Transcriptome-wide organization of subcellular microenvironments revealed by ATLAS-Seq

Abstract Subcellular organization of RNAs and proteins is critical for cell function, but we still lack global maps and conceptual frameworks for how these molecules are localized in cells and tissues. Here, we introduce ATLAS-Seq, which generates transcriptomes and proteomes from detergent-free tissue lysates fractionated across a sucrose gradient. Proteomic analysis of fractions confirmed separation of subcellular compartments. Unexpectedly, RNAs tended to co-sediment with other RNAs in similar protein complexes, cellular compartments, or with similar biological functions. With the exception of those encoding secreted proteins, most RNAs sedimented differently than their encoded protein counterparts. To identify RNA binding proteins potentially driving these patterns, we correlated their sedimentation profiles to all RNAs, confirming known interactions and predicting new associations. Hundreds of alternative RNA isoforms exhibited distinct sedimentation patterns across the gradient, despite sharing most of their coding sequence. These observations suggest that transcriptomes can be organized into networks of co-segregating mRNAs encoding functionally related proteins and provide insights into the establishment and maintenance of subcellular organization.

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RBP2GO: a comprehensive pan-species database on RNA-binding proteins, their interactions and functions

Nucleic Acids Research ◽

10.1093/nar/gkaa1040 ◽

2020 ◽

Vol 49 (D1) ◽

pp. D425-D436 ◽

Cited By ~ 1

Author(s):

Maiwen Caudron-Herger ◽

Ralf E Jansen ◽

Elsa Wassmer ◽

Sven Diederichs

Keyword(s):

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

Protein Complexes ◽

Rapid Evolution ◽

Cellular Processes ◽

Advanced Search ◽

New Research ◽

Cellular Compartments ◽

User Friendly

Abstract RNA–protein complexes have emerged as central players in numerous key cellular processes with significant relevance in health and disease. To further deepen our knowledge of RNA-binding proteins (RBPs), multiple proteome-wide strategies have been developed to identify RBPs in different species leading to a large number of studies contributing experimentally identified as well as predicted RBP candidate catalogs. However, the rapid evolution of the field led to an accumulation of isolated datasets, hampering the access and comparison of their valuable content. Moreover, tools to link RBPs to cellular pathways and functions were lacking. Here, to facilitate the efficient screening of the RBP resources, we provide RBP2GO (https://RBP2GO.DKFZ.de), a comprehensive database of all currently available proteome-wide datasets for RBPs across 13 species from 53 studies including 105 datasets identifying altogether 22 552 RBP candidates. These are combined with the information on RBP interaction partners and on the related biological processes, molecular functions and cellular compartments. RBP2GO offers a user-friendly web interface with an RBP scoring system and powerful advanced search tools allowing forward and reverse searches connecting functions and RBPs to stimulate new research directions.

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

Molecules ◽

10.3390/molecules26082270 ◽

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.

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

Non-Coding RNA ◽

10.3390/ncrna7010011 ◽

2021 ◽

Vol 7 (1) ◽

pp. 11 ◽

Cited By ~ 1

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.

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Identification of mRNAs Associated with αCP2-Containing RNP Complexes

Molecular and Cellular Biology ◽

10.1128/mcb.23.19.7055-7067.2003 ◽

2003 ◽

Vol 23 (19) ◽

pp. 7055-7067 ◽

Cited By ~ 45

Author(s):

Shelly A. Waggoner ◽

Stephen A. Liebhaber

Keyword(s):

Binding Proteins ◽

Rna Binding ◽

Cell Function ◽

Rna Binding Proteins ◽

Translation Control ◽

Viral Mrnas ◽

Posttranscriptional Gene Regulation ◽

Direct Binding

ABSTRACT Posttranscriptional controls in higher eukaryotes are central to cell differentiation and developmental programs. These controls reflect sequence-specific interactions of mRNAs with one or more RNA binding proteins. The α-globin poly(C) binding proteins (αCPs) comprise a highly abundant subset of K homology (KH) domain RNA binding proteins and have a characteristic preference for binding single-stranded C-rich motifs. αCPs have been implicated in translation control and stabilization of multiple cellular and viral mRNAs. To explore the full contribution of αCPs to cell function, we have identified a set of mRNAs that associate in vivo with the major αCP2 isoforms. One hundred sixty mRNA species were consistently identified in three independent analyses of αCP2-RNP complexes immunopurified from a human hematopoietic cell line (K562). These mRNAs could be grouped into subsets encoding cytoskeletal components, transcription factors, proto-oncogenes, and cell signaling factors. Two mRNAs were linked to ceroid lipofuscinosis, indicating a potential role for αCP2 in this infantile neurodegenerative disease. Surprisingly, αCP2 mRNA itself was represented in αCP2-RNP complexes, suggesting autoregulatory control of αCP2 expression. In vitro analyses of representative target mRNAs confirmed direct binding of αCP2 within their 3′ untranslated regions. These data expand the list of mRNAs that associate with αCP2 in vivo and establish a foundation for modeling its role in coordinating pathways of posttranscriptional gene regulation.

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Current Understanding of Circular RNAs in Systemic Lupus Erythematosus

Frontiers in Immunology ◽

10.3389/fimmu.2021.628872 ◽

2021 ◽

Vol 12 ◽

Author(s):

Hongjiang Liu ◽

Yundong Zou ◽

Chen Chen ◽

Yundi Tang ◽

Jianping Guo

Keyword(s):

Systemic Lupus Erythematosus ◽

Lupus Erythematosus ◽

Rna Binding ◽

Rna Binding Proteins ◽

Protein Complexes ◽

Expression Patterns ◽

Regulation Of Gene Expression ◽

Current Understanding ◽

Circular Rnas ◽

Systemic Lupus

Systemic lupus erythematosus (SLE) is a common and potentially fatal autoimmune disease that affects multiple organs. To date, its etiology and pathogenesis remains elusive. Circular RNAs (circRNAs) are a novel class of endogenous non-coding RNAs with covalently closed loop structure. Growing evidence has demonstrated that circRNAs may play an essential role in regulation of gene expression and transcription by acting as microRNA (miRNA) sponges, impacting cell survival and proliferation by interacting with RNA binding proteins (RBPs), and strengthening mRNA stability by forming RNA-protein complexes duplex structures. The expression patterns of circRNAs exhibit tissue-specific and pathogenesis-related manner. CircRNAs have implicated in the development of multiple autoimmune diseases, including SLE. In this review, we summarize the characteristics, biogenesis, and potential functions of circRNAs, its impact on immune responses and highlight current understanding of circRNAs in the pathogenesis of SLE.

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Systematic discovery of endogenous human ribonucleoprotein complexes

10.1101/480061 ◽

2018 ◽

Cited By ~ 2

Author(s):

Anna L. Mallam ◽

Wisath Sae-Lee ◽

Jeffrey M. Schaub ◽

Fan Tu ◽

Anna Battenhouse ◽

...

Keyword(s):

Rna Binding ◽

Rna Binding Proteins ◽

Protein Complexes ◽

Embryonic Stem ◽

Human Protein ◽

Disease States ◽

Ribonucleoprotein Complexes ◽

Associated Proteins ◽

Domains Of Life ◽

Factor C

AbstractRNA-binding proteins (RBPs) play essential roles in biology and are frequently associated with human disease. While recent studies have systematically identified individual RBPs, their higher order assembly intoRibonucleoprotein (RNP) complexes has not been systematically investigated. Here, we describe a proteomics method for systematic identification of RNP complexes in human cells. We identify 1,428 protein complexes that associate with RNA, indicating that over 20% of known human protein complexes contain RNA. To explore the role of RNA in the assembly of each complex, we identify complexes that dissociate, change composition, or form stable protein-only complexes in the absence of RNA. Importantly, these data also provide specific novel insights into the function of well-studied protein complexes not previously known to associate with RNA, including replication factor C (RFC) and cytokinetic centralspindlin complex. Finally, we use our method to systematically identify cell-type specific RNA-associated proteins in mouse embryonic stem cells. We distribute these data as a resource, rna.MAP (rna.proteincomplexes.org) which provides a comprehensive dataset for the study of RNA-associated protein complexes. Our system thus provides a novel methodology for further explorations across human tissues and disease states, as well as throughout all domains of life.SummaryAn exploration of human protein complexes in the presence and absence of RNA reveals endogenous ribonucleoprotein complexes

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Multiple roles of heterogeneous nuclear ribonucleoprotein K during skeletal muscle cell differentiation

10.1101/2021.07.09.451593 ◽

2021 ◽

Author(s):

Keisuke Hitachi ◽

Yuri Kiyofuji ◽

Masashi Nakatani ◽

Kunihiro Tsuchida

Keyword(s):

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

Protein Complexes ◽

Myogenic Differentiation ◽

Cell Physiology ◽

Transcriptional Level ◽

Loss Of Function ◽

Independent Manner ◽

Multiple Functions

RNA-binding proteins (RBPs) regulate cell physiology via the formation of ribonucleic-protein complexes with coding and non-coding RNAs. RBPs have multiple functions in the same cells; however, the precise mechanism through which their pleiotropic functions are determined remains unknown. In this study, we revealed the multiple inhibitory functions of hnRNPK for myogenic differentiation. We first identified hnRNPK as a lncRNA Myoparr binding protein. Gain- and loss-of-function experiments showed that hnRNPK repressed the expression of myogenin at the transcriptional level via binding to Myoparr. Moreover, hnRNPK repressed the expression of a set of genes coding for aminoacyl-tRNA synthetases in a Myoparr-independent manner. Mechanistically, hnRNPK regulated the eIF2α/Atf4 pathway, one branch of the intrinsic pathways of the endoplasmic reticulum sensors, in differentiating myoblasts. Thus, our findings demonstrate that hnRNPK plays multiple lncRNA-dependent and -independent roles in the inhibition of myogenic differentiation, indicating that the analysis of lncRNA-binding proteins will be useful for elucidating both the physiological functions of lncRNAs and the multiple functions of RBPs.

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Quantitative Proteomics Reveals the Defense Response of Wheat against Puccinia striiformis f. sp. tritici

Scientific Reports ◽

10.1038/srep34261 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 12

Author(s):

Yuheng Yang ◽

Yang Yu ◽

Chaowei Bi ◽

Zhensheng Kang

Keyword(s):

Quantitative Proteomics ◽

Rna Binding ◽

Rna Binding Proteins ◽

Puccinia Striiformis ◽

Post Inoculation ◽

Control Groups ◽

Stimulus Response ◽

Wheat Gene ◽

Related Proteins ◽

And Control

Abstract Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is considered one of the most aggressive diseases to wheat production. In this study, we used an iTRAQ-based approach for the quantitative proteomic comparison of the incompatible Pst race CYR23 in infected and non-infected leaves of the wheat cultivar Suwon11. A total of 3,475 unique proteins were identified from three key stages of interaction (12, 24, and 48 h post-inoculation) and control groups. Quantitative analysis showed that 530 proteins were differentially accumulated by Pst infection (fold changes >1.5, p < 0.05). Among these proteins, 10.54% was classified as involved in the immune system process and stimulus response. Intriguingly, bioinformatics analysis revealed that a set of reactive oxygen species metabolism-related proteins, peptidyl–prolyl cis–trans isomerases (PPIases), RNA-binding proteins (RBPs), and chaperonins was involved in the response to Pst infection. Our results were the first to show that PPIases, RBPs, and chaperonins participated in the regulation of the immune response in wheat and even in plants. This study aimed to provide novel routes to reveal wheat gene functionality and better understand the early events in wheat–Pst incompatible interactions.

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RNA regulons are essential in intestinal homeostasis

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00403.2017 ◽

2019 ◽

Vol 316 (1) ◽

pp. G197-G204 ◽

Cited By ~ 2

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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The Regulatory Functions of Circular RNAs in Digestive System Cancers

Cancers ◽

10.3390/cancers12030770 ◽

2020 ◽

Vol 12 (3) ◽

pp. 770 ◽

Cited By ~ 4

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.

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