scholarly journals Multifunctional RNA-binding proteins influence mRNA abundance and translational efficiency of distinct sets of target genes

2021 ◽  
Vol 17 (12) ◽  
pp. e1009658
Author(s):  
Valentin Schneider-Lunitz ◽  
Jorge Ruiz-Orera ◽  
Norbert Hubner ◽  
Sebastiaan van Heesch
Keyword(s):  
Translational Regulation ◽  
Binding Proteins ◽  
Target Genes ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Translational Efficiency ◽  
Splicing Factors ◽  
Biological Processes ◽  
Differential Affinity

RNA-binding proteins (RBPs) can regulate more than a single aspect of RNA metabolism. We searched for such previously undiscovered multifunctionality within a set of 143 RBPs, by defining the predictive value of RBP abundance for the transcription and translation levels of known RBP target genes across 80 human hearts. This led us to newly associate 27 RBPs with cardiac translational regulation in vivo. Of these, 21 impacted both RNA expression and translation, albeit for virtually independent sets of target genes. We highlight a subset of these, including G3BP1, PUM1, UCHL5, and DDX3X, where dual regulation is achieved through differential affinity for target length, by which separate biological processes are controlled. Like the RNA helicase DDX3X, the known splicing factors EFTUD2 and PRPF8—all identified as multifunctional RBPs by our analysis—selectively influence target translation rates depending on 5’ UTR structure. Our analyses identify dozens of RBPs as being multifunctional and pinpoint potential novel regulators of translation, postulating unanticipated complexity of protein-RNA interactions at consecutive stages of gene expression.

scholarly journals Dual-function RNA-binding proteins influence mRNA abundance and translational efficiency of distinct sets of target genes.

2021 ◽  
Author(s):  
Valentin Schneider-Lunitz ◽  
Jorge Ruiz-Orera ◽  
Norbert Hubner ◽  
Sebastiaan van Heesch
Keyword(s):  
Translational Control ◽  
Binding Proteins ◽  
Target Genes ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Dual Function ◽  
Translational Efficiency ◽  
Splicing Factors ◽  
Mrna Levels

RNA-binding proteins (RBPs) are key regulators of RNA metabolism. Many RBPs possess uncharacterized RNA-binding domains and localize to multiple subcellular compartments, suggesting their involvement in multiple biological processes. We searched for such multifunctionality within a set of 143 RBPs by integrating experimentally validated target genes with the transcriptomes and translatomes of 80 human hearts. This revealed that RBP abundance is predictive of the extent of target regulation in vivo, leading us to newly associate 27 RBPs with translational control. Amongst those were several splicing factors, of which the muscle specific RBM20 modulated target translation rates through switches in isoform production. For 21 RBPs, we newly observed dual regulatory effects impacting both mRNA levels and translation rates, albeit for virtually independent sets of target genes. We highlight a subset, including G3BP1, PUM1, UCHL5, and DDX3X, where dual regulation is achieved by differential affinity for targets of distinct length and functionality. Strikingly, in a manner very similar to DDX3X, the known splicing factors EFTUD2 and PRPF8 selectively influence target translation rates depending on 5' UTR structure. Our results indicate unanticipated complexity of protein-RNA interactions at consecutive stages of gene expression and implicate multiple core splicing factors as key regulators of translational output.

scholarly journals The Role of Pumilio RNA Binding Protein in Plants

Biomolecules ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1851
Author(s):  
Sung Un Huh
Keyword(s):  
Abiotic Stresses ◽  
Translational Regulation ◽  
Binding Proteins ◽  
Target Genes ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Specific Binding ◽  
Regulation System ◽  
Translational Efficiency ◽  
Control Target

Eukaryotic organisms have a posttranscriptional/translational regulation system for the control of translational efficiency. RNA binding proteins (RBPs) have been known to control target genes. One type of protein, Pumilio (Pum)/Puf family RNA binding proteins, show a specific binding of 3′ untranslational region (3′ UTR) of target mRNA and function as a post-transcriptional/translational regulator in eukaryotic cells. Plant Pum protein is involved in development and biotic/abiotic stresses. Interestingly, Arabidopsis Pum can control target genes in a sequence-specific manner and rRNA processing in a sequence-nonspecific manner. As shown in in silico Pum gene expression analysis, Arabidopsis and rice Pum genes are responsive to biotic/abiotic stresses. Plant Pum can commonly contribute to host gene regulation at the post-transcriptional/translational step, as can mammalian Pum. However, the function of plant Pum proteins is not yet fully known. In this review, we briefly summarize the function of plant Pum in defense, development, and environmental responses via recent research and bioinformatics data.

scholarly journals An interaction network of RNA-binding proteins involved in Drosophila oogenesis

2020 ◽  
Author(s):  
Prashali Bansal ◽  
Johannes Madlung ◽  
Kristina Schaaf ◽  
Boris Macek ◽  
Fulvia Bono
Keyword(s):  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Direct Interaction ◽  
Interaction Network ◽  
Splicing Factors ◽  
Mrna Regulation ◽  
Interaction Partners ◽  
Maternal Transcripts

AbstractDuring Drosophila oogenesis, the localization and translational regulation of maternal transcripts relies on RNA-binding proteins (RBPs). Many of these RBPs localize several mRNAs and may have additional direct interaction partners to regulate their functions. Using immunoprecipitation from whole Drosophila ovaries coupled to mass spectrometry, we examined protein-protein associations of 6 GFP-tagged RBPs expressed at physiological levels. Analysis of the interaction network and further validation in human cells allowed us to identify 26 previously unknown associations, besides recovering several well characterized interactions. We identified interactions between RBPs and several splicing factors, providing links between nuclear and cytoplasmic events of mRNA regulation. Additionally, components of the translational and RNA decay machineries were selectively co-purified with some baits, suggesting a mechanism for how RBPs may regulate maternal transcripts. Given the evolutionary conservation of the studied RBPs, the interaction network presented here provides the foundation for future functional and structural studies of mRNA localization across metazoans.

scholarly journals An Interaction Network of RNA-Binding Proteins Involved in Drosophila Oogenesis

2020 ◽  
Vol 19 (9) ◽  
pp. 1485-1502
Author(s):  
Prashali Bansal ◽  
Johannes Madlung ◽  
Kristina Schaaf ◽  
Boris Macek ◽  
Fulvia Bono
Keyword(s):  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Direct Interaction ◽  
Interaction Network ◽  
Splicing Factors ◽  
Mrna Regulation ◽  
Interaction Partners ◽  
Maternal Transcripts

During Drosophila oogenesis, the localization and translational regulation of maternal transcripts relies on RNA-binding proteins (RBPs). Many of these RBPs localize several mRNAs and may have additional direct interaction partners to regulate their functions. Using immunoprecipitation from whole Drosophila ovaries coupled to mass spectrometry, we examined protein-protein associations of 6 GFP-tagged RBPs expressed at physiological levels. Analysis of the interaction network and further validation in human cells allowed us to identify 26 previously unknown associations, besides recovering several well characterized interactions. We identified interactions between RBPs and several splicing factors, providing links between nuclear and cytoplasmic events of mRNA regulation. Additionally, components of the translational and RNA decay machineries were selectively co-purified with some baits, suggesting a mechanism for how RBPs may regulate maternal transcripts. Given the evolutionary conservation of the studied RBPs, the interaction network presented here provides the foundation for future functional and structural studies of mRNA localization across metazoans.

scholarly journals An in vivo binding assay for RNA-binding proteins based on repression of a reporter gene

2017 ◽  
Author(s):  
Noa Katz ◽  
Roni Cohen ◽  
Oz Solomon ◽  
Beate Kaufmann ◽  
Noa Eden ◽  
...  
Keyword(s):  
Reporter Gene ◽  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Coat Proteins ◽  
High Throughput Assay

ABSTRACTWe employ a reporter assay and Selective 2′-hydroxyl acylation analysed by primer extension sequencing (SHAPE-seq) to study translational regulation by RNA-binding proteins, in bacteria. We designed 82 constructs, each with a single hairpin based on the binding sites of the RNA-binding coat proteins of phages MS2, PP7, GA, and Qβ, at various positions within the N-terminus of a reporter gene. In the absence of RNA-binding proteins, the translation level depends on hairpin location, and exhibits a three-nucleotide periodicity. For hairpin positions within the initiation region, we observe strong translational repression in the presence of its cognate RNA-binding protein. In vivo SHAPE-seq results for a representative construct indicate that the repression phenomenon correlates with a wide-swath of protection, including the hairpin and extending past the ribosome binding site. Consequently, our data suggest that the protection provided by the RBP-hairpin complex inhibits ribosomal initiation. Finally, utilizing the repression phenomenon for quantifying protein-RNA binding affinity in vivo, we both observe partially contrasting results to previous in vitro and in situ studies, and additionally, show that this method can be used in a high-throughput assay for a quantitative study of protein-RNA binding in vivo.

scholarly journals Proteins binding to 5' untranslated region sites: a general mechanism for translational regulation of mRNAs in human and yeast cells.

1994 ◽  
Vol 14 (9) ◽  
pp. 5898-5909 ◽  
Author(s):  
R Stripecke ◽  
C C Oliveira ◽  
J E McCarthy ◽  
M W Hentze
Keyword(s):  
Binding Sites ◽  
Untranslated Region ◽  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Yeast Cells ◽  
General Mechanism ◽  
Translational Repressor

We demonstrate that a bacteriophage protein and a spliceosomal protein can be converted into eukaryotic translational repressor proteins. mRNAs with binding sites for the bacteriophage MS2 coat protein or the spliceosomal human U1A protein were expressed in human HeLa cells and yeast. The presence of the appropriate binding protein resulted in specific, dose-dependent translational repression when the binding sites were located in the 5' untranslated region (UTR) of the reporter mRNAs. Neither mRNA export from the nucleus to the cytoplasm nor mRNA stability was demonstrably affected by the binding proteins. The data thus reveal a general mechanism for translational regulation: formation of mRNA-protein complexes in the 5' UTR controls translation initiation by steric blockage of a sensitive step in the initiation pathway. Moreover, the findings establish the basis for novel strategies to study RNA-protein interactions in vivo and to clone RNA-binding proteins.

Proteins binding to 5' untranslated region sites: a general mechanism for translational regulation of mRNAs in human and yeast cells

1994 ◽  
Vol 14 (9) ◽  
pp. 5898-5909
Author(s):  
R Stripecke ◽  
C C Oliveira ◽  
J E McCarthy ◽  
M W Hentze
Keyword(s):  
Binding Sites ◽  
Untranslated Region ◽  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Yeast Cells ◽  
General Mechanism ◽  
Translational Repressor

We demonstrate that a bacteriophage protein and a spliceosomal protein can be converted into eukaryotic translational repressor proteins. mRNAs with binding sites for the bacteriophage MS2 coat protein or the spliceosomal human U1A protein were expressed in human HeLa cells and yeast. The presence of the appropriate binding protein resulted in specific, dose-dependent translational repression when the binding sites were located in the 5' untranslated region (UTR) of the reporter mRNAs. Neither mRNA export from the nucleus to the cytoplasm nor mRNA stability was demonstrably affected by the binding proteins. The data thus reveal a general mechanism for translational regulation: formation of mRNA-protein complexes in the 5' UTR controls translation initiation by steric blockage of a sensitive step in the initiation pathway. Moreover, the findings establish the basis for novel strategies to study RNA-protein interactions in vivo and to clone RNA-binding proteins.

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 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]

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