scholarly journals RNA splicing in human disease and in the clinic

2017 ◽  
Vol 131 (5) ◽  
pp. 355-368 ◽  
Author(s):  
Diana Baralle ◽  
Emanuele Buratti
Keyword(s):  
Human Disease ◽  
Rna Splicing ◽  
High Throughput Sequencing ◽  
Genomic Sequence ◽  
Disease Onset ◽  
Mrna Splicing ◽  
Sequence Variant ◽  
Disease Mutations

Defects at the level of the pre-mRNA splicing process represent a major cause of human disease. Approximately 15–50% of all human disease mutations have been shown to alter functioning of basic and auxiliary splicing elements. These elements are required to ensure proper processing of pre-mRNA splicing molecules, with their disruption leading to misprocessing of the pre-mRNA molecule and disease. The splicing process is a complex process, with much still to be uncovered before we are able to accurately predict whether a reported genomic sequence variant (GV) represents a splicing-associated disease mutation or a harmless polymorphism. Furthermore, even when a mutation is correctly identified as affecting the splicing process, there still remains the difficulty of providing an exact evaluation of the potential impact on disease onset, severity and duration. In this review, we provide a brief overview of splicing diagnostic methodologies, from in silico bioinformatics approaches to wet lab in vitro and in vivo systems to evaluate splicing efficiencies. In particular, we provide an overview of how the latest developments in high-throughput sequencing can be applied to the clinic, and are already changing clinical approaches.

scholarly journals GPKOW is essential for pre-mRNA splicing in vitro and suppresses splicing defect caused by dominant-negative DHX16 mutation in vivo

2014 ◽  
Vol 34 (6) ◽  
Author(s):  
Shengbing Zang ◽  
Ting-Yu Lin ◽  
Xinji Chen ◽  
Marieta Gencheva ◽  
Alain N. S. Newo ◽  
...  
Keyword(s):  
Rna Splicing ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Mrna Splicing ◽  
Dominant Negative ◽  
Splicing Defect ◽  
Similarity And Difference

Using biochemical, mutation, and cellular analyses, we characterized important domains involved in the functionality of a RNA-binding protein in RNA splicing. We also showed the similarity and difference between yeast and human counterparts.

scholarly journals The methyltransferase SETD2 couples transcription and splicing by engaging mRNA processing factors through its SHI domain

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Saikat Bhattacharya ◽  
Michaella J. Levy ◽  
Ning Zhang ◽  
Hua Li ◽  
Laurence Florens ◽  
...  
Keyword(s):  
Rna Splicing ◽  
Rna Binding ◽  
Mrna Processing ◽  
Key Factors ◽  
Pol Ii ◽  
Mrna Transcripts ◽  
Hnrnp Protein ◽  
Processing Factors

AbstractHeterogeneous ribonucleoproteins (hnRNPs) are RNA binding molecules that are involved in key processes such as RNA splicing and transcription. One such hnRNP protein, hnRNP L, regulates alternative splicing (AS) by binding to pre-mRNA transcripts. However, it is unclear what factors contribute to hnRNP L-regulated AS events. Using proteomic approaches, we identified several key factors that co-purify with hnRNP L. We demonstrate that one such factor, the histone methyltransferase SETD2, specifically interacts with hnRNP L in vitro and in vivo. This interaction occurs through a previously uncharacterized domain in SETD2, the SETD2-hnRNP Interaction (SHI) domain, the deletion of which, leads to a reduced H3K36me3 deposition. Functionally, SETD2 regulates a subset of hnRNP L-targeted AS events. Our findings demonstrate that SETD2, by interacting with Pol II as well as hnRNP L, can mediate the crosstalk between the transcription and the splicing machinery.

scholarly journals EJE Prize 2013: Regulation of aldosterone secretion: from physiology to disease

2013 ◽  
Vol 168 (6) ◽  
pp. R85-R93 ◽  
Author(s):  
Felix Beuschlein
Keyword(s):  
High Throughput Sequencing ◽  
Secondary Hypertension ◽  
Small Sample ◽  
Cellular Systems ◽  
Common Disease ◽  
Aldosterone Secretion ◽  
Technical Developments ◽  
Starting Point

Arterial hypertension is a major cardiovascular risk factor that affects between 10 and 40% of the population in industrialized countries. Primary aldosteronism (PA) is the most common form of secondary hypertension with an estimated prevalence of around 10% in referral centers and 4% in a primary care setting. Despite its high prevalence until recently, the underlying genetic and molecular basis of this common disease had remained largely obscure. Over the past decade, a number of insights have been achieved that have relied onin vitrocellular systems, wild-type and genetically modifiedin vivomodels, as well as clinical studies in well-characterized patient populations. This progress has been made possible by a number of independent technical developments including that of specific hormone assays that allow measurement in small sample volumes as well as genetic techniques that enable high-throughput sequencing of a large number of samples. Furthermore, animal models have provided important insights into the physiology of aldosterone regulation that have served as a starting point for investigation of mechanisms involved in autonomous aldosterone secretion. Finally, national and international networks that have built up registries and biobanks have been instrumental in fostering translational research endeavors in PA. Therefore, it is to be expected that in the near future, further pathophysiological mechanisms that result in autonomous aldosterone secretion will be unraveled.

scholarly journals Genomic Sequence of Rhesus Cytomegalovirus 180.92: Insights into the Coding Potential of Rhesus Cytomegalovirus

2006 ◽  
Vol 80 (8) ◽  
pp. 4179-4182 ◽  
Author(s):  
Pierre Rivailler ◽  
Amitinder Kaur ◽  
R. Paul Johnson ◽  
Fred Wang
Keyword(s):  
Genomic Sequence ◽  
Open Reading Frames ◽  
Rhesus Cytomegalovirus ◽  
Human Cmv ◽  
Coding Capacity ◽  
Coding Potential ◽  
Reading Frames ◽  
Genetic Rearrangements

ABSTRACT A pathogenic isolate of rhesus cytomegalovirus (rhCMV 180.92) was cloned, sequenced, and annotated. Comparisons with the published rhCMV 68.1 genome revealed 8 open reading frames (ORFs) in isolate 180.92 that are absent in 68.1, 10 ORFs in 68.1 that are absent in 180.92, and 34 additional ORFs that were not previously annotated. Most of the differences appear to be due to genetic rearrangements in both isolates from a region that is frequently altered in human CMV (hCMV) during in vitro passage. These results indicate that the rhCMV ORF repertoire is larger than previously recognized. Like hCMV, understanding of the complete coding capacity of rhCMV is complicated by genomic instability and may require comparisons with additional isolates in vitro and in vivo.

FKBP51 Modulates Hippocampal Size and Function in Post-translational Regulation of Parkin

2021 ◽  
Author(s):  
Bin Qiu ◽  
Zhaohui Zhong ◽  
Shawn Righter ◽  
Yuxue Xu ◽  
Jun Wang ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Translational Regulation ◽  
Disease Onset ◽  
Fold Increase ◽  
Knock Out ◽  
Fk506 Binding Protein ◽  
Protein Levels ◽  
And Function

Abstract FK506-binding protein 51 (encoded by Fkpb51) has been associated with stress-related mental illness. To identify its function, we studied the morphological consequences of Fkbp51 deletion. Artificial Intelligence-assist morphological analysis identified that Fkbp51 knock-out (KO) mice possess more elongated CA and DG but shorter in height in coronal section when compared to WT. Primary cultured Fkbp51 KO hippocampal neurons were shown to exhibit larger dendritic outgrowth than wild-type (WT) controls, pharmacological manipulation experiments suggest that this may occur through regulation of microtubule-associated protein. Both in vitro primary culture and in vivo labeling support that FKBP51 regulates microtubule-associated protein expression. Furthermore, in the absence of differences in mRNA expression, Fkbp51 KO hippocampus exhibited decreases in βIII-tubulin, MAP2, and Tau protein levels, but a greater than 2.5-fold increase in Parkin protein. Overexpression and knock-down FKBP51 demonstrated that FKBP51 negatively regulates Parkin in a dose-dependent and ubiquitin-mediated manner. These results indicate a potential novel post-translational regulatory of Parkin by FKBP51 and significance of their interaction on disease onset.

The phylogenetically invariant ACAGAGA and AGC sequences of U6 small nuclear RNA are more tolerant of mutation in human cells than in Saccharomyces cerevisiae

1993 ◽  
Vol 13 (9) ◽  
pp. 5377-5382
Author(s):  
B Datta ◽  
A M Weiner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transient Expression ◽  
Human Cells ◽  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
Transient Expression Assay ◽  
U6 Snrna ◽  
Nuclear Rna

U6 small nuclear RNA (snRNA) is the most highly conserved of the five spliceosomal snRNAs that participate in nuclear mRNA splicing. The proposal that U6 snRNA plays a key catalytic role in splicing [D. Brow and C. Guthrie, Nature (London) 337:14-15, 1989] is supported by the phylogenetic conservation of U6, the sensitivity of U6 to mutation, cross-linking of U6 to the vicinity of the 5' splice site, and genetic evidence for extensive base pairing between U2 and U6 snRNAs. We chose to mutate the phylogenetically invariant 41-ACAGAGA-47 and 53-AGC-55 sequences of human U6 because certain point mutations within the homologous regions of Saccharomyces cerevisiae U6 selectively block the first or second step of mRNA splicing. We found that both sequences are more tolerant to mutation in human cells (assayed by transient expression in vivo) than in S. cerevisiae (assayed by effects on growth or in vitro splicing). These differences may reflect different rate-limiting steps in the particular assays used or differential reliance on redundant RNA-RNA or RNA-protein interactions. The ability of mutations in U6 nucleotides A-45 and A-53 to selectively block step 2 of splicing in S. cerevisiae had previously been construed as evidence that these residues might participate directly in the second chemical step of splicing; an indirect, structural role seems more likely because the equivalent mutations have no obvious phenotype in the human transient expression assay.

scholarly journals CDK10 Regulates Gastric Cancer Metastasis By Inhibiting lncRNA-C5ORF42-5 Via Phosphorylation Level of AKT/ERK

2021 ◽  
Author(s):  
Zi-Jian Deng ◽  
Dong-Wen Chen ◽  
Xi-Jie Chen ◽  
Jia-Ming Fang ◽  
Liang Xv ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cancer Cells ◽  
Cell Line ◽  
High Throughput ◽  
Cancer Metastasis ◽  
High Throughput Sequencing ◽  
Gastric Cancer Cells ◽  
Related Proteins

Abstract Background: Gastric cancer is the fourth most common malignant disease. Both CDK10 and long noncoding RNAs (lncRNAs) have been found to exert biological functions in multiple cancers. However, it is still unclear whether CDK10 represses tumor progression in gastric cancer by reducing potential targeting lncRNAs.Methods: The functions of CDK10 and lncRNA-C5ORF42-5 in proliferation, invasion and migration were assessed by MTS assays, colony formation assays, cell cycle and apoptosis assays, Transwell assays, wound healing assays and animal experiments. We used high-throughput sequencing to confirm the existence of lncRNA-C5ORF42-5 and quantitative real-time PCR was used to evaluate lncRNA expression. Then, with RNA-seq sequencing as well as GO function and KEGG enrichment analysis, we identified the signaling pathways in which lncRNA-C5ORF42-5 was involved in gastric cancer. Finally, western blotting was used to identify the genes regulated by lncRNA-C5ORF42-5.Results: Our results showed that CDK10 is expressed at relatively low levels in gastric cancer cell lines and inhibits the progression of gastric cancer cells both in vitro and in vivo. Next, based on high-throughput sequencing, we identified a novel lncRNA, lncRNA-C5ORF42-5, in the stable CDK10-overexpressing cell line compared with the CDK-knockdown cell line and their controls. Additionally, we confirmed that lncRNA-C5ORF42-5 acts as an oncogene to promote metastasis in gastric cancer in vitro and in vivo. We then ascertained that lncRNA-C5ORF42-5 is a major contributor to the function of CDK10 in gastric cancer metastasis by upregulating lncRNA-C5ORF42-5 to reverse the effects of CDK10 overexpression. Finally, we explored the mechanism by which lncRNA-C5ORF42-5 overexpression affects gastric cancer cells to elucidate whether lncRNA-C5ORF42-5 may increase the activity of the SMAD pathway of BMP signaling and promote the expression of EMT-related proteins, such as E-cadherin. Additionally, overexpression of lncRNA-C5ORF42-5 affected the phosphorylation levels of AKT and ERK.Conclusion: Our findings suggest that CDK10 overexpression represses gastric cancer tumor progression by reducing lncRNA-C5ORF42-5 and hindering activation of the related proteins in metastatic signaling pathways, which provides new insight into developing effective therapeutic strategies in the treatment of metastatic gastric cancer.

scholarly journals Elucidation of the BMI1 interactome identifies novel regulatory roles in glioblastoma

NAR Cancer ◽  
2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Verónica Freire-Benéitez ◽  
Nicola Pomella ◽  
Thomas O Millner ◽  
Anaëlle A Dumas ◽  
Maria Victoria Niklison-Chirou ◽  
...  
Keyword(s):  
Chromatin Immunoprecipitation ◽  
Protein Composition ◽  
Mrna Splicing ◽  
Cholesterol Transport ◽  
Normal Brain ◽  
Regulatory Functions ◽  
Polycomb Repressive Complex 1 ◽  
Normal Brain Development

Abstract Glioblastoma (GBM) is the most common and aggressive intrinsic brain tumour in adults. Epigenetic mechanisms controlling normal brain development are often dysregulated in GBM. Among these, BMI1, a structural component of the Polycomb Repressive Complex 1 (PRC1), which promotes the H2AK119ub catalytic activity of Ring1B, is upregulated in GBM and its tumorigenic role has been shown in vitro and in vivo. Here, we have used protein and chromatin immunoprecipitation followed by mass spectrometry (MS) analysis to elucidate the protein composition of PRC1 in GBM and transcriptional silencing of defining interactors in primary patient-derived GIC lines to assess their functional impact on GBM biology. We identify novel regulatory functions in mRNA splicing and cholesterol transport which could represent novel targetable mechanisms in GBM.

scholarly journals Transcriptome Profiling of Panc-1 Spheroid Cells with Pancreatic Cancer Stem Cells Properties Cultured by a Novel 3D Semi-Solid System

2018 ◽  
Vol 47 (5) ◽  
pp. 2109-2125 ◽  
Author(s):  
Zhaocong Yang ◽  
Yanfeng Zhang ◽  
Tingting Tang ◽  
Qinhua Zhu ◽  
Wanyue Shi ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Culture System ◽  
High Throughput Sequencing ◽  
Semi Solid ◽  
Pancreatic Cancer Stem Cells ◽  
Solid System

Background/Aims: Pancreatic cancer remains one of the deadliest human malignancies, the lethality of which may be attributed to the presence of pancreatic cancer stem cells (PCSCs), a small subpopulation of cells existing within pancreatic tumor with high carcinogenesis. Therefore, it is crucial to establish an efficient enrichment and culture system of PCSCs and identify the key genes involved in the regulation of PCSCs. The three-dimensional (3D) liquid suspension mammosphere culture system has been established for enrichment and culture of PCSCs in vitro as the cell spheres are likely to originate from individual cell clone, but it has been challenged because the cell spheroids could be a result of cell aggregation. Methods: We optimized the existing culture system by adding methylcellulose to create a 3D semi-solid system which prevented the non-specific aggregation. Then we identified the CSC properties of Panc-1 spheroid cells cultured by this system by detecting the genes associated with stemness and by evaluation of the tumorigenicity in vitro and in vivo through invasion, migration and xenograft experiments methods. Subsequently, we performed high-throughput sequencing (HTS) of the Panc-1 spheroid cells. Results: We confirmed the PCSCs properties and high malignancy of the Panc-1 spheroid cells enriched by our novel 3D semi-solid system both in vitro and in vivo. Hundreds of mRNA, microRNA (miRNA) and dozens of long non-coding RNA (LncRNA) were identified to be differentially regulated in PCSCs-like Panc-1 spheroid cells compared with their parental cells by HTS. Conclusions: Our results demonstrate an efficient enrichment and culture system for Panc-1 spheroid cells with the PCSCs properties. The differentially expressed genes and their targets identified by the HTS of the Panc-1 spheroid cells can serve as new potential biomarkers for pancreatic cancer diagnosis and targeted therapy.

YY1 and RTCB in mouse uterine decidualization and embryo implantation

Reproduction ◽  
2021 ◽  
Author(s):  
Ran Li ◽  
Xiao-Tong Song ◽  
Si-Wei Guo ◽  
Na Zhao ◽  
Mei He ◽  
...  
Keyword(s):  
Early Pregnancy ◽  
Embryo Implantation ◽  
Mrna Splicing ◽  
Proximal Promoter ◽  
Mouse Uterus ◽  
Rna Ligase ◽  
Xbp1 Mrna ◽  
Transcription Factor Yy1

As a multifunctional transcription factor, YY1 regulates the expression of many genes essential for early embryonic development. RTCB is an RNA ligase that plays a role in tRNA maturation and Xbp1 mRNA splicing. YY1 can bind in vitro to the response element in the proximal promoter of Rtcb and regulate Rtcb promoter activity. However, the in vivo regulation and whether these two genes are involved in the mother-fetal dialogue during early pregnancy remain unclear. In this study, we validated that YY1 bound in vivo to the proximal promoter of Rtcb in mouse uterus of early pregnancy. Moreover, via building a variety of animal models, our study suggested that both YY1 and RTCB might play a role in mouse uterus decidualization and embryo implantation during early pregnancy.

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