scholarly journals Genes with 5′ terminal oligopyrimidine tracts preferentially escape global suppression of translation by the SARS-CoV-2 Nsp1 protein

2020 ◽  
Author(s):  
Shilpa Rao ◽  
Ian Hoskins ◽  
P. Daniela Garcia ◽  
Tori Tonn ◽  
Hakan Ozadam ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Ribosome Profiling ◽  
Fine Tuning ◽  
Host Protein ◽  
Structural Protein ◽  
Translation Machinery ◽  
Human Genes ◽  
Repressive Effect

AbstractViruses rely on the host translation machinery to synthesize their own proteins. Consequently, they have evolved varied mechanisms to co-opt host translation for their survival. SARS-CoV-2 relies on a non-structural protein, Nsp1, for shutting down host translation. Despite this, it is currently unknown how viral proteins and host factors critical for viral replication can escape a global shutdown of host translation. Here, using a novel FACS-based assay called MeTAFlow, we report a dose-dependent reduction in both nascent protein synthesis and mRNA abundance in cells expressing Nsp1. We perform RNA-Seq and matched ribosome profiling experiments to identify gene-specific changes both at the mRNA expression and translation level. We discover a functionally-coherent subset of human genes preferentially translated in the context of Nsp1 expression. These genes include the translation machinery components, RNA binding proteins, and others important for viral pathogenicity. Importantly, we uncovered a remarkable enrichment of 5′ terminal oligo-pyrimidine (TOP) tracts among preferentially translated genes. Using reporter assays, we validated that 5’ UTRs of TOP transcripts can drive preferential protein synthesis in the presence of NSP1. Collectively, our study suggests fine tuning of host gene expression and translation by Nsp1 despite its global repressive effect on host protein synthesis.

scholarly journals Angiogenin (ANG)—Ribonuclease Inhibitor (RNH1) System in Protein Synthesis and Disease

2021 ◽  
Vol 22 (3) ◽  
pp. 1287
Author(s):  
Mayuresh Anant Sarangdhar ◽  
Ramanjaneyulu Allam
Keyword(s):  
Protein Synthesis ◽  
Ribosome Biogenesis ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Therapeutic Potential ◽  
Translation Regulation ◽  
Future Research ◽  
Ribonuclease Inhibitor ◽  
Cell Homeostasis ◽  
Translation Machinery

Protein synthesis is a highly complex process executed by well-organized translation machinery. Ribosomes, tRNAs and mRNAs are the principal components of this machinery whereas RNA binding proteins and ribosome interacting partners act as accessory factors. Angiogenin (ANG)—Ribonuclease inhibitor (RNH1) system is one such accessory part of the translation machinery that came into focus afresh due to its unconventional role in the translation. ANG is conventionally known for its ability to induce blood vessel formation and RNH1 as a “sentry” to protect RNAs from extracellular RNases. However, recent studies suggest them to be important in translation regulation. During cell homeostasis, ANG in the nucleus promotes rRNA transcription. While under stress, ANG translocates to the cytosol and cleaves tRNA into fragments which inhibit ribosome biogenesis and protein synthesis. RNH1, which intimately interacts with ANG to inhibit its ribonucleolytic activity, can also bind to the 40S ribosomes and control translation by yet to be known mechanisms. Here, we review recent advancement in the knowledge of translation regulation by the ANG-RNH1 system. We also gather information about this system in cell homeostasis as well as in pathological conditions such as cancer and ribosomopathies. Additionally, we discuss the future research directions and therapeutic potential of this system.

scholarly journals 5′ Untranslated Region Elements Show High Abundance and Great Variability in Homologous ABCA Subfamily Genes

2020 ◽  
Vol 21 (22) ◽  
pp. 8878
Author(s):  
Pavel Dvorak ◽  
Viktor Hlavac ◽  
Pavel Soucek
Keyword(s):  
Cancer Progression ◽  
Dna Sequences ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Ribosome Profiling ◽  
Open Reading Frames ◽  
Fine Tuning ◽  
Breast Cancer Patients ◽  
Protein Levels ◽  
Reading Frames

The 12 members of the ABCA subfamily in humans are known for their ability to transport cholesterol and its derivatives, vitamins, and xenobiotics across biomembranes. Several ABCA genes are causatively linked to inborn diseases, and the role in cancer progression and metastasis is studied intensively. The regulation of translation initiation is implicated as the major mechanism in the processes of post-transcriptional modifications determining final protein levels. In the current bioinformatics study, we mapped the features of the 5′ untranslated regions (5′UTR) known to have the potential to regulate translation, such as the length of 5′UTRs, upstream ATG codons, upstream open-reading frames, introns, RNA G-quadruplex-forming sequences, stem loops, and Kozak consensus motifs, in the DNA sequences of all members of the subfamily. Subsequently, the conservation of the features, correlations among them, ribosome profiling data as well as protein levels in normal human tissues were examined. The 5′UTRs of ABCA genes contain above-average numbers of upstream ATGs, open-reading frames and introns, as well as conserved ones, and these elements probably play important biological roles in this subfamily, unlike RG4s. Although we found significant correlations among the features, we did not find any correlation between the numbers of 5′UTR features and protein tissue distribution and expression scores. We showed the existence of single nucleotide variants in relation to the 5′UTR features experimentally in a cohort of 105 breast cancer patients. 5′UTR features presumably prepare a complex playground, in which the other elements such as RNA binding proteins and non-coding RNAs play the major role in the fine-tuning of protein expression.

scholarly journals Musashi proteins are post-transcriptional regulators of the epithelial-luminal cell state

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Yarden Katz ◽  
Feifei Li ◽  
Nicole J Lambert ◽  
Ethan S Sokol ◽  
Wai-Leong Tam ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Cell Biology ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
State Regulation ◽  
Ribosome Profiling ◽  
Luminal Cell ◽  
Mesenchymal Transition ◽  
Cell State

The conserved Musashi (Msi) family of RNA binding proteins are expressed in stem/progenitor and cancer cells, but generally absent from differentiated cells, consistent with a role in cell state regulation. We found that Msi genes are rarely mutated but frequently overexpressed in human cancers and are associated with an epithelial-luminal cell state. Using ribosome profiling and RNA-seq analysis, we found that Msi proteins regulate translation of genes implicated in epithelial cell biology and epithelial-to-mesenchymal transition (EMT), and promote an epithelial splicing pattern. Overexpression of Msi proteins inhibited the translation of Jagged1, a factor required for EMT, and repressed EMT in cell culture and in mammary gland in vivo. Knockdown of Msis in epithelial cancer cells promoted loss of epithelial identity. Our results show that mammalian Msi proteins contribute to an epithelial gene expression program in neural and mammary cell types.

scholarly journals Ras Suppresses TXNIP Expression by Restricting Ribosome Translocation

2018 ◽  
Author(s):  
Zhizhou Ye ◽  
Donald E. Ayer
Keyword(s):  
Protein Synthesis ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Aerobic Glycolysis ◽  
Protein Translation ◽  
Underlying Mechanism ◽  
Metabolic Reprogramming ◽  
Coding Region ◽  
Interacting Protein ◽  
Thioredoxin Interacting Protein

ABSTRACTOncogenic Ras upregulates aerobic glycolysis to meet the bioenergetic and biosynthetic demands of rapidly growing cells. In contrast, Thioredoxin interacting protein (TXNIP) is a potent inhibitor of glucose uptake and is frequently downregulated in human cancers. Our lab previously discovered that Ras activation suppresses TXNIP transcription and translation. In this report, we developed a system to study how Ras affects TXNIP translation in the absence of transcriptional affects. We show that whereas Ras drives a global increase in protein translation, it suppresses TXNIP protein synthesis by reducing the rate at which ribosomes transit the coding region of TXNIP mRNA. To investigate the underlying mechanism(s), we randomized or optimized the codons in the TXNIP message without altering the TXNIP primary amino acid sequence. Translation from these mRNA variants is still repressed by Ras, intimating that mRNA secondary structure, miRNAs, RNA binding proteins, or codon usage do not contribute to the blockade of TXNIP synthesis. Rather, we show that the N-terminus of the growing TXNIP polypeptide is the target for Ras-dependent translational repression. Our work demonstrates how Ras suppresses TXNIP translation elongation in the face of a global upregulation of protein synthesis and provides new insight into Ras-dependent metabolic reprogramming.

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 Posttranscriptional Mechanism That Controls Ptbp1 Abundance in the Xenopus Epidermis

2014 ◽  
Vol 35 (4) ◽  
pp. 758-768 ◽  
Author(s):  
Agnès Méreau ◽  
Vincent Anquetil ◽  
Hubert Lerivray ◽  
Justine Viet ◽  
Claire Schirmer ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fine Tuning ◽  
Splice Isoforms ◽  
Content Type ◽  
Splicing Pattern ◽  
Exon 11 ◽  
Splicing Patterns ◽  
Different Tissues

The output of alternative splicing depends on the cooperative or antagonistic activities of several RNA-binding proteins (RBPs), like Ptbp1 and Esrp1 inXenopus. Fine-tuning of the RBP abundance is therefore of prime importance to achieve tissue- or cell-specific splicing patterns. Here, we addressed the mechanisms leading to the high expression of theptbp1gene, which encodes Ptbp1, inXenopusepidermis. Two splice isoforms ofptbp1mRNA differ by the presence of an alternative exon 11, and only the isoform including exon 11 can be translated to a full-length protein.In vivominigene assays revealed that the nonproductive isoform was predominantly produced. Knockdown experiments demonstrated that Esrp1, which is specific to the epidermis, strongly stimulated the expression ofptbp1by favoring the productive isoform. Consequently, knocking downesrp1phenocopiedptbp1inactivation. Conversely, Ptbp1 repressed the expression of its own gene by favoring the nonproductive isoform. Hence, a complex posttranscriptional mechanism controls Ptbp1 abundance inXenopusepidermis: skipping of exon 11 is the default splicing pattern, but Esrp1 stimulatesptbp1expression by favoring the inclusion of exon 11 up to a level that is limited by Ptbp1 itself. These results decipher a posttranscriptional mechanism that achieves various abundances of the ubiquitous RBP Ptbp1 in different tissues.

scholarly journals Posttranscriptional regulation of human endogenous retroviruses by RNA-binding motif protein 4, RBM4

2020 ◽  
Vol 117 (42) ◽  
pp. 26520-26530
Author(s):  
Amir K. Foroushani ◽  
Bryan Chim ◽  
Madeline Wong ◽  
Andre Rastegar ◽  
Patrick T. Smith ◽  
...  
Keyword(s):  
Human Genome ◽  
Posttranscriptional Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Ectopic Expression ◽  
Transcript Level ◽  
Endogenous Retroviruses ◽  
Host Protein ◽  
Binding Motif ◽  
Sequencing Data

The human genome encodes for over 1,500 RNA-binding proteins (RBPs), which coordinate regulatory events on RNA transcripts. Most studies of RBPs have concentrated on their action on host protein-encoding mRNAs, which constitute a minority of the transcriptome. A widely neglected subset of our transcriptome derives from integrated retroviral elements, termed endogenous retroviruses (ERVs), that comprise ∼8% of the human genome. Some ERVs have been shown to be transcribed under physiological and pathological conditions, suggesting that sophisticated regulatory mechanisms to coordinate and prevent their ectopic expression exist. However, it is unknown how broadly RBPs and ERV transcripts directly interact to provide a posttranscriptional layer of regulation. Here, we implemented a computational pipeline to determine the correlation of expression between individual RBPs and ERVs from single-cell or bulk RNA-sequencing data. One of our top candidates for an RBP negatively regulating ERV expression was RNA-binding motif protein 4 (RBM4). We used photoactivatable ribonucleoside-enhanced cross-linking and immunoprecipitation to demonstrate that RBM4 indeed bound ERV transcripts at CGG consensus elements. Loss of RBM4 resulted in an elevated transcript level of bound ERVs of the HERV-K and -H families, as well as increased expression of HERV-K envelope protein. We pinpointed RBM4 regulation of HERV-K to a CGG-containing element that is conserved in the LTRs of HERV-K-10, -K-11, and -K-20, and validated the functionality of this site using reporter assays. In summary, we systematically identified RBPs that may regulate ERV function and demonstrate a role for RBM4 in controlling ERV expression.

scholarly journals miR-503 represses CUG-binding protein 1 translation by recruiting CUGBP1 mRNA to processing bodies

2012 ◽  
Vol 23 (1) ◽  
pp. 151-162 ◽  
Author(s):  
Yu-Hong Cui ◽  
Lan Xiao ◽  
Jaladanki N. Rao ◽  
Tongtong Zou ◽  
Lan Liu ◽  
...  
Keyword(s):  
Rna Binding ◽  
De Novo ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Mrna Levels ◽  
Intestinal Epithelial ◽  
Coding Region ◽  
Processing Bodies ◽  
Target Mrnas ◽  
Repressive Effect

microRNAs (miRNAs) and RNA-binding proteins (RBPs) jointly regulate gene expression at the posttranscriptional level and are involved in many aspects of cellular functions. The RBP CUG-binding protein 1 (CUGBP1) destabilizes and represses the translation of several target mRNAs, but the exact mechanism that regulates CUGBP1 abundance remains elusive. In this paper, we show that miR-503, computationally predicted to associate with three sites of the CUGBP1 mRNA, represses CUGBP1 expression. Overexpression of an miR-503 precursor (pre-miR-503) reduced the de novo synthesis of CUGBP1 protein, whereas inhibiting miR-503 by using an antisense RNA (antagomir) enhanced CUGBP1 biosynthesis and elevated its abundance; neither intervention changed total CUGBP1 mRNA levels. Studies using heterologous reporter constructs revealed a greater repressive effect of miR-503 through the CUGBP1 coding region sites than through the single CUGBP1 3′-untranslated region target site. CUGBP1 mRNA levels in processing bodies (P-bodies) increased in cells transfected with pre-miR-503, while silencing P-body resident proteins Ago2, RCK, or LSm4 decreased miR-503–mediated repression of CUGBP1 expression. Decreasing the levels of cellular polyamines reduced endogenous miR-503 levels and promoted CUGBP1 expression, an effect that was prevented by ectopic miR-503 overexpression. Repression of CUGBP1 by miR-503 in turn altered the expression of CUGBP1 target mRNAs and thus increased the sensitivity of intestinal epithelial cells to apoptosis. These findings identify miR-503 as both a novel regulator of CUGBP1 expression and a modulator of intestinal epithelial homoeostasis.

scholarly journals Pan-cancer analysis of RNA binding proteins (RBPs) reveals the involvement of the core pre-mRNA splicing and translation machinery in tumorigenesis

2018 ◽  
Author(s):  
Bin Zhang ◽  
Kamesh R. Babu ◽  
Chun You Lim ◽  
Zhi Hao Kwok ◽  
Jia Li ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Copy Number ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Human Cancer ◽  
Mrna Splicing ◽  
Regulatory Mechanisms ◽  
Translation Machinery ◽  
The Core ◽  
Cancer Types

AbstractRNA binding proteins (RBPs) are key regulators of posttranscriptional processes such as RNA maturation, transport, localization, turnover and translation. Despite their dysregulation in various diseases including cancer, the landscape of RBP expression and regulatory mechanisms in human cancer has not been well characterized. Here, we analyzed mRNA expression of 1487 RBPs in ~6700 clinical samples across 16 human cancer types and found that there were significantly more upregulated RBPs than downregulated ones in tumors when compared to their adjacent normal tissues. Across almost all of the 16 cancer types, 109 RBPs were consistently upregulated (cuRBPs) while only 41 RBPs were consistently downregulated (cdRBPs). Integrating expression with the copy number and DNA methylation data, we found that the overexpression of cuRBPs is largely associated with the amplification of copy number, whereas the downregulation of cdRBPs may be a result of epigenetic silencing mediated by DNA methylation. Furthermore, our results indicated that cuRBPs could work together to promote cancer progression potentially through the involvement of splicing and translation machinery, while cdRBPs might function independently to suppress tumorigenesis. Additionally, we focused on colon cancer and identified several novel potential oncogenic RBPs, such as PABPC1L which might promote cancer development via regulating the core splicing machinery. In summary, we showed distinct expression landscapes, regulatory mechanisms and characteristics of cuRBPs and cdRBPs and implicated several novel RBPs in cancer pathogenesis. Moreover, our results suggest that the involvement of the core pre-mRNA splicing and translation machinery could be critical in tumorigenesis.

scholarly journals SARS-CoV-2 nucleocapsid protein undergoes liquid-liquid phase separation stimulated by RNA and partitions into phases of human ribonucleoproteins

2020 ◽  
Author(s):  
Theodora Myrto Perdikari ◽  
Anastasia C. Murthy ◽  
Veronica H. Ryan ◽  
Scott Watters ◽  
Mandar T. Naik ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Nucleocapsid Protein ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Liquid Phase Separation ◽  
Host Protein ◽  
Host Cells ◽  
Liquid Liquid Phase Separation

AbstractTightly packed complexes of nucleocapsid protein and genomic RNA form the core of viruses and may assemble within viral factories, dynamic compartments formed within the host cells. Here, we examine the possibility that the multivalent RNA-binding nucleocapsid protein (N) from the severe acute respiratory syndrome coronavirus (SARS-CoV-2) compacts RNA via protein-RNA liquid-liquid phase separation (LLPS) and that N interactions with host RNA-binding proteins are mediated by phase separation. To this end, we created a construct expressing recombinant N fused to a N-terminal maltose binding protein tag which helps keep the oligomeric N soluble for purification. Using in vitro phase separation assays, we find that N is assembly-prone and phase separates avidly. Phase separation is modulated by addition of RNA and changes in pH and is disfavored at high concentrations of salt. Furthermore, N enters into in vitro phase separated condensates of full-length human hnRNPs (TDP-43, FUS, and hnRNPA2) and their low complexity domains (LCs). However, N partitioning into the LC of FUS, but not TDP-43 or hnRNPA2, requires cleavage of the solubilizing MBP fusion. Hence, LLPS may be an essential mechanism used for SARS-CoV-2 and other RNA viral genome packing and host protein co-opting, functions necessary for viral replication and hence infectivity.

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