scholarly journals The RNA-Protein Interactome of Differentiated Kidney Tubular Epithelial Cells

2019 ◽  
Vol 30 (4) ◽  
pp. 564-576 ◽  
Author(s):  
Michael Ignarski ◽  
Constantin Rill ◽  
Rainer W.J. Kaiser ◽  
Madlen Kaldirim ◽  
René Neuhaus ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Collecting Duct ◽  
Rna Stability ◽  
Tubular Epithelial Cells ◽  
Experimental Conditions ◽  
Rna Molecules ◽  
Ciliated Epithelial Cells

BackgroundRNA-binding proteins (RBPs) are fundamental regulators of cellular biology that affect all steps in the generation and processing of RNA molecules. Recent evidence suggests that regulation of RBPs that modulate both RNA stability and translation may have a profound effect on the proteome. However, regulation of RBPs in clinically relevant experimental conditions has not been studied systematically.MethodsWe used RNA interactome capture, a method for the global identification of RBPs to characterize the global RNA‐binding proteome (RBPome) associated with polyA-tailed RNA species in murine ciliated epithelial cells of the inner medullary collecting duct. To study regulation of RBPs in a clinically relevant condition, we analyzed hypoxia-associated changes of the RBPome.ResultsWe identified >1000 RBPs that had been previously found using other systems. In addition, we found a number of novel RBPs not identified by previous screens using mouse or human cells, suggesting that these proteins may be specific RBPs in differentiated kidney epithelial cells. We also found quantitative differences in RBP-binding to mRNA that were associated with hypoxia versus normoxia.ConclusionsThese findings demonstrate the regulation of RBPs through environmental stimuli and provide insight into the biology of hypoxia-response signaling in epithelial cells in the kidney. A repository of the RBPome and proteome in kidney tubular epithelial cells, derived from our findings, is freely accessible online, and may contribute to a better understanding of the role of RNA-protein interactions in kidney tubular epithelial cells, including the response of these cells to hypoxia.

scholarly journals Towards an Ideal In Cell Hybridization-Based Strategy to Discover Protein Interactomes of Selected RNA Molecules

2022 ◽  
Vol 23 (2) ◽  
pp. 942
Author(s):  
Michele Spiniello ◽  
Mark Scalf ◽  
Amelia Casamassimi ◽  
Ciro Abbondanza ◽  
Lloyd M. Smith
Keyword(s):  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Cell Hybridization ◽  
Rna Molecules ◽  
Cellular Context ◽  
Protein Binders ◽  
Non Coding Rnas ◽  
Technical Features ◽  
Experimental Strategies

RNA-binding proteins are crucial to the function of coding and non-coding RNAs. The disruption of RNA–protein interactions is involved in many different pathological states. Several computational and experimental strategies have been developed to identify protein binders of selected RNA molecules. Amongst these, ‘in cell’ hybridization methods represent the gold standard in the field because they are designed to reveal the proteins bound to specific RNAs in a cellular context. Here, we compare the technical features of different ‘in cell’ hybridization approaches with a focus on their advantages, limitations, and current and potential future applications.

scholarly journals The Landscape of RNA-Protein Interactions in Plants: Approaches and Current Status

2021 ◽  
Vol 22 (6) ◽  
pp. 2845
Author(s):  
Vesper Burjoski ◽  
Anireddy S. N. Reddy
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Translational Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Current Status ◽  
Protein Coding ◽  
Rna Molecules ◽  
Cellular Processes ◽  
Binding Domains

RNAs transmit information from DNA to encode proteins that perform all cellular processes and regulate gene expression in multiple ways. From the time of synthesis to degradation, RNA molecules are associated with proteins called RNA-binding proteins (RBPs). The RBPs play diverse roles in many aspects of gene expression including pre-mRNA processing and post-transcriptional and translational regulation. In the last decade, the application of modern techniques to identify RNA–protein interactions with individual proteins, RNAs, and the whole transcriptome has led to the discovery of a hidden landscape of these interactions in plants. Global approaches such as RNA interactome capture (RIC) to identify proteins that bind protein-coding transcripts have led to the identification of close to 2000 putative RBPs in plants. Interestingly, many of these were found to be metabolic enzymes with no known canonical RNA-binding domains. Here, we review the methods used to analyze RNA–protein interactions in plants thus far and highlight the understanding of plant RNA–protein interactions these techniques have provided us. We also review some recent protein-centric, RNA-centric, and global approaches developed with non-plant systems and discuss their potential application to plants. We also provide an overview of results from classical studies of RNA–protein interaction in plants and discuss the significance of the increasingly evident ubiquity of RNA–protein interactions for the study of gene regulation and RNA biology in plants.

PlncRNADB: A Repository of Plant lncRNAs and lncRNA-RBP Protein Interactions

2019 ◽  
Vol 14 (7) ◽  
pp. 621-627 ◽  
Author(s):  
Youhuang Bai ◽  
Xiaozhuan Dai ◽  
Tiantian Ye ◽  
Peijing Zhang ◽  
Xu Yan ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Populus Trichocarpa ◽  
Noncoding Rnas ◽  
Reference Database ◽  
Protein Coding ◽  
Arabidopsis Lyrata ◽  
User Friendly

Background: Long noncoding RNAs (lncRNAs) are endogenous noncoding RNAs, arbitrarily longer than 200 nucleotides, that play critical roles in diverse biological processes. LncRNAs exist in different genomes ranging from animals to plants. Objective: PlncRNADB is a searchable database of lncRNA sequences and annotation in plants. Methods: We built a pipeline for lncRNA prediction in plants, providing a convenient utility for users to quickly distinguish potential noncoding RNAs from protein-coding transcripts. Results: More than five thousand lncRNAs are collected from four plant species (Arabidopsis thaliana, Arabidopsis lyrata, Populus trichocarpa and Zea mays) in PlncRNADB. Moreover, our database provides the relationship between lncRNAs and various RNA-binding proteins (RBPs), which can be displayed through a user-friendly web interface. Conclusion: PlncRNADB can serve as a reference database to investigate the lncRNAs and their interaction with RNA-binding proteins in plants. The PlncRNADB is freely available at http://bis.zju.edu.cn/PlncRNADB/.

scholarly journals Compendium of Methods to Uncover RNA-Protein Interactions In Vivo

2021 ◽  
Vol 4 (1) ◽  
pp. 22
Author(s):  
Mrinmoyee Majumder ◽  
Viswanathan Palanisamy
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Expression Patterns ◽  
Control Of Gene Expression ◽  
Regulatory Pathways ◽  
Advantages And Disadvantages ◽  
Protein Nucleic Acid

Control of gene expression is critical in shaping the pro-and eukaryotic organisms’ genotype and phenotype. The gene expression regulatory pathways solely rely on protein–protein and protein–nucleic acid interactions, which determine the fate of the nucleic acids. RNA–protein interactions play a significant role in co- and post-transcriptional regulation to control gene expression. RNA-binding proteins (RBPs) are a diverse group of macromolecules that bind to RNA and play an essential role in RNA biology by regulating pre-mRNA processing, maturation, nuclear transport, stability, and translation. Hence, the studies aimed at investigating RNA–protein interactions are essential to advance our knowledge in gene expression patterns associated with health and disease. Here we discuss the long-established and current technologies that are widely used to study RNA–protein interactions in vivo. We also present the advantages and disadvantages of each method discussed in the review.

scholarly journals RNA-Centric Approaches to Profile the RNA–Protein Interaction Landscape on Selected RNAs

2021 ◽  
Vol 7 (1) ◽  
pp. 11 ◽  
Author(s):  
André P. Gerber
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Regulatory Networks ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Protein Complexes ◽  
Cell Protein ◽  
Transcriptional Regulatory Networks ◽  
Technological Advances

RNA–protein interactions frame post-transcriptional regulatory networks and modulate transcription and epigenetics. While the technological advances in RNA sequencing have significantly expanded the repertoire of RNAs, recently developed biochemical approaches combined with sensitive mass-spectrometry have revealed hundreds of previously unrecognized and potentially novel RNA-binding proteins. Nevertheless, a major challenge remains to understand how the thousands of RNA molecules and their interacting proteins assemble and control the fate of each individual RNA in a cell. Here, I review recent methodological advances to approach this problem through systematic identification of proteins that interact with particular RNAs in living cells. Thereby, a specific focus is given to in vivo approaches that involve crosslinking of RNA–protein interactions through ultraviolet irradiation or treatment of cells with chemicals, followed by capture of the RNA under study with antisense-oligonucleotides and identification of bound proteins with mass-spectrometry. Several recent studies defining interactomes of long non-coding RNAs, viral RNAs, as well as mRNAs are highlighted, and short reference is given to recent in-cell protein labeling techniques. These recent experimental improvements could open the door for broader applications and to study the remodeling of RNA–protein complexes upon different environmental cues and in disease.

scholarly journals LPI-HyADBS: a hybrid framework for lncRNA-protein interaction prediction integrating feature selection and classification

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Liqian Zhou ◽  
Qi Duan ◽  
Xiongfei Tian ◽  
He Xu ◽  
Jianxin Tang ◽  
...  
Keyword(s):  
Feature Selection ◽  
Protein Interactions ◽  
Cross Validation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Biological Information ◽  
Hybrid Framework ◽  
Protein Interaction Prediction ◽  
Single Dataset ◽  
Feature Selection Approach

Abstract Background Long noncoding RNAs (lncRNAs) have dense linkages with a plethora of important cellular activities. lncRNAs exert functions by linking with corresponding RNA-binding proteins. Since experimental techniques to detect lncRNA-protein interactions (LPIs) are laborious and time-consuming, a few computational methods have been reported for LPI prediction. However, computation-based LPI identification methods have the following limitations: (1) Most methods were evaluated on a single dataset, and researchers may thus fail to measure their generalization ability. (2) The majority of methods were validated under cross validation on lncRNA-protein pairs, did not investigate the performance under other cross validations, especially for cross validation on independent lncRNAs and independent proteins. (3) lncRNAs and proteins have abundant biological information, how to select informative features need to further investigate. Results Under a hybrid framework (LPI-HyADBS) integrating feature selection based on AdaBoost, and classification models including deep neural network (DNN), extreme gradient Boost (XGBoost), and SVM with a penalty Coefficient of misclassification (C-SVM), this work focuses on finding new LPIs. First, five datasets are arranged. Each dataset contains lncRNA sequences, protein sequences, and an LPI network. Second, biological features of lncRNAs and proteins are acquired based on Pyfeat. Third, the obtained features of lncRNAs and proteins are selected based on AdaBoost and concatenated to depict each LPI sample. Fourth, DNN, XGBoost, and C-SVM are used to classify lncRNA-protein pairs based on the concatenated features. Finally, a hybrid framework is developed to integrate the classification results from the above three classifiers. LPI-HyADBS is compared to six classical LPI prediction approaches (LPI-SKF, LPI-NRLMF, Capsule-LPI, LPI-CNNCP, LPLNP, and LPBNI) on five datasets under 5-fold cross validations on lncRNAs, proteins, lncRNA-protein pairs, and independent lncRNAs and independent proteins. The results show LPI-HyADBS has the best LPI prediction performance under four different cross validations. In particular, LPI-HyADBS obtains better classification ability than other six approaches under the constructed independent dataset. Case analyses suggest that there is relevance between ZNF667-AS1 and Q15717. Conclusions Integrating feature selection approach based on AdaBoost, three classification techniques including DNN, XGBoost, and C-SVM, this work develops a hybrid framework to identify new linkages between lncRNAs and proteins.

Development of an RNA-protein crosslinker to capture protein interactions with diverse RNA structures in cells

RNA ◽  
2021 ◽  
pp. rna.078896.121
Author(s):  
Yan Han ◽  
Xuzhen Guo ◽  
Tiancai Zhang ◽  
Jiangyun Wang ◽  
Keqiong Ye
Keyword(s):  
Protein Interactions ◽  
Rna Structure ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
High Specificity ◽  
Ester Group ◽  
Yeast Cells ◽  
Primary Amines ◽  
Low Efficiency ◽  
In Cells

Characterization of RNA-protein interaction is fundamental for understanding metabolism and function of RNA. UV crosslinking has been widely used to map the targets of RNA-binding proteins, but is limited by low efficiency, requirement for zero-distance contact and biases for single-stranded RNA structure and certain residues of RNA and protein. Here, we report the development of an RNA-protein crosslinker (AMT-NHS) composed of a psoralen derivative and an N-hydroxysuccinimide ester group, which react with RNA bases and primary amines of protein, respectively. We show that AMT-NHS can penetrate into living yeast cells and crosslink Cbf5 to H/ACA snoRNAs with high specificity. The crosslinker induced different crosslinking patterns than UV and targeted both single- and double-stranded regions of RNA. The crosslinker provides a new tool to capture diverse RNA-protein interactions in cells.

A synthetic RNA editing factor edits its target site in chloroplasts and bacteria

2020 ◽  
Author(s):  
Santana Royan ◽  
Bernard Gutmann ◽  
Catherine Colas des Francs-Small ◽  
Suvi Honkanen ◽  
Jason Schmidberger ◽  
...  
Keyword(s):  
Rna Editing ◽  
Protein Interactions ◽  
Rna Binding ◽  
Catalytic Domain ◽  
Rna Binding Proteins ◽  
Cytidine Deaminase ◽  
Land Plant ◽  
Target Sequence ◽  
Pentatricopeptide Repeat

Abstract Targeted cytidine to uridine RNA editing is a widespread phenomenon throughout the land plant lineage. Members of the pentatricopeptide repeat (PPR) protein family act as the specificity factors in this process. These proteins consist of helix-turn-helix domains, each of which recognises a single RNA nucleotide following a well-elucidated code. A cytidine deaminase-like domain (present at the C-terminus of some PPR editing factors or provided in trans via protein-protein interactions) is the catalytic domain in the process. The huge expansion of the PPR superfamily in land plants provides the sequence variation required for design of novel consensus-based RNA-binding proteins. We used this approach to construct a synthetic RNA editing factor designed to target one of the two sites in the Arabidopsis chloroplast transcriptome naturally recognised by the RNA editing factor CHLOROPLAST BIOGENESIS 19 (CLB19). We show that this designed editing factor specifically recognises the target sequence in in vitro binding assays and can partially complement a clb19 mutant. The designed factor is specific for the target rpoA site and does not recognise or edit the other site recognised by CLB19 in the clpP1 transcript. We show that the designed editing factor can function equally specifically in the bacterium E. coli, and shows some activity even in the absence of the editing cofactors that are often required for natural editing factor activity in plants. This study serves as a successful pilot into the design and application of programmable RNA editing factors based on plant PPR proteins.

scholarly journals The structural basis for RNA selectivity by the IMP family of RNA-binding proteins

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Jeetayu Biswas ◽  
Vivek L. Patel ◽  
Varun Bhaskar ◽  
Jeffrey A. Chao ◽  
Robert H. Singer ◽  
...  
Keyword(s):  
Neural Development ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Stability ◽  
Underlying Mechanism ◽  
Structural Basis ◽  
General Mechanism ◽  
Binding Domains ◽  
Variable Loops

Abstract The IGF2 mRNA-binding proteins (ZBP1/IMP1, IMP2, IMP3) are highly conserved post-transcriptional regulators of RNA stability, localization and translation. They play important roles in cell migration, neural development, metabolism and cancer cell survival. The knockout phenotypes of individual IMP proteins suggest that each family member regulates a unique pool of RNAs, yet evidence and an underlying mechanism for this is lacking. Here, we combine systematic evolution of ligands by exponential enrichment (SELEX) and NMR spectroscopy to demonstrate that the major RNA-binding domains of the two most distantly related IMPs (ZBP1 and IMP2) bind to different consensus sequences and regulate targets consistent with their knockout phenotypes and roles in disease. We find that the targeting specificity of each IMP is determined by few amino acids in their variable loops. As variable loops often differ amongst KH domain paralogs, we hypothesize that this is a general mechanism for evolving specificity and regulation of the transcriptome.

scholarly journals RNA regulons are essential in intestinal homeostasis

2019 ◽  
Vol 316 (1) ◽  
pp. G197-G204 ◽  
Author(s):  
Louis R. Parham ◽  
Patrick A. Williams ◽  
Priya Chatterji ◽  
Kelly A. Whelan ◽  
Kathryn E. Hamilton
Keyword(s):  
Epithelial Cells ◽  
Rna Binding ◽  
Environmental Changes ◽  
Cell Function ◽  
Rna Binding Proteins ◽  
Cell Types ◽  
Intestinal Epithelial ◽  
Cell Dynamics ◽  
Functional Roles ◽  
Proliferating Cell

Intestinal epithelial cells are among the most rapidly proliferating cell types in the human body. There are several different subtypes of epithelial cells, each with unique functional roles in responding to the ever-changing environment. The epithelium’s ability for rapid and customized responses to environmental changes requires multitiered levels of gene regulation. An emerging paradigm in gastrointestinal epithelial cells is the regulation of functionally related mRNA families, or regulons, via RNA-binding proteins (RBPs). RBPs represent a rapid and efficient mechanism to regulate gene expression and cell function. In this review, we will provide an overview of intestinal epithelial RBPs and how they contribute specifically to intestinal epithelial stem cell dynamics. In addition, we will highlight key gaps in knowledge in the global understanding of RBPs in gastrointestinal physiology as an opportunity for future studies.

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