scholarly journals The mechanism of Ccr4-Not recruitment to specific mRNAs involves sequence-selective tethering by RNA-binding proteins

2018 ◽  
Author(s):  
Michael W Webster ◽  
James A W Stowell ◽  
Lori A Passmore
Keyword(s):  
Mrna Stability ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Low Complexity ◽  
Dissociation Rate ◽  
Multiple Regions ◽  
Specific Mrnas ◽  
Dissociation Rate Constants

AbstractThe Ccr4-Not complex removes mRNA poly(A) tails to regulate eukaryotic mRNA stability and translation. RNA-binding proteins contribute to specificity 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) act as molecular tethers: They recruit Ccr4-Not via multiple regions within low-complexity sequences, and bind specific RNA sequences via RNA-binding domains. 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. This is primarily due to differences in the dissociation rate constants. As a result, motif quality is a major determinant of mRNA stability for Puf3 targets in vivo. Together, we provide new insight into the selective deadenylation of specific mRNAs by Ccr4-Not, and the prediction of targeted mRNAs.

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 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.

RNA-Binding Proteins and Translation in Neurodegenerative Disease

2020 ◽  
Author(s):  
Kent E. Duncan
Keyword(s):  
Neurodegenerative Diseases ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Potential Contribution ◽  
Fused In Sarcoma ◽  
Specific Mrnas ◽  
Ataxia Syndrome ◽  
Research Frontiers

Both RNA-binding proteins (RBPs) and translation are increasingly implicated in several neurodegenerative diseases, but their specific roles in promoting disease are not yet fully defined. This chapter critically evaluates the evidence that altered translation of specific mRNAs mediated by RNA-binding proteins plays an important role in driving specific neurodegenerative diseases. First, diseases are discussed where a causal role for RNA-binding proteins in disease appears solid, but whether this involves altered translation is less clear. The main foci here are TAR DNA-binding protein (TDP-43) and fused in sarcoma (FUS) in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Subsequently, diseases are presented where altered translation is believed to contribute, but involvement of RNA-binding proteins is less clear. These include Huntington’s and other repeat expansion disorders such as fragile X tremor/ataxia syndrome (FXTAS), where repeat-induced non-AUG-initiated (RAN) translation is a focus. The potential contribution of both canonical and non-canonical RBPs to altered translation in Parkinson’s disease is discussed. The chapter closes by proposing key research frontiers for the field to explore and outlining methodological advances that could help to address them.

scholarly journals Identification of mRNAs Associated with αCP2-Containing RNP Complexes

2003 ◽  
Vol 23 (19) ◽  
pp. 7055-7067 ◽  
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.

scholarly journals RNA Binding Proteins and the Pathogenesis of Frontotemporal Lobar Degeneration

2019 ◽  
Vol 14 (1) ◽  
pp. 469-495 ◽  
Author(s):  
Jeffrey W. Hofmann ◽  
William W. Seeley ◽  
Eric J. Huang
Keyword(s):  
Frontotemporal Lobar Degeneration ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Low Complexity ◽  
Compelling Evidence ◽  
Abnormal Protein ◽  
Early Onset Dementia ◽  
Cellular Pathways ◽  
Liquid Liquid Phase Separation

Frontotemporal dementia is a group of early onset dementia syndromes linked to underlying frontotemporal lobar degeneration (FTLD) pathology that can be classified based on the formation of abnormal protein aggregates involving tau and two RNA binding proteins, TDP-43 and FUS. Although elucidation of the mechanisms leading to FTLD pathology is in progress, recent advances in genetics and neuropathology indicate that a majority of FTLD cases with proteinopathy involving RNA binding proteins show highly congruent genotype–phenotype correlations. Specifically, recent studies have uncovered the unique properties of the low-complexity domains in RNA binding proteins that can facilitate liquid–liquid phase separation in the formation of membraneless organelles. Furthermore, there is compelling evidence that mutations in FTLD genes lead to dysfunction in diverse cellular pathways that converge on the endolysosomal pathway, autophagy, and neuroinflammation. Together, these results provide key mechanistic insights into the pathogenesis and potential therapeutic targets of FTLD.

scholarly journals SSMART: Sequence-structure motif identification for RNA-binding proteins

2018 ◽  
Author(s):  
Alina Munteanu ◽  
Neelanjan Mukherjee ◽  
Uwe Ohler
Keyword(s):  
Binding Sites ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Sequence Motif ◽  
Primary Sequence ◽  
Sequence Structure ◽  
Rna Targets

AbstractMotivationRNA-binding proteins (RBPs) regulate every aspect of RNA metabolism and function. There are hundreds of RBPs encoded in the eukaryotic genomes, and each recognize its RNA targets through a specific mixture of RNA sequence and structure properties. For most RBPs, however, only a primary sequence motif has been determined, while the structure of the binding sites is uncharacterized.ResultsWe developed SSMART, an RNA motif finder that simultaneously models the primary sequence and the structural properties of the RNA targets sites. The sequence-structure motifs are represented as consensus strings over a degenerate alphabet, extending the IUPAC codes for nucleotides to account for secondary structure preferences. Evaluation on synthetic data showed that SSMART is able to recover both sequence and structure motifs implanted into 3‘UTR-like sequences, for various degrees of structured/unstructured binding sites. In addition, we successfully used SSMART on high-throughput in vivo and in vitro data, showing that we not only recover the known sequence motif, but also gain insight into the structural preferences of the RBP.AvailabilitySSMART is freely available at https://ohlerlab.mdc-berlin.de/software/SSMART_137/[email protected]

scholarly journals TRIBE editing reveals specific mRNA targets of eIF4E-BP in Drosophila and in mammals

2020 ◽  
Author(s):  
Hua Jin ◽  
Daxiang Na ◽  
Reazur Rahman ◽  
Weijin Xu ◽  
Allegra Fieldsend ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Growth Control ◽  
Ribosome Profiling ◽  
Translational Efficiency ◽  
Mtor Inhibition ◽  
Mrna Targets ◽  
Specific Mrnas ◽  
Specific Mrna

Abstract4E-BP (eIF4E-BP) represses translation initiation by binding to the 5’cap-binding protein eIF4E and inhibiting its activity. Although 4E-BP has been shown to be important in growth control, stress response, cancer, neuronal activity and mammalian circadian rhythms, it is not understood how it preferentially represses a subset of mRNAs. We successfully used hyperTRIBE (Targets of RNA-binding proteins identified by editing) to identify in vivo 4E-BP mRNA targets in both Drosophila and mammals under conditions known to activate 4E-BP. The protein associates with specific mRNAs, and ribosome profiling data show that mTOR inhibition changes the translational efficiency of 4E-BP TRIBE targets compared to non-targets. In both systems, these targets have specific motifs and are enriched in translation-related pathways, which correlate well with the known activity of 4E-BP and suggest that it modulates the binding specificity of eIF4E and contributes to mTOR translational specificity.

scholarly journals A unique ribonucleoprotein complex assembles preferentially on ecdysone-responsive sites in Drosophila melanogaster.

1993 ◽  
Vol 13 (9) ◽  
pp. 5323-5330 ◽  
Author(s):  
S A Amero ◽  
M J Matunis ◽  
E L Matunis ◽  
J W Hockensmith ◽  
G Raychaudhuri ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Polytene Chromosomes ◽  
Cell Extract ◽  
Sequence Motifs ◽  
Drosophila Cell ◽  
Rnase Digestion

The protein on ecdysone puffs (PEP) is associated preferentially with active ecdysone-inducible puffs on Drosophila polytene chromosomes and contains sequence motifs characteristic of transcription factors and RNA-binding proteins (S. A. Amero, S. C. R. Elgin, and A. L. Beyer, Genes Dev. 5:188-200, 1991). PEP is associated with RNA in vivo, as demonstrated here by the sensitivity of PEP-specific chromosomal immunostaining in situ to RNase digestion and by the immunopurification of PEP in Drosophila cell extract with heterogeneous nuclear ribonucleoprotein (hnRNP) complexes. As revealed by sequential immunostaining, PEP is found on a subset of chromosomal sites bound by the HRB (heterogeneous nuclear RNA-binding) proteins, which are basic Drosophila hnRNPs. These observations lead us to suggest that a unique, PEP-containing hnRNP complex assembles preferentially on the transcripts of ecdysone-regulated genes in Drosophila melanogaster presumably to expedite the transcription and/or processing of these transcripts.

scholarly journals RNA-binding proteins with prion-like domains in health and disease

2017 ◽  
Vol 474 (8) ◽  
pp. 1417-1438 ◽  
Author(s):  
Alice Ford Harrison ◽  
James Shorter
Keyword(s):  
Phase Transitions ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Low Complexity ◽  
Fused In Sarcoma ◽  
Heterogeneous Nuclear Ribonucleoproteins ◽  
Health And Disease ◽  
Lateral Sclerosis

Approximately 70 human RNA-binding proteins (RBPs) contain a prion-like domain (PrLD). PrLDs are low-complexity domains that possess a similar amino acid composition to prion domains in yeast, which enable several proteins, including Sup35 and Rnq1, to form infectious conformers, termed prions. In humans, PrLDs contribute to RBP function and enable RBPs to undergo liquid–liquid phase transitions that underlie the biogenesis of various membraneless organelles. However, this activity appears to render RBPs prone to misfolding and aggregation connected to neurodegenerative disease. Indeed, numerous RBPs with PrLDs, including TDP-43 (transactivation response element DNA-binding protein 43), FUS (fused in sarcoma), TAF15 (TATA-binding protein-associated factor 15), EWSR1 (Ewing sarcoma breakpoint region 1), and heterogeneous nuclear ribonucleoproteins A1 and A2 (hnRNPA1 and hnRNPA2), have now been connected via pathology and genetics to the etiology of several neurodegenerative diseases, including amyotrophic lateral sclerosis, frontotemporal dementia, and multisystem proteinopathy. Here, we review the physiological and pathological roles of the most prominent RBPs with PrLDs. We also highlight the potential of protein disaggregases, including Hsp104, as a therapeutic strategy to combat the aberrant phase transitions of RBPs with PrLDs that likely underpin neurodegeneration.

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