scholarly journals Gain-of-function cardiomyopathic mutations in RBM20 rewire splicing regulation and re-distribute ribonucleoprotein aggregates within processing bodies

2021 ◽  
Author(s):  
Aidan M Fenix ◽  
Yuichiro Miyaoka ◽  
Alessandro Bertero ◽  
Steven Blue ◽  
Matthew J Spindler ◽  
...  
Keyword(s):  
Rna Binding ◽  
Acute Stress ◽  
Rna Binding Proteins ◽  
Super Resolution ◽  
Circular Rna ◽  
Binding Motif ◽  
Missense Mutations ◽  
Gain Of Function ◽  
Processing Bodies ◽  
Heart Pathology

RNA binding motif protein 20 (RBM20) is a key regulator of alternative splicing in the heart, and its mutation leads to malignant dilated cardiomyopathy (DCM). To understand the mechanism of RBM20-associated DCM, we engineered isogenic human induced pluripotent stem cells (iPSCs) with heterozygous or homozygous DCM-associated missense mutations in RBM20 (R636S) as well as RBM20 knockout (KO) iPSCs. iPSC-derived engineered heart tissues made from these cell lines recapitulated contractile dysfunction of RBM20-associated DCM and revealed greater dysfunction with missense mutations than KO. Analysis of RBM20 RNA binding by eCLIP revealed a gain-of-function preference of mutant RBM20 for 3′ UTR sequences that are shared with amyotrophic lateral sclerosis (ALS) and processing-body associated RNA binding proteins (FUS, DDX6). Deep RNA sequencing revealed that the RBM20 R636S mutant has unique gene, splicing, polyadenylation and circular RNA defects that differ from RBM20 KO, impacting distinct cardiac signaling pathways. Splicing defects specific to KO or R636S mutations were supported by data from R636S gene-edited pig hearts and eCLIP. Super-resolution microscopy verified that mutant RBM20 maintains limited nuclear localization potential; rather, the mutant protein associates with cytoplasmic processing bodies (DDX6) under basal conditions, and with stress granules (G3BP1) following acute stress. Taken together, our results highlight a novel pathogenic mechanism in cardiac disease through splicing-dependent and -independent pathways that are likely to mediate differential contractile phenotypes and stress-associated heart pathology.

scholarly journals Gain-of-function cardiomyopathic mutations in RBM20 rewire splicing regulation and re-distribute ribonucleoprotein granules within processing bodies

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Aidan M. Fenix ◽  
Yuichiro Miyaoka ◽  
Alessandro Bertero ◽  
Steven M. Blue ◽  
Matthew J. Spindler ◽  
...  
Keyword(s):  
Rna Binding ◽  
Acute Stress ◽  
Rna Binding Proteins ◽  
Super Resolution ◽  
Circular Rna ◽  
Missense Mutations ◽  
Gain Of Function ◽  
Processing Bodies ◽  
Gene Splicing ◽  
Processing Body

AbstractMutations in the cardiac splicing factor RBM20 lead to malignant dilated cardiomyopathy (DCM). To understand the mechanism of RBM20-associated DCM, we engineered isogenic iPSCs with DCM-associated missense mutations in RBM20 as well as RBM20 knockout (KO) iPSCs. iPSC-derived engineered heart tissues made from these cell lines recapitulate contractile dysfunction of RBM20-associated DCM and reveal greater dysfunction with missense mutations than KO. Analysis of RBM20 RNA binding by eCLIP reveals a gain-of-function preference of mutant RBM20 for 3′ UTR sequences that are shared with amyotrophic lateral sclerosis (ALS) and processing-body associated RNA binding proteins (FUS, DDX6). Deep RNA sequencing reveals that the RBM20 R636S mutant has unique gene, splicing, polyadenylation and circular RNA defects that differ from RBM20 KO. Super-resolution microscopy verifies that mutant RBM20 maintains very limited nuclear localization potential; rather, the mutant protein associates with cytoplasmic processing bodies (DDX6) under basal conditions, and with stress granules (G3BP1) following acute stress. Taken together, our results highlight a pathogenic mechanism in cardiac disease through splicing-dependent and -independent pathways.

scholarly journals Identification of Novel RNA Binding Proteins Influencing Circular RNA Expression in Hepatocellular Carcinoma

2021 ◽  
Vol 22 (14) ◽  
pp. 7477
Author(s):  
Rok Razpotnik ◽  
Petra Nassib ◽  
Tanja Kunej ◽  
Damjana Rozman ◽  
Tadeja Režen
Keyword(s):  
Hepatocellular Carcinoma ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Regulatory Protein ◽  
Circular Rna ◽  
Differentially Expressed ◽  
Circular Rnas ◽  
Bioinformatics Analyses ◽  
Published Evidence

Circular RNAs (circRNAs) are increasingly recognized as having a role in cancer development. Their expression is modified in numerous cancers, including hepatocellular carcinoma (HCC); however, little is known about the mechanisms of their regulation. The aim of this study was to identify regulators of circRNAome expression in HCC. Using publicly available datasets, we identified RNA binding proteins (RBPs) with enriched motifs around the splice sites of differentially expressed circRNAs in HCC. We confirmed the binding of some of the candidate RBPs using ChIP-seq and eCLIP datasets in the ENCODE database. Several of the identified RBPs were found to be differentially expressed in HCC and/or correlated with the overall survival of HCC patients. According to our bioinformatics analyses and published evidence, we propose that NONO, PCPB2, PCPB1, ESRP2, and HNRNPK are candidate regulators of circRNA expression in HCC. We confirmed that the knocking down the epithelial splicing regulatory protein 2 (ESRP2), known to be involved in the maintenance of the adult liver phenotype, significantly changed the expression of candidate circRNAs in a model HCC cell line. By understanding the systemic changes in transcriptome splicing, we can identify new proteins involved in the molecular pathways leading to HCC development and progression.

scholarly journals FUS driven circCNOT6L biogenesis in mouse and human spermatozoa supports zygote development

2021 ◽  
Author(s):  
Teresa Chioccarelli ◽  
Geppino Falco ◽  
Donato Cappetta ◽  
Antonella De Angelis ◽  
Luca Roberto ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Rna Binding ◽  
Cannabinoid Receptor ◽  
Rna Binding Proteins ◽  
Embryonic Stem ◽  
Circular Rna ◽  
Physical Interaction ◽  
Knock Out ◽  
Maternal Contribution ◽  
Zygote Development

AbstractCircular RNA (circRNA) biogenesis requires a backsplicing reaction, promoted by inverted repeats in cis-flanking sequences and trans factors, such as RNA-binding proteins (RBPs). Among these, FUS plays a key role. During spermatogenesis and sperm maturation along the epididymis such a molecular mechanism has been poorly explored. With this in mind, we chose circCNOT6L as a study case and wild-type (WT) as well as cannabinoid receptor type-1 knock-out (Cb1−/−) male mice as animal models to analyze backsplicing mechanisms. Our results suggest that spermatozoa (SPZ) have an endogenous skill to circularize mRNAs, choosing FUS as modulator of backsplicing and under CB1 stimulation. A physical interaction between FUS and CNOT6L as well as a cooperation among FUS, RNA Polymerase II (RNApol2) and Quaking (QKI) take place in SPZ. Finally, to gain insight into FUS involvement in circCNOT6L biogenesis, FUS expression was reduced through RNA interference approach. Paternal transmission of FUS and CNOT6L to oocytes during fertilization was then assessed by using murine unfertilized oocytes (NF), one-cell zygotes (F) and murine oocytes undergoing parthenogenetic activation (PA) to exclude a maternal contribution. The role of circCNOT6L as an active regulator of zygote transition toward the 2-cell-like state was suggested using the Embryonic Stem Cell (ESC) system. Intriguingly, human SPZ exactly mirror murine SPZ.

scholarly journals The effective function of circular RNA in colorectal cancer

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Mandana Ameli-Mojarad ◽  
Melika Ameli-Mojarad ◽  
Mahrooyeh Hadizadeh ◽  
Chris Young ◽  
Hosna Babini ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Circular Rna ◽  
Circular Rnas ◽  
Colorectal Tumorigenesis ◽  
Non Coding Rnas ◽  
Mirna Sponges ◽  
Mechanisms Of Development

AbstractColorectal cancer (CRC) is the 3rd most common type of cancer worldwide. Late detection plays role in one-third of annual mortality due to CRC. Therefore, it is essential to find a precise and optimal diagnostic and prognostic biomarker for the identification and treatment of colorectal tumorigenesis. Covalently closed, circular RNAs (circRNAs) are a class of non-coding RNAs, which can have the same function as microRNA (miRNA) sponges, as regulators of splicing and transcription, and as interactors with RNA-binding proteins (RBPs). Therefore, circRNAs have been investigated as specific targets for diagnostic and prognostic detection of CRC. These non-coding RNAs are also linked to metastasis, proliferation, differentiation, migration, angiogenesis, apoptosis, and drug resistance, illustrating the importance of understanding their involvement in the molecular mechanisms of development and progression of CRC. In this review, we present a detailed summary of recent findings relating to the dysregulation of circRNAs and their potential role in CRC.

Molecular dissection of a genome defense pathway in Neurospora crassa

2015 ◽  
Author(s):  
◽  
Erin C. Boone
Keyword(s):  
Neurospora Crassa ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Nuclear Periphery ◽  
Bimolecular Fluorescence Complementation ◽  
Proper Function ◽  
Rnai Pathway ◽  
Binding Motif ◽  
Perinuclear Region ◽  
A Genome

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Meiotic silencing by unpaired DNA (MSUD) is an RNA interference (RNAi) pathway in Neurospora crassa that detects genes without a homologous partner and silences them for the duration of sexual development. In this study, we have further elucidated the function of known MSUD proteins, identified novel proteins that are required for MSUD, and demonstrated the conservation of RNAi-related processes at the nuclear periphery. We began by showing SAD-2 is crucial for the localization of other MSUD proteins in the perinuclear region. These data suggest that SAD-2 works as a scaffold protein and that proper function of MSUD, like other germline RNAi-like systems, is reliant on the presence of silencing proteins in the perinuclear region. An MSUD suppression assay identified two novel MSUD proteins, SAD-Y and SAD-B'. Even though SAD-Y and its homologs contain a conserved putative RNA- binding motif, they have yet to be assigned to a biochemical pathway. Our work here has linked silencing to SAD-Y-like proteins. SAD-Y has been shown to interact with other MSUD factors in both the nucleus and at the nuclear periphery. SAD-B's homolog has been found in the nuage, an epicenter for RNA-binding proteins involved in post-transcriptional regulation for Drosophila germline cells. SAD-B interacts with core MSUD proteins and has an especially intimate association with SMS-2, which requires it for localization. Furthermore, bimolecular fluorescence complementation (BiFC) revealed that SAD-B' interacts with a Golgi retrograde transport protein and an autophagy marker protein, suggesting the importance of the endomembrane system in this RNAi process.

Roles for myosin Va in RNA transport and turnover

2012 ◽  
Vol 40 (6) ◽  
pp. 1416-1420 ◽  
Author(s):  
Mary W. McCaffrey ◽  
Andrew J. Lindsay
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Class V ◽  
Processing Bodies ◽  
Surface Receptors ◽  
P Bodies ◽  
Myosin Va ◽  
Myosin Vb ◽  
Actin Cables ◽  
The Brain

Mammals express three class V myosins. Myosin Va is widely expressed, but enriched in the brain, testes and melanocytes, myosin Vb is expressed ubiquitously, and myosin Vc is believed to be epithelium-specific. Myosin Va is the best characterized of the three and plays a key role in the transport of cargo to the plasma membrane. Its cargo includes cell-surface receptors, pigment and organelles such as the endoplasmic reticulum. It is also emerging that RNA and RNA-BPs (RNA-binding proteins) make up another class of myosin Va cargo. It has long been established that the yeast class V myosin, Myo4p, transports mRNAs along actin cables into the growing bud, and now several groups have reported a similar role for class V myosins in higher eukaryotes. Myosin Va has also been implicated in the assembly and maintenance of P-bodies (processing bodies), cytoplasmic foci that are involved in mRNA storage and degradation. The present review examines the evidence that myosin Va plays a role in the transport and turnover of mRNA.

scholarly journals The Regulatory Functions of Circular RNAs in Digestive System Cancers

Cancers ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 770 ◽  
Author(s):  
Xiao Yuan ◽  
Ya Yuan ◽  
Zhi He ◽  
Diyan Li ◽  
Bo Zeng ◽  
...  
Keyword(s):  
Digestive System ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Circular Rna ◽  
Research Progress ◽  
Circular Rnas ◽  
Biological Functions ◽  
Ribonucleic Acids ◽  
Mirna Sponges ◽  
And Biological Markers

Circular ribonucleic acids (circRNAs), which are a type of covalently closed circular RNA, are receiving increasing attention. An increasing amount of evidence suggests that circRNAs are involved in the biogenesis and development of multiple diseases such as digestive system cancers. Dysregulated circRNAs have been found to act as oncogenes or tumour suppressors in digestive system cancers. Moreover, circRNAs are related to ageing and a wide variety of processes in tumour cells, such as cell apoptosis, invasion, migration, and proliferation. Moreover, circRNAs can perform a remarkable multitude of biological functions, such as regulating splicing or transcription, binding RNA-binding proteins to enable function, acting as microRNA (miRNA) sponges, and undergoing translated into proteins. However, in digestive system cancers, circRNAs function mainly as miRNA sponges. Herein, we summarise the latest research progress on biological functions of circRNAs in digestive system cancers. This review serves as a synopsis of potential therapeutic targets and biological markers for digestive system cancer.

scholarly journals A comprehensive review of circRNA: from purification and identification to disease marker potential

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5503 ◽  
Author(s):  
Sheng Xu ◽  
LuYu Zhou ◽  
Murugavel Ponnusamy ◽  
LiXia Zhang ◽  
YanHan Dong ◽  
...  
Keyword(s):  
Noncoding Rna ◽  
High Stability ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Neurological Diseases ◽  
Circular Rna ◽  
Biological Functions ◽  
Disease Marker ◽  
Pathological Conditions ◽  
Microrna Sponge

Circular RNA (circRNA) is an endogenous noncoding RNA with a covalently closed cyclic structure. Based on their components, circRNAs are divided into exonic circRNAs, intronic circRNAs, and exon-intron circRNAs. CircRNAs have well-conserved sequences and often have high stability due to their resistance to exonucleases. Depending on their sequence, circRNAs are involved in different biological functions, including microRNA sponge activity, modulation of alternative splicing or transcription, interaction with RNA-binding proteins, and rolling translation, and are a derivative of pseudogenes. CircRNAs are involved in the development of a variety of pathological conditions, such as cardiovascular diseases, diabetes, neurological diseases, and cancer. Emerging evidence has shown that circRNAs are likely to be new potential clinical diagnostic markers or treatments for many diseases. Here we describe circRNA research methods and biological functions, and discuss the potential relationship between circRNAs and disease progression.

scholarly journals Posttranscriptional regulation of human endogenous retroviruses by RNA-binding motif protein 4, RBM4

2020 ◽  
Vol 117 (42) ◽  
pp. 26520-26530
Author(s):  
Amir K. Foroushani ◽  
Bryan Chim ◽  
Madeline Wong ◽  
Andre Rastegar ◽  
Patrick T. Smith ◽  
...  
Keyword(s):  
Human Genome ◽  
Posttranscriptional Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Ectopic Expression ◽  
Transcript Level ◽  
Endogenous Retroviruses ◽  
Host Protein ◽  
Binding Motif ◽  
Sequencing Data

The human genome encodes for over 1,500 RNA-binding proteins (RBPs), which coordinate regulatory events on RNA transcripts. Most studies of RBPs have concentrated on their action on host protein-encoding mRNAs, which constitute a minority of the transcriptome. A widely neglected subset of our transcriptome derives from integrated retroviral elements, termed endogenous retroviruses (ERVs), that comprise ∼8% of the human genome. Some ERVs have been shown to be transcribed under physiological and pathological conditions, suggesting that sophisticated regulatory mechanisms to coordinate and prevent their ectopic expression exist. However, it is unknown how broadly RBPs and ERV transcripts directly interact to provide a posttranscriptional layer of regulation. Here, we implemented a computational pipeline to determine the correlation of expression between individual RBPs and ERVs from single-cell or bulk RNA-sequencing data. One of our top candidates for an RBP negatively regulating ERV expression was RNA-binding motif protein 4 (RBM4). We used photoactivatable ribonucleoside-enhanced cross-linking and immunoprecipitation to demonstrate that RBM4 indeed bound ERV transcripts at CGG consensus elements. Loss of RBM4 resulted in an elevated transcript level of bound ERVs of the HERV-K and -H families, as well as increased expression of HERV-K envelope protein. We pinpointed RBM4 regulation of HERV-K to a CGG-containing element that is conserved in the LTRs of HERV-K-10, -K-11, and -K-20, and validated the functionality of this site using reporter assays. In summary, we systematically identified RBPs that may regulate ERV function and demonstrate a role for RBM4 in controlling ERV expression.

scholarly journals Biogenesis mechanisms of circular RNA can be categorized through feature extraction of a machine learning model

2019 ◽  
Vol 35 (23) ◽  
pp. 4867-4870
Author(s):  
Chengyu Liu ◽  
Yu-Chen Liu ◽  
Hsien-Da Huang ◽  
Wei Wang
Keyword(s):  
Feature Extraction ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Cpg Island ◽  
Cpg Islands ◽  
Circular Rna ◽  
Training Data ◽  
Supplementary Information ◽  
Circular Rnas ◽  
Genomic Locus

Abstract Motivation In recent years, multiple circular RNAs (circRNA) biogenesis mechanisms have been discovered. Although each reported mechanism has been experimentally verified in different circRNAs, no single biogenesis mechanism has been proposed that can universally explain the biogenesis of all tens of thousands of discovered circRNAs. Under the hypothesis that human circRNAs can be categorized according to different biogenesis mechanisms, we designed a contextual regression model trained to predict the formation of circular RNA from a random genomic locus on human genome, with potential biogenesis factors of circular RNA as the features of the training data. Results After achieving high prediction accuracy, we found through the feature extraction technique that the examined human circRNAs can be categorized into seven subgroups, according to the presence of the following sequence features: RNA editing sites, simple repeat sequences, self-chains, RNA binding protein binding sites and CpG islands within the flanking regions of the circular RNA back-spliced junction sites. These results support all of the previously reported biogenesis mechanisms of circRNA and solidify the idea that multiple biogenesis mechanisms co-exist for different subset of human circRNAs. Furthermore, we uncover a potential new links between circRNA biogenesis and flanking CpG island. We have also identified RNA binding proteins putatively correlated with circRNA biogenesis. Availability and implementation Scripts and tutorial are available at http://wanglab.ucsd.edu/star/circRNA. This program is under GNU General Public License v3.0. Supplementary information Supplementary data are available at Bioinformatics online.

Sign in / Sign up

Export Citation Format

Share Document