to cause autosomal-dominant macrothrombocytopenia with a grey-platelet syndrome phenotype, implying that the mutant protein acts in a dominant-negative manner. We report on a Chechen family from eastern Georgia whose affected family members all present with severe, life-threatening bleeding diathesis. The female index patient had recurrent hematomata and multiple petechiae since childhood. Both of her children (age 9 and 7) present with very low platelet counts (below 45/nL) and a similar cutaneous bleeding pattern like her mother. The brother also had thrombocytopenia and died at age 33 in reponse to a spontaneous cerebral hemorrhage. In contrast, the index patient's husband, her parents and the children of the deceased brother were clinically unaffected. Blood smears of affected patients showed macrothrombocytopenia with reduced May-Grünwald-Giema staining and decreased staining for alpha-granule markers von Willebrand factor (vWF) and P-selectin (CD62P). Platelet function testing revealed reduced responses to ADP, collagen, TRAP-6 and arachidonic acid. White and red blood cell parameters were overall normal in the index patient and the two affected children. We analyzed DNA from the index patient by targeted next generation sequencing for 59 genes relevant for platelet formation or function. We found a novel homozygous single nucleotide insertion in GFI1B (NM_004188.5; c.551insG), which was confirmed by Sanger sequencing and is expected to cause a premature stop-codon. The homozygous mutation co-segregated with the phenotype. The unaffected mother, the husband and two unaffected nephews were heterozygous, suggesting a local founder variant and an unexpected autosomal-recessive trait. Bone marrow analysis showed unaffected myeloid and erythroid cells, but dysplastic micromegakaryocytes with increased CD34 staining. Peripheral blood platelets were also positive for CD34. We performed quantitative real-time PCR of platelet RNA and found residual homozygous c.551_G insertion in the p37 transcript and an unexpected expression of the p32 variant. The p37 transcript was markedly reduced in context with an increased p32/p37 ratio compared to controls. Our findings indicate that the mutated transcript was not completely degraded by nonsense-mediated decay, but mostly subjected to alternative splicing skipping the mutated exon 9. Our findings imply that the first two zinc fingers of GFI1B are dispensable for human erythropoiesis, but essential for normal megakaryopoiesis and the production of functional platelets. While previous mutations affect both isoforms, the insertion variant presented here, results in a premature stop-codon and affects only the p37 isoform due to alternative splicing. This splice variant defines an important node at the megakaryocytic-erythroid progenitor stage and we conclude that the transcriptional regulation of erythropoiesis is uncoupled from that of megakaryopoiesis through alternative splicing of GFI1B. Disclosures No relevant conflicts of interest to declare.

scholarly journals Interferon lambda 4 expression is suppressed by the host during viral infection

2016 ◽  
Vol 213 (12) ◽  
pp. 2539-2552 ◽  
Author(s):  
MeeAe Hong ◽  
Johannes Schwerk ◽  
Chrissie Lim ◽  
Alison Kell ◽  
Abigail Jarret ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Splice Variants ◽  
Stop Codon ◽  
Association Studies ◽  
Premature Stop Codon ◽  
Genetic Association Studies ◽  
Protein Isoforms ◽  
Translation Efficiency ◽  
Regulation Of Expression ◽  
Protein Coding

Interferon (IFN) lambdas are critical antiviral effectors in hepatic and mucosal infections. Although IFNλ1, IFNλ2, and IFNλ3 act antiviral, genetic association studies have shown that expression of the recently discovered IFNL4 is detrimental to hepatitis C virus (HCV) infection through a yet unknown mechanism. Intriguingly, human IFNL4 harbors a genetic variant that introduces a premature stop codon. We performed a molecular and biochemical characterization of IFNλ4 to determine its role and regulation of expression. We found that IFNλ4 exhibits similar antiviral activity to IFNλ3 without negatively affecting antiviral IFN activity or cell survival. We show that humans deploy several mechanisms to limit expression of functional IFNλ4 through noncoding splice variants and nonfunctional protein isoforms. Furthermore, protein-coding IFNL4 mRNA are not loaded onto polyribosomes and lack a strong polyadenylation signal, resulting in poor translation efficiency. This study provides mechanistic evidence that humans suppress IFNλ4 expression, suggesting that immune function is dependent on other IFNL family members.

scholarly journals Alternative splicing of the proadrenomedullin gene results in differential expression of gene products

2001 ◽  
Vol 27 (1) ◽  
pp. 31-41 ◽  
Author(s):  
A Martinez ◽  
DL Hodge ◽  
M Garayoa ◽  
HA Young ◽  
F Cuttitta
Keyword(s):  
Alternative Splicing ◽  
Bioactive Peptides ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Ribonuclease Protection Assay ◽  
Gene Products ◽  
Protection Assay ◽  
The Third ◽  
Ribonuclease Protection ◽  
Fourth Exon

The adrenomedullin (AM) gene codifies for two bioactive peptides, AM and proAM N-terminal 20 peptide (PAMP). We have found two forms of the AM mRNA. Form A is devoid of introns and results in a prohormone containing both peptides. Form B retains the third intron, which introduces a premature stop codon, producing a shorter prohormone with only PAMP. Tissues with a higher B/A ratio were more immunoreactive for PAMP than for AM. The form B message was found in the cytoplasmic compartment, thus excluding that the longer message was a result of contaminating nuclear mRNA. Form B was found in cells that express PAMP but not AM. mRNA expression in a variety of cell lines was investigated by ribonuclease protection assay and form B was found in significant amounts in two of them. Treatments that modify AM expression, such as exposure to hypoxia, were shown to change the B/A ratio and the relative secretion of AM and PAMP, indicating that the splicing mechanism for AM can be modulated and is physiologically relevant. Analysis of the sequence of the third intron and the fourth exon of the AM gene found motifs compatible with a highly regulated alternative splicing mechanism.

scholarly journals Antisense Oligonucleotide-Mediated Splice Switching: Potential Therapeutic Approach for Cancer Mitigation

Cancers ◽  
2021 ◽  
Vol 13 (21) ◽  
pp. 5555
Author(s):  
Prithi Raguraman ◽  
Akilandeswari Ashwini Balachandran ◽  
Suxiang Chen ◽  
Sarah D. Diermeier ◽  
Rakesh N. Veedu
Keyword(s):  
Cancer Progression ◽  
Messenger Rna ◽  
Splice Variants ◽  
Stop Codon ◽  
Mrna Level ◽  
Premature Stop Codon ◽  
Exon Skipping ◽  
Protein Isoforms ◽  
Delivery Strategies ◽  
And Function

Splicing is an essential process wherein precursor messenger RNA (pre-mRNA) is reshaped into mature mRNA. In alternative splicing, exons of any pre-mRNA get rearranged to form mRNA variants and subsequently protein isoforms, which are distinct both by structure and function. On the other hand, aberrant splicing is the cause of many disorders, including cancer. In the past few decades, developments in the understanding of the underlying biological basis for cancer progression and therapeutic resistance have identified many oncogenes as well as carcinogenic splice variants of essential genes. These transcripts are involved in various cellular processes, such as apoptosis, cell signaling and proliferation. Strategies to inhibit these carcinogenic isoforms at the mRNA level are promising. Antisense oligonucleotides (AOs) have been developed to inhibit the production of alternatively spliced carcinogenic isoforms through splice modulation or mRNA degradation. AOs can also be used to induce splice switching, where the expression of an oncogenic protein can be inhibited by the induction of a premature stop codon. In general, AOs are modified chemically to increase their stability and binding affinity. One of the major concerns with AOs is efficient delivery. Strategies for the delivery of AOs are constantly being evolved to facilitate the entry of AOs into cells. In this review, the different chemical modifications employed and delivery strategies applied are discussed. In addition to that various AOs in clinical trials and their efficacy are discussed herein with a focus on six distinct studies that use AO-mediated exon skipping as a therapeutic strategy to combat cancer.

scholarly journals Regulation of FXR1 by alternative splicing is required for muscle development and controls liquid-like condensates in muscle cells

2019 ◽  
Author(s):  
Jean A. Smith ◽  
Ennessa G. Curry ◽  
R. Eric Blue ◽  
Christine Roden ◽  
Samantha E. R. Dundon ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Muscle Development ◽  
Rna Binding ◽  
Fragile X ◽  
Protein Isoforms ◽  
List Type ◽  
Tissue Specific ◽  
Intrinsically Disordered ◽  
Multiple Protein

SUMMARYFragile-X mental retardation autosomal homolog-1 (FXR1) is a muscle-enriched RNA-binding protein. FXR1 depletion is perinatally lethal in mice, Xenopus, and zebrafish; however, the mechanisms driving these phenotypes remain unclear. The FXR1 gene undergoes alternative splicing, producing multiple protein isoforms and mis-splicing has been implicated in disease. Furthermore, mutations that cause frameshifts in muscle-specific isoforms result in congenital multi-minicore myopathy. We observed that FXR1 alternative splicing is pronounced in the serine and arginine-rich intrinsically-disordered domain; these domains are known to promote biomolecular condensation. Here, we show that tissue-specific splicing of fxr1 is required for Xenopus development and alters the disordered domain of FXR1. FXR1 isoforms vary in the formation of RNA-dependent biomolecular condensates in cells and in vitro. This work shows that regulation of tissue-specific splicing can influence FXR1 condensates in muscle development and how mis-splicing promotes disease.HIGHLIGHTSThe muscle-specific exon 15 impacts FXR1 functionsAlternative splicing of FXR1 is tissue- and developmental stage specificFXR1 forms RNA-dependent condensatesSplicing regulation changes FXR1 condensate properties

Alternative splicing generates a smaller assortment of CaV2.1 transcripts in cerebellar Purkinje cells than in the cerebellum

2006 ◽  
Vol 24 (2) ◽  
pp. 86-96 ◽  
Author(s):  
Srinivasan Kanumilli ◽  
Elizabeth W. Tringham ◽  
C. Elizabeth Payne ◽  
Jonathan R. B. Dupere ◽  
Kanamarlapudi Venkateswarlu ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Calcium Channels ◽  
Purkinje Cells ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Brain Regions ◽  
Rt Pcr ◽  
Brain Neurons ◽  
Cerebellar Purkinje Cells ◽  
P Type

P/Q-type calcium channels control many calcium-driven functions in the brain. The CACNA1A gene encoding the pore-forming CaV2.1 (α1A) subunit of P/Q-type channels undergoes alternative splicing at multiple loci. This results in channel variants with different phenotypes. However, the combinatorial patterns of alternative splice events at two or more loci, and hence the diversity of CaV2.1 transcripts, are incompletely defined for specific brain regions and types of brain neurons. Using RT-PCR and splice variant-specific primers, we have identified multiple CaV2.1 transcript variants defined by different pairs of splice events in the cerebellum of adult rat. We have uncovered new splice variations between exons 28 and 34 (some of which predict a premature stop codon) and a new variation in exon 47 (which predicts a novel extended COOH-terminus). Single cell RT-PCR reveals that each individual cerebellar Purkinje neuron also expresses multiple alternative CaV2.1 transcripts, but the assortment is smaller than in the cerebellum. Two of these variants encode different extended COOH-termini which are not the same as those previously reported in Purkinje cells of the mouse. Our patch-clamp recordings show that calcium channel currents in the soma and dendrites of Purkinje cells are largely inhibited by a concentration of ω-agatoxin IVA selective for P-type over Q-type channels, suggesting that the different transcripts may form phenotypic variants of P-type calcium channels in Purkinje cells. These results expand the known diversity of CaV2.1 transcripts in cerebellar Purkinje cells, and propose the selective expression of distinct assortments of CaV2.1 transcripts in different brain neurons and species.

scholarly journals Alternative Splicing: Recent Insights into Mechanisms and Functional Roles

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2327
Author(s):  
Claudia Ghigna ◽  
Maria Paola Paronetto
Keyword(s):  
Alternative Splicing ◽  
Human Cells ◽  
Protein Isoforms ◽  
Primary Transcript ◽  
Functional Roles ◽  
Multiple Protein

Alternative splicing generates multiple protein isoforms from one primary transcript and represents one of the major drivers of proteomic diversity in human cells [...]

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