scholarly journals Structural Transition, Function and Dysfunction of TDP-43 in Neurodegenerative Diseases

2019 ◽  
Vol 73 (5) ◽  
pp. 380-390 ◽  
Author(s):  
Tariq Afroz ◽  
Manuela Pérez-Berlanga ◽  
Magdalini Polymenidou
Keyword(s):  
Neurodegenerative Diseases ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Low Complexity ◽  
Transition Function ◽  
Common Mechanism ◽  
Cellular Functions

Altered cellular localization and pathologic aggregation of RNA binding proteins (RPBs) containing low complexity regions (LCRs) is a hallmark of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Given the importance of RBPs in maintaining a healthy RNA homeostasis, a common mechanism in disease progression is the loss of RNA-related cellular functions. In this review, we summarize and discuss the knowledge gained in the recent years on the molecular mechanisms of TDP-43 proteinopathies that comprise a set of neurodegenerative diseases characterized by the mislocalization and aggregation of the RNA-binding protein TDP-43. Based on biophysical, biochemical and in vivo data, we highlight pathways that are misregulated early in disease and contribute to its progression, thereby representing attractive therapeutic targets.

scholarly journals Challenges and perspectives for structural biology of lncRNAs—the example of the Xist lncRNA A-repeats

2019 ◽  
Vol 11 (10) ◽  
pp. 845-859 ◽  
Author(s):  
Alisha N Jones ◽  
Michael Sattler
Keyword(s):  
High Resolution ◽  
Structural Biology ◽  
Recent Progress ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Structural Features ◽  
Molecular Features

Abstract Following the discovery of numerous long non-coding RNA (lncRNA) transcripts in the human genome, their important roles in biology and human disease are emerging. Recent progress in experimental methods has enabled the identification of structural features of lncRNAs. However, determining high-resolution structures is challenging as lncRNAs are expected to be dynamic and adopt multiple conformations, which may be modulated by interaction with protein binding partners. The X-inactive specific transcript (Xist) is necessary for X inactivation during dosage compensation in female placental mammals and one of the best-studied lncRNAs. Recent progress has provided new insights into the domain organization, molecular features, and RNA binding proteins that interact with distinct regions of Xist. The A-repeats located at the 5′ end of the transcript are of particular interest as they are essential for mediating silencing of the inactive X chromosome. Here, we discuss recent progress with elucidating structural features of the Xist lncRNA, focusing on the A-repeats. We discuss the experimental and computational approaches employed that have led to distinct structural models, likely reflecting the intrinsic dynamics of this RNA. The presence of multiple dynamic conformations may also play an important role in the formation of the associated RNPs, thus influencing the molecular mechanism underlying the biological function of the Xist A-repeats. We propose that integrative approaches that combine biochemical experiments and high-resolution structural biology in vitro with chemical probing and functional studies in vivo are required to unravel the molecular mechanisms of lncRNAs.

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

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.

scholarly journals Positional motif analysis reveals the extent of specificity of protein-RNA interactions observed by CLIP

2021 ◽  
Author(s):  
Klara Kuret ◽  
Aram Gustav Amalietti ◽  
Jernej Ule
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Data Interpretation ◽  
Low Complexity ◽  
Motif Analysis ◽  
Link Type ◽  
Motif Enrichment ◽  
The Impact

AbstractBackgroundCrosslinking and immunoprecipitation (CLIP) is a method used to identify in vivo RNA– protein binding sites on a transcriptome-wide scale. With the increasing amounts of available data for RNA-binding proteins (RBPs), it is important to understand to what degree the enriched motifs specify the RNA binding profiles of RBPs in cells.ResultsWe develop positionally-enriched k-mer analysis (PEKA), a computational tool for efficient analysis of enriched motifs from individual CLIP datasets, which minimises the impact of technical and regional genomic biases by internal data normalisation. We cross-validate PEKA with mCross, and show that background correction by size-matched input doesn’t generally improve the specificity of detected motifs. We identify motif classes with common enrichment patterns across eCLIP datasets and across RNA regions, while also observing variations in the specificity and the extent of motif enrichment across eCLIP datasets, between variant CLIP protocols, and between CLIP and in vitro binding data. Thereby we gain insights into the contributions of technical and regional genomic biases to the enriched motifs, and find how motif enrichment features relate to the domain composition and low-complexity regions (LCRs) of the studied proteins.ConclusionsOur study provides insights into the overall contributions of regional binding preferences, protein domains and LCRs to the specificity of protein-RNA interactions, and shows the value of cross-motif and cross-RBP comparison for data interpretation. Our results are presented for exploratory analysis via an online platform in an RBP-centric and motif-centric manner (https://imaps.goodwright.com/apps/peka/). PEKA is available from https://github.com/ulelab/peka.

scholarly journals Prevention of EloR/KhpA heterodimerization by introduction of site-specific amino acid substitutions renders the essential elongasome protein PBP2b redundant in Streptococcus pneumoniae

2018 ◽  
Author(s):  
Anja Ruud Winther ◽  
Morten Kjos ◽  
Gro Anita Stamsås ◽  
Leiv Sigve Håvarstein ◽  
Daniel Straume
Keyword(s):  
Cell Division ◽  
Cell Elongation ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Cell Region ◽  
Protein Protein Interaction

AbstractThe RNA binding proteins EloR and KhpA are important components of the regulatory network that controls and coordinates cell elongation and division in S. pneumoniae. Loss of either protein reduce cell length, and makes the essential elongasome proteins PBP2b and RodA dispensable. It has been shown previously in formaldehyde crosslinking experiments that EloR co-precipitates with KhpA, indicating that they form a complex in vivo. In the present study, we used 3D modeling and site directed mutagenesis in combination with protein crosslinking to further study the relationship between EloR and KhpA. Protein-protein interaction studies demonstrated that KhpA forms homodimers and that KhpA in addition binds strongly to the KH-II domain of EloR. Site directed mutagenesis identified isoleucine 61 (I61) as crucial for KhpA homodimerization. When substituting I61 with phenylalanine, KhpA lost the ability to homodimerize, while it still interacted strongly with EloR. In contrast, both homo- and heterodimerization were lost when I61 was substituted with tyrosine. By expressing these KhpA versions in S. pneumoniae, we were able to show that disruption of EloR/KhpA heterodimerization makes the elongasome redundant in S. pneumoniae. Of note, loss of KhpA homodimerization did not give rise to this phenotype, demonstrating that the EloR/KhpA complex is crucial for regulating the activity of the elongasome. In support of this conclusion, we found that localization of KhpA to the pneumococcal mid-cell region depends on its interaction with EloR. Furthermore, we found that the EloR/KhpA complex co-localizes with FtsZ throughout the cell cycle.ImportanceTo ensure correct cell division, bacteria need to monitor the progression of cell division and coordinate the activities of cell division proteins accordingly. Understanding the molecular mechanisms behind these regulatory systems is of high academic interest and might facilitate the development of new therapeutics and strategies to combat pathogens. EloR and KhpA form a heterodimer that is part of a signaling pathway controlling cell elongation in the human pathogen S. pneumoniae. Here we have identified amino acids that are crucial for EloR/KhpA heterodimerization, and demonstrated that disruption of the EloR/KhpA interaction renders the cells independent of a functional elongasome. Furthermore, we found the EloR/KhpA complex to co-localize with the division ring (FtsZ) during cell division.

Modelling FUSopathies: focus on protein aggregation

2013 ◽  
Vol 41 (6) ◽  
pp. 1613-1617 ◽  
Author(s):  
Tatyana A. Shelkovnikova
Keyword(s):  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Pathological Process ◽  
Model Organisms ◽  
In Vivo Models ◽  
Fused In Sarcoma ◽  
Amyloid Aggregates ◽  
Irreversible Aggregation

The discovery of a causative link between dysfunction of a number of RNA-binding proteins with prion-like domains and the development of certain (neuro)degenerative diseases has completely changed our perception of molecular mechanisms instigating pathological process in these disorders. Irreversible aggregation of these proteins is a crucial pathogenic event delineating a type of proteinopathy. FUS (fused in sarcoma) is a prototypical member of the class, and studies into the causes and consequences of FUSopathies have been instrumental in characterizing the processes leading to deregulation of RNA metabolism in neurodegeneration. In vivo models of FUSopathy have provided critical insights into the mechanisms of FUS toxicity and clues on the role of non-amyloid aggregates, which are hallmarks of these diseases. The present review summarizes the data on FUS aggregation signatures in available model organisms on the basis of overexpression of FUS variants.

scholarly journals TriPepSVM - de novo prediction of RNA-binding proteins based on short amino acid motifs

2018 ◽  
Author(s):  
Annkatrin Bressin ◽  
Roman Schulte-Sasse ◽  
Davide Figini ◽  
Erika C Urdaneta ◽  
Benedikt M Beckmann ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
De Novo ◽  
Rna Binding Proteins ◽  
Low Complexity ◽  
Structural Features ◽  
Support Vector ◽  
String Kernels ◽  
Rna Binders

In recent years hundreds of novel RNA-binding proteins (RBPs) have been identified leading to the discovery of novel RNA-binding domains (RBDs). Furthermore, unstructured or disordered low-complexity regions of RBPs have been identified to play an important role in interactions with nucleic acids. However, these advances in understanding RBPs are limited mainly to eukaryotic species and we only have limited tools to faithfully predict RNA-binders from bacteria. Here, we describe a support vector machine (SVM)-based method, called TriPepSVM, for the classification of RNA-binding proteins and non-RBPs. TriPepSVM applies string kernels to directly handle protein sequences using tri-peptide frequencies. Testing the method in human and bacteria, we find that several RBP-enriched tripeptides occur more often in structurally disordered regions of RBPs. TriPepSVM outperforms existing applications, which consider classical structural features of RNA-binding or homology, in the task of RBP prediction in both human and bacteria. Finally, we predict 66 novel RBPs in Salmonella Typhimurium and validate the bacterial proteins ClpX, DnaJ and UbiG to associate with RNA in vivo.

scholarly journals RNA Proximity Labeling: A New Detection Tool for RNA–Protein Interactions

Molecules ◽  
2021 ◽  
Vol 26 (8) ◽  
pp. 2270
Author(s):  
Ronja Weissinger ◽  
Lisa Heinold ◽  
Saira Akram ◽  
Ralf-Peter Jansen ◽  
Orit Hermesh
Keyword(s):  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Protein Complexes ◽  
Affinity Purification ◽  
Cellular Functions ◽  
Cellular Compartments ◽  
Rna Protein Complexes

Multiple cellular functions are controlled by the interaction of RNAs and proteins. Together with the RNAs they control, RNA interacting proteins form RNA protein complexes, which are considered to serve as the true regulatory units for post-transcriptional gene expression. To understand how RNAs are modified, transported, and regulated therefore requires specific knowledge of their interaction partners. To this end, multiple techniques have been developed to characterize the interaction between RNAs and proteins. In this review, we briefly summarize the common methods to study RNA–protein interaction including crosslinking and immunoprecipitation (CLIP), and aptamer- or antisense oligonucleotide-based RNA affinity purification. Following this, we focus on in vivo proximity labeling to study RNA–protein interactions. In proximity labeling, a labeling enzyme like ascorbate peroxidase or biotin ligase is targeted to specific RNAs, RNA-binding proteins, or even cellular compartments and uses biotin to label the proteins and RNAs in its vicinity. The tagged molecules are then enriched and analyzed by mass spectrometry or RNA-Seq. We highlight the latest studies that exemplify the strength of this approach for the characterization of RNA protein complexes and distribution of RNAs in vivo.

scholarly journals Molecular functions of nuclear actin in transcription

2006 ◽  
Vol 172 (7) ◽  
pp. 967-971 ◽  
Author(s):  
Piergiorgio Percipalle ◽  
Neus Visa
Keyword(s):  
Rna Polymerase Ii ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
In Vivo Studies ◽  
Macromolecular Assemblies ◽  
In Vivo Experiments ◽  
Specific Subset

Actin is not only a major cytoskeletal component in all eukaryotic cells but also a nuclear protein that plays a role in gene transcription. We put together data from in vitro and in vivo experiments that begin to provide insights into the molecular mechanisms by which actin functions in transcription. Recent studies performed in vitro have suggested that actin, in direct contact with the transcription apparatus, is required in an early step of transcription that is common to all three eukaryotic RNA polymerases. In addition, there is evidence from in vivo studies that actin is involved in the transcription elongation of class II genes. In this case, actin is bound to a specific subset of premessenger RNA binding proteins, and the actin–messenger RNP complex may constitute a molecular platform for recruitment of histone-modifying enzymes. We discuss a general model for actin in RNA polymerase II transcription whereby actin works as a conformational switch in conjunction with specific adaptors to facilitate the remodeling of large macromolecular assemblies at the promoter and along the active gene.

scholarly journals RNA-binding proteins distinguish between similar sequence motifs to promote targeted deadenylation by Ccr4-Not

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Michael W Webster ◽  
James AW Stowell ◽  
Lori A Passmore
Keyword(s):  
Mrna Stability ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Low Complexity ◽  
Dissociation Rate ◽  
Sequence Motifs ◽  
Similar Sequence ◽  
Mrna Targets

The Ccr4-Not complex removes mRNA poly(A) tails to regulate eukaryotic mRNA stability and translation. RNA-binding proteins contribute to specificity by interacting with both Ccr4-Not and target mRNAs, but this is not fully understood. Here, we reconstitute accelerated and selective deadenylation of RNAs containing AU-rich elements (AREs) and Pumilio-response elements (PREs). We find that the fission yeast homologues of Tristetraprolin/TTP and Pumilio/Puf (Zfs1 and Puf3) interact with Ccr4-Not via multiple regions within low-complexity sequences, suggestive of a multipartite interface that extends beyond previously defined interactions. Using a two-color assay to simultaneously monitor poly(A) tail removal from different RNAs, we demonstrate that Puf3 can distinguish between RNAs of very similar sequence. Analysis of binding kinetics reveals that this is primarily due to differences in dissociation rate constants. Consequently, motif quality is a major determinant of mRNA stability for Puf3 targets in vivo and can be used for the prediction of mRNA targets.

scholarly journals Active role of free RNA during the early phase of proteostasis stress

2019 ◽  
Author(s):  
Marion Alriquet ◽  
Adrían Martínez-Limón ◽  
Gerd Hanspach ◽  
Martin Hengesbach ◽  
Gian G. Tartaglia ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Active Form ◽  
Active Role ◽  
Structural Features ◽  
Cellular Functions ◽  
Granule Formation

ABSTRACTTransient sequestration of proteins and RNA is an essential principle of cellular reaction to stress. Compared to polypeptides, less is known about the role of RNA released from polysomes during acute proteostasis stress. Using quantitative mass spectrometry, we identified a set of proteins assembled by free RNA in the heat-shocked mammalian cytosol. RNA-associated proteins displayed higher disorder and larger size, which supports the role of multivalent interactions during the initial phase of the RNA granule formation. Structural features of the free RNA interactors defined them as a subset of RNA-binding proteins. The interactome contained preferentially the active form of eIF2α. The interaction between assembled proteins in vivo required RNA. The reconstitution of the association process in vitro indicated to the multimolecular basis for the increased binding to RNA upon heat shock in the cytosol. Our results reveal how free RNA can participate in reorganization of cellular functions during proteostasis stress.

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