scholarly journals GC content shapes mRNA decay and storage in human cells

2018 ◽  
Author(s):  
Maïté Courel ◽  
Yves Clément ◽  
Dominika Foretek ◽  
Olivia Vidal Cruchez ◽  
Zhou Yi ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Mrna Decay ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Gc Content ◽  
Mrna Translation ◽  
Human Cells ◽  
Protein Yield ◽  
Fine Tuning ◽  
Rnp Granules

SummaryControl of protein expression results from the fine tuning of mRNA synthesis, decay and translation. These processes, which are controlled by a large number of RNA-binding proteins and by localization in RNP granules such as P-bodies, appear often intimately linked although the rules of this interplay are not well understood. In this study, we combined our recent P-body transcriptome with various transcriptomes obtained following silencing of broadly acting mRNA decay and repression factors. This analysis revealed the central role of GC content in mRNA fate, in terms of P-body localization, mRNA translation and mRNA decay. It also rationalized why PBs mRNAs have a strikingly low protein yield. We report too the existence of distinct mRNA decay pathways with preference for AU-rich or GC-rich transcripts. Compared to this impact of the GC content, sequence-specific RBPs and miRNAs appeared to have only modest additional effects on their bulk targets. Altogether, these results lead to an integrated view of post-transcriptional control in human cells where most regulation at the level of translation is dedicated to AU-rich mRNAs, which have a limiting protein yield, whereas regulation at the level of 5’ decay applies to GC-rich mRNAs, whose translation is optimal.

scholarly journals GC content shapes mRNA storage and decay in human cells

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Maïté Courel ◽  
Yves Clément ◽  
Clémentine Bossevain ◽  
Dominika Foretek ◽  
Olivia Vidal Cruchez ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Gc Content ◽  
Mrna Translation ◽  
Human Cells ◽  
Translation Regulation ◽  
P Bodies ◽  
Related Data ◽  
Low Protein ◽  
Mrna Fate

mRNA translation and decay appear often intimately linked although the rules of this interplay are poorly understood. In this study, we combined our recent P-body transcriptome with transcriptomes obtained following silencing of broadly acting mRNA decay and repression factors, and with available CLIP and related data. This revealed the central role of GC content in mRNA fate, in terms of P-body localization, mRNA translation and mRNA stability: P-bodies contain mostly AU-rich mRNAs, which have a particular codon usage associated with a low protein yield; AU-rich and GC-rich transcripts tend to follow distinct decay pathways; and the targets of sequence-specific RBPs and miRNAs are also biased in terms of GC content. Altogether, these results suggest an integrated view of post-transcriptional control in human cells where most translation regulation is dedicated to inefficiently translated AU-rich mRNAs, whereas control at the level of 5’ decay applies to optimally translated GC-rich mRNAs.

scholarly journals RNA-binding proteins La and HuR cooperatively modulate translation repression of PDCD4 mRNA

2020 ◽  
pp. jbc.RA120.014894
Author(s):  
Ravi Kumar ◽  
Dipak Kumar Poria ◽  
Partho Sarothi Ray
Keyword(s):  
Inflammatory Response ◽  
Chronic Inflammation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Critical Role ◽  
Regulation Of Gene Expression ◽  
Mrna Translation ◽  
Suppressor Gene ◽  
Cooperative Action ◽  
Translation Repression

Post-transcriptional regulation of gene expression plays a critical role in controlling the inflammatory response. An uncontrolled inflammatory response results in chronic inflammation, often leading to tumorigenesis. Programmed cell death 4 (PDCD4) is a pro-inflammatory tumor-suppressor gene which helps to prevent the transition from chronic inflammation to cancer. PDCD4 mRNA translation is regulated by an interplay between the oncogenic microRNA miR-21 and the RNA-binding protein (RBP) HuR in response to LPS stimulation, but the role of other regulatory factors remain unknown. Here we report that the RBP Lupus antigen (La) interacts with the 3’UTR of PDCD4 mRNA and prevents miR-21-mediated translation repression. While LPS causes nuclear-cytoplasmic translocation of HuR, it enhances cellular La expression. Remarkably, La and HuR were found to bind cooperatively to the PDCD4 mRNA and mitigate miR-21-mediated translation repression. The cooperative action of La and HuR reduced cell proliferation and enhanced apoptosis, reversing the pro-oncogenic function of miR-21. Together, these observations demonstrate a cooperative interplay between two RBPs, triggered differentially by the same stimulus, which exerts a synergistic effect on PDCD4 expression and thereby helps maintain a balance between inflammation and tumorigenesis.

scholarly journals IDH1 fine-tunes cap-dependent translation initiation

2019 ◽  
Vol 11 (10) ◽  
pp. 816-828 ◽  
Author(s):  
Lichao Liu ◽  
J Yuyang Lu ◽  
Fajin Li ◽  
Xudong Xing ◽  
Tong Li ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Ribosomal Proteins ◽  
Rna Binding ◽  
Embryonic Stem ◽  
Mrna Translation ◽  
Fine Tuning ◽  
Start Codon ◽  
Metabolic Enzyme ◽  
Oxidative Decarboxylation ◽  
Dependent Manner

Abstract The metabolic enzyme isocitrate dehydrogenase 1 (IDH1) catalyzes the oxidative decarboxylation of isocitrate to α-ketoglutarate (α-KG). Its mutation often leads to aberrant gene expression in cancer. IDH1 was reported to bind thousands of RNA transcripts in a sequence-dependent manner; yet, the functional significance of this RNA-binding activity remains elusive. Here, we report that IDH1 promotes mRNA translation via direct associations with polysome mRNA and translation machinery. Comprehensive proteomic analysis in embryonic stem cells (ESCs) revealed striking enrichment of ribosomal proteins and translation regulators in IDH1-bound protein interactomes. We performed ribosomal profiling and analyzed mRNA transcripts that are associated with actively translating polysomes. Interestingly, knockout of IDH1 in ESCs led to significant downregulation of polysome-bound mRNA in IDH1 targets and subtle upregulation of ribosome densities at the start codon, indicating inefficient translation initiation upon loss of IDH1. Tethering IDH1 to a luciferase mRNA via the MS2-MBP system promotes luciferase translation, independently of the catalytic activity of IDH1. Intriguingly, IDH1 fails to enhance luciferase translation driven by an internal ribosome entry site. Together, these results reveal an unforeseen role of IDH1 in fine-tuning cap-dependent translation via the initiation step.

scholarly journals Deep analysis of RNA N6-adenosine methylation (m6A) patterns in human cells

2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Jun Wang ◽  
Liangjiang Wang
Keyword(s):  
Rna Binding ◽  
Data Transfer ◽  
Rna Binding Proteins ◽  
Consensus Sequence ◽  
Model Performance ◽  
Cell Types ◽  
Human Cells ◽  
Rna Modification ◽  
Cell Type ◽  
Cell Type Specific

Abstract N6-adenosine methylation (m6A) is the most abundant internal RNA modification in eukaryotes, and affects RNA metabolism and non-coding RNA function. Previous studies suggest that m6A modifications in mammals occur on the consensus sequence DRACH (D = A/G/U, R = A/G, H = A/C/U). However, only about 10% of such adenosines can be m6A-methylated, and the underlying sequence determinants are still unclear. Notably, the regulation of m6A modifications can be cell-type-specific. In this study, we have developed a deep learning model, called TDm6A, to predict RNA m6A modifications in human cells. For cell types with limited availability of m6A data, transfer learning may be used to enhance TDm6A model performance. We show that TDm6A can learn common and cell-type-specific motifs, some of which are associated with RNA-binding proteins previously reported to be m6A readers or anti-readers. In addition, we have used TDm6A to predict m6A sites on human long non-coding RNAs (lncRNAs) for selection of candidates with high levels of m6A modifications. The results provide new insights into m6A modifications on human protein-coding and non-coding transcripts.

scholarly journals G3BP1-linked mRNA partitioning supports selective protein synthesis in response to oxidative stress

2020 ◽  
Vol 48 (12) ◽  
pp. 6855-6873 ◽  
Author(s):  
Syam Prakash Somasekharan ◽  
Fan Zhang ◽  
Neetu Saxena ◽  
Jia Ni Huang ◽  
I-Chih Kuo ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mrna Translation ◽  
Stress Adaptation ◽  
Rna Granules ◽  
Before And After ◽  
Selective Translation ◽  
Energy Consuming ◽  
Limit Energy

Abstract Cells limit energy-consuming mRNA translation during stress to maintain metabolic homeostasis. Sequestration of mRNAs by RNA binding proteins (RBPs) into RNA granules reduces their translation, but it remains unclear whether RBPs also function in partitioning of specific transcripts to polysomes (PSs) to guide selective translation and stress adaptation in cancer. To study transcript partitioning under cell stress, we catalogued mRNAs enriched in prostate carcinoma PC-3 cell PSs, as defined by polysome fractionation and RNA sequencing (RNAseq), and compared them to mRNAs complexed with the known SG-nucleator protein, G3BP1, as defined by spatially-restricted enzymatic tagging and RNAseq. By comparing these compartments before and after short-term arsenite-induced oxidative stress, we identified three major categories of transcripts, namely those that were G3BP1-associated and PS-depleted, G3BP1-dissociated and PS-enriched, and G3BP1-associated but also PS-enriched. Oxidative stress profoundly altered the partitioning of transcripts between these compartments. Under arsenite stress, G3BP1-associated and PS-depleted transcripts correlated with reduced expression of encoded mitochondrial proteins, PS-enriched transcripts that disassociated from G3BP1 encoded cell cycle and cytoprotective proteins whose expression increased, while transcripts that were both G3BP1-associated and PS-enriched encoded proteins involved in diverse stress response pathways. Therefore, G3BP1 guides transcript partitioning to reprogram mRNA translation and support stress adaptation.

scholarly journals RNA-Binding Proteins HuB, HuC, and HuD are Distinctly Regulated in Dorsal Root Ganglia Neurons from STZ-Sensitive Compared to STZ-Resistant Diabetic Mice

2019 ◽  
Vol 20 (8) ◽  
pp. 1965 ◽  
Author(s):  
Cosmin Cătălin Mustăciosu ◽  
Adela Banciu ◽  
Călin Mircea Rusu ◽  
Daniel Dumitru Banciu ◽  
Diana Savu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Body Weight ◽  
Sensory Neurons ◽  
Dorsal Root Ganglia ◽  
Dorsal Root ◽  
Transcriptional Control ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Diabetic Mice

The neuron-specific Elav-like Hu RNA-binding proteins were described to play an important role in neuronal differentiation and plasticity by ensuring the post-transcriptional control of RNAs encoding for various proteins. Although Elav-like Hu proteins alterations were reported in diabetes or neuropathy, little is known about the regulation of neuron-specific Elav-like Hu RNA-binding proteins in sensory neurons of dorsal root ganglia (DRG) due to the diabetic condition. The goal of our study was to analyze the gene and protein expression of HuB, HuC, and HuD in DRG sensory neurons in diabetes. The diabetic condition was induced in CD-1 adult male mice with single-intraperitoneal injection of streptozotocin (STZ, 150 mg/kg), and 8-weeks (advanced diabetes) after induction was quantified the Elav-like proteins expression. Based on the glycemia values, we identified two types of responses to STZ, and mice were classified in STZ-resistant (diabetic resistant, glycemia < 260 mg/dL) and STZ-sensitive (diabetic, glycemia > 260 mg/dL). Body weight measurements indicated that 8-weeks after STZ-induction of diabetes, control mice have a higher increase in body weight compared to the diabetic and diabetic resistant mice. Moreover, after 8-weeks, diabetic mice (19.52 ± 3.52 s) have longer paw withdrawal latencies in the hot-plate test than diabetic resistant (11.36 ± 1.92 s) and control (11.03 ± 1.97 s) mice, that correlates with the installation of warm hypoalgesia due to the diabetic condition. Further on, we evidenced the decrease of Elav-like gene expression in DRG neurons of diabetic mice (Elavl2, 0.68 ± 0.05 fold; Elavl3, 0.65 ± 0.01 fold; Elavl4, 0.53 ± 0.07 fold) and diabetic resistant mice (Ealvl2, 0.56 ± 0.07 fold; Elavl3, 0.32 ± 0.09 fold) compared to control mice. Interestingly, Elav-like genes have a more accentuated downregulation in diabetic resistant than in diabetic mice, although hypoalgesia was evidenced only in diabetic mice. The Elav-like gene expression changes do not always correlate with the Hu protein expression changes. To detail, HuB is upregulated and HuD is downregulated in diabetic mice, while HuB, HuC, and HuD are downregulated in diabetic resistant mice compared to control mice. To resume, we demonstrated HuD downregulation and HuB upregulation in DRG sensory neurons induced by diabetes, which might be correlated with altered post-transcriptional control of RNAs involved in the regulation of thermal hypoalgesia condition caused by the advanced diabetic neuropathy.

scholarly journals A potential role for a novel ZC3H5 complex in regulating mRNA translation in Trypanosoma brucei

2020 ◽  
Vol 295 (42) ◽  
pp. 14291-14304
Author(s):  
Kathrin Bajak ◽  
Kevin Leiss ◽  
Christine Clayton ◽  
Esteban Erben
Keyword(s):  
Gene Expression ◽  
Trypanosoma Brucei ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Affinity Purification ◽  
Critical Role ◽  
Rna Binding Protein ◽  
Mrna Translation

In Trypanosoma brucei and related kinetoplastids, gene expression regulation occurs mostly posttranscriptionally. Consequently, RNA-binding proteins play a critical role in the regulation of mRNA and protein abundance. Yet, the roles of many RNA-binding proteins are not understood. Our previous research identified the RNA-binding protein ZC3H5 as possibly involved in gene repression, but its role in controlling gene expression was unknown. We here show that ZC3H5 is an essential cytoplasmic RNA-binding protein. RNAi targeting ZC3H5 causes accumulation of precytokinetic cells followed by rapid cell death. Affinity purification and pairwise yeast two-hybrid analysis suggest that ZC3H5 forms a complex with three other proteins, encoded by genes Tb927.11.4900, Tb927.8.1500, and Tb927.7.3040. RNA immunoprecipitation revealed that ZC3H5 is preferentially associated with poorly translated, low-stability mRNAs, the 5′-untranslated regions and coding regions of which are enriched in the motif (U/A)UAG(U/A). As previously found in high-throughput analyses, artificial tethering of ZC3H5 to a reporter mRNA or other complex components repressed reporter expression. However, depletion of ZC3H5 in vivo caused only very minor decreases in a few targets, marked increases in the abundances of very stable mRNAs, an increase in monosomes at the expense of large polysomes, and appearance of “halfmer” disomes containing two 80S subunits and one 40S subunit. We speculate that the ZC3H5 complex might be implicated in quality control during the translation of suboptimal open reading frames.

Targeting of transcripts encoding membrane proteins in polarized epithelia: RNA–protein binding studies of the SGLT1 3′-UTR

2008 ◽  
Vol 36 (3) ◽  
pp. 525-527 ◽  
Author(s):  
Christopher M. Pedder ◽  
Dianne Ford ◽  
John E. Hesketh
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Protein Localization ◽  
Complex Structure ◽  
Mrna Translation ◽  
Drosophila Embryo ◽  
Binding Studies ◽  
Polarized Epithelia ◽  
Apical Localization

mRNA stability, mRNA translation and spatial localization of mRNA species within a cell can be governed by signals in the 3′-UTR (3′-untranslated region). Local translation of proteins is essential for the development of many eukaryotic cell types, such as the Drosophila embryo, where the spatial and temporal localization of bicoid and gurken mRNAs, among others, is required to establish morphogen gradients. More recent studies have suggested that mRNA localization also occurs with transcripts coding for membrane-based or secreted proteins, and that localization at organelles such as the endoplasmic reticulum directs translation more efficiently to specific subdomains, so as to aid correct protein localization. In human epithelial cells, the mRNA coding for SGLT1 (sodium–glucose co-transporter 1), an apical membrane protein, has been shown to be localized apically in polarized cells. However, the nature of the signals and RNA-binding proteins involved are unknown. Ongoing work is aimed at identifying the localization signals in the SGLT1 3′-UTR and the corresponding binding proteins. Using a protein extract from polarized Caco-2 cells, both EMSAs (electrophoretic mobility-shift assays) and UV cross-linking assays have shown that a specific protein complex is formed with the first 300 bases of the 3′-UTR sequence. MFold predictions suggest that this region folds into a complex structure and ongoing studies using a series of strategic deletions are being carried out to identify the precise nature of the motif involved, particularly the role of the sequence or RNA secondary structure, as well as to identify the main proteins present within the complex. Such information will provide details of the post-transcriptional events that lead to apical localization of the SGLT1 transcript and may reveal mechanisms of more fundamental importance in the apical localization of proteins in polarized epithelia.

scholarly journals Two Yeast La Motif-containing Proteins Are RNA-binding Proteins that Associate with Polyribosomes

1999 ◽  
Vol 10 (11) ◽  
pp. 3849-3862 ◽  
Author(s):  
Suzanne G. Sobel ◽  
Sandra L. Wolin
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Polymerase Iii ◽  
Mrna Translation ◽  
Protein Synthesis Inhibitors ◽  
La Protein ◽  
Evolutionarily Conserved ◽  
Synthetically Lethal ◽  
Type Strains

We have characterized two Saccharomyces cerevisiaeproteins, Sro9p and Slf1p, which contain a highly conserved motif found in all known La proteins. Originally described as an autoantigen in patients with rheumatic disease, the La protein binds to newly synthesized RNA polymerase III transcripts. In yeast, the La protein homologue Lhp1p is required for the normal pathway of tRNA maturation and also stabilizes newly synthesized U6 RNA. We show that deletions in both SRO9 and SLF1 are not synthetically lethal with a deletion in LHP1, indicating that the three proteins do not function in a single essential process. Indirect immunofluorescence microscopy reveals that although Lhp1p is primarily localized to the nucleus, Sro9p is cytoplasmic. We demonstrate that Sro9p and Slf1p are RNA-binding proteins that associate preferentially with translating ribosomes. Consistent with a role in translation, strains lacking either Sro9p or Slf1p are less sensitive than wild-type strains to certain protein synthesis inhibitors. Thus, Sro9p and Slf1p define a new and possibly evolutionarily conserved class of La motif-containing proteins that may function in the cytoplasm to modulate mRNA translation.

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