Translation control of Enterovirus A71 gene expression

AbstractUpon EV-A71 infection of a host cell, EV-A71 RNA is translated into a viral polyprotein. Although EV-A71 can use the cellular translation machinery to produce viral proteins, unlike cellular translation, which is cap-dependent, the viral RNA genome of EV-A71 does not contain a 5′ cap and the translation of EV-A71 protein is cap-independent, which is mediated by the internal ribosomal entry site (IRES) located in the 5′ UTR of EV-A71 mRNA. Like many other eukaryotic viruses, EV-A71 manipulates the host cell translation devices, using an elegant RNA-centric strategy in infected cells. During viral translation, viral RNA plays an important role in controlling the stage of protein synthesis. In addition, due to the cellular defense mechanism, viral replication is limited by down-regulating translation. EV-A71 also utilizes protein factors in the host to overcome antiviral responses or even use them to promote viral translation rather than host cell translation. In this review, we provide an introduction to the known strategies for EV-A71 to exploit cellular translation mechanisms.

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Heat Shock Protein A6, a Novel HSP70, Is Induced During Enterovirus A71 Infection to Facilitate Internal Ribosomal Entry Site-Mediated Translation

Frontiers in Microbiology ◽

10.3389/fmicb.2021.664955 ◽

2021 ◽

Vol 12 ◽

Author(s):

Yu-Siang Su ◽

Lih-Hwa Hwang ◽

Chi-Ju Chen

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Neurological Diseases ◽

Internal Ribosomal Entry Site ◽

Viral Proteins ◽

Cellular Protein ◽

Encephalomyocarditis Virus ◽

Entry Site ◽

Enterovirus A71 ◽

Ribosomal Entry

Enterovirus A71 (EV-A71) is a human pathogen causing hand, foot, and mouth disease (HFMD) in children. Its infection can lead to severe neurological diseases or even death in some cases. While being produced in a large quantity during infection, viral proteins often require the assistance from cellular chaperones for proper folding. In this study, we found that heat shock protein A6 (HSPA6), whose function in viral life cycle is scarcely studied, was induced and functioned as a positive regulator for EV-A71 infection. Depletion of HSPA6 led to the reductions of EV-A71 viral proteins, viral RNA and virions as a result of the downregulation of internal ribosomal entry site (IRES)-mediated translation. Unlike other HSP70 isoforms such as HSPA1, HSPA8, and HSPA9, which regulate all phases of the EV-A71 life, HSPA6 was required for the IRES-mediated translation only. Unexpectedly, the importance of HSPA6 in the IRES activity could be observed in the absence of viral proteins, suggesting that HSPA6 facilitated IRES activity through cellular factor(s) instead of viral proteins. Intriguingly, the knockdown of HSPA6 also caused the reduction of luciferase activity driven by the IRES from coxsackievirus A16, echovirus 9, encephalomyocarditis virus, or hepatitis C virus, supporting that HSPA6 may assist the function of a cellular protein generally required for viral IRES activities.

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A Moonlighting microRNA: Mechanism(s) of miR-122-Mediated Viral RNA Accumulation

Proceedings ◽

10.3390/proceedings2020050131 ◽

2020 ◽

Vol 50 (1) ◽

pp. 131

Author(s):

Jasmin Chahal ◽

Luca FR Gebert ◽

Hin Hark Gan ◽

Kristin C Gunsalus ◽

Ian J MacRae ◽

...

Keyword(s):

Cell Culture ◽

Viral Genome ◽

Rna Virus ◽

Internal Ribosomal Entry Site ◽

Viral Rna ◽

Antiviral Resistance ◽

Hcv Rna ◽

Entry Site ◽

Rna Chaperone ◽

Rna Accumulation

Hepatitis C virus (HCV) is a positive-sense RNA virus that interacts with a human-liver-specific microRNA, termed miR-122. miR-122 binds to two sites in the 5' untranslated region (UTR) of the viral genome, and this interaction promotes HCV RNA accumulation. This interaction is important for viral RNA accumulation in cell culture, and miR-122 inhibitors have been demonstrated to be efficacious in reducing HCV titers in chronic HCV-infected patients. However, the precise mechanism(s) of miR-122-mediated viral RNA accumulation have remained elusive. We have used biophysical analysis and assays for viral replication in cell culture to understand the interactions between the human Argonaute 2 (hAgo2):miR-122 complex and the HCV genome. In addition, we have analyzed several resistance-associated variants which were isolated from patients who underwent miR-122 inhibitor-based therapy in order to shed light on novel mechanisms of antiviral resistance. Our results provide a new model for miR-122:HCV RNA interactions and demonstrate that miR-122 plays at least three roles in the HCV life cycle: (1) miR-122 acts as an RNA chaperone to suppress an energetically favorable secondary structure and allows the viral internal ribosomal entry site (IRES) to form; (2) miR-122 binding to the 5' terminus protects the genome from the activity of cellular pyrophosphatases (DOM3Z and DUSP11) and subsequent exonuclease-mediated decay; and (3) the Argonaute (Ago) protein at Site 2 makes direct contact with the HCV IRES, enhancing viral translation. In addition, analyses of several resistance-associated variants that were isolated from patients that underwent miR-122 inhibitor-based therapy suggests that mutations in the 5' terminus alter the structure of the 5' UTR in a manner that promotes RNA chaperone activity or viral genome stability, even in the absence of miR-122. Taken together, these findings provide insight into the mechanism(s) of miR-122-mediated viral RNA accumulation and suggest new mechanisms of antiviral resistance which are mediated by changes in RNA structure.

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Far upstream element binding protein 1 binds the internal ribosomal entry site of enterovirus 71 and enhances viral translation and viral growth

Nucleic Acids Research ◽

10.1093/nar/gkr682 ◽

2011 ◽

Vol 39 (22) ◽

pp. 9633-9648 ◽

Cited By ~ 55

Author(s):

P.-N. Huang ◽

J.-Y. Lin ◽

N. Locker ◽

Y.-A. Kung ◽

C.-T. Hung ◽

...

Keyword(s):

Binding Protein ◽

Internal Ribosomal Entry Site ◽

Enterovirus 71 ◽

Entry Site ◽

Viral Translation ◽

Viral Growth ◽

Element Binding Protein ◽

Upstream Element ◽

Ribosomal Entry

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Translocating lipopolysaccharide correlates with the severity of enterovirus A71-induced HFMD by promoting pro-inflammation and viral IRES activity

10.21203/rs.3.rs-806112/v1 ◽

2021 ◽

Author(s):

Yuya Wang ◽

Xiaoling Xue ◽

Kena Dan ◽

Xiongbo Yang ◽

Qingqing Yang ◽

...

Keyword(s):

Inflammatory Mediators ◽

Internal Ribosomal Entry Site ◽

Foot And Mouth Disease ◽

Serum Levels ◽

Interferon Γ ◽

Healthy Children ◽

Entry Site ◽

Fatty Acid Binding ◽

Enterovirus A71

Abstract BackgroundThe increase of inflammation-inducing enterobacteria was recently observed in severe hand, foot, and mouth disease (HFMD) caused by Enterovirus A71 (EV-A71). This study aimed to verify the occurrence of bacterial translocation (BT) and further explore the contributory role of BT to severity of EV-A71-mediated HFMD cases. MethodsSerum specimens from 65 mild and 65 severe EV-A71-associated HFMD cases and 65 healthy children were collected. EV-A71 VP1 in serum, inﬂammatory mediators including C-reactive protein, IL-1β, IL-6, interferon-γ and tumor necrosis factor-α, BT related biomarkers including Claudin-3, intestinal fatty acid binding protein, lipopolysaccharide (LPS), soluble CD14(sCD14) and endotoxin core antibody were measured by ELISA. Bacterial DNA (BactDNA) fragments were quantified by quantified PCR (qPCR). Rhabdomyosarcoma (RD) cells, infected with LPS-pre-incubated EV-A71 or transfected with plasmid containing viral 2A pro or mRNA containing viral internal ribosomal entry site (IRES), were post-treated with or without LPS in vitro . EV-A71 RNA and viral or cellular proteins were determined by qPCR and western blot, respectively. ResultsCompared to mild HFMD patients, remarkably higher inﬂammatory mediators as well as BT-related biomarkers except BactDNA were observed in severe HFMD cases (all P <0.05). In severe HFMD group, circulating concentrations of LPS and sCD14 showed statistical correlations with inflammation indices (all P <0.05), serum levels of EV-A71 VP1 were found to be positively correlated with serum LPS ( r =0.341, P =0.005) and serum sCD14( r =0.458, P <0.001). In vitro , EV-A71 attachment and internalization were only slightly promoted by LPS pre-incubation; however, EV-A71 proliferation and viral 2A pro -mediated IRES activity were significantly accelerated by LPS post-treatment. ConclusionsOur results collectively indicate that gut-derived translocating LPS contributes to the severity of EV-A71-induced HFMD by driving inflammatory response and viral proliferation via viral 2A pro -mediated IRES.

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Far upstream element binding protein 2 interacts with enterovirus 71 internal ribosomal entry site and negatively regulates viral translation

Nucleic Acids Research ◽

10.1093/nar/gkn901 ◽

2008 ◽

Vol 37 (1) ◽

pp. 47-59 ◽

Cited By ~ 100

Author(s):

Jing-Yi Lin ◽

Mei-Ling Li ◽

Shin-Ru Shih

Keyword(s):

Binding Protein ◽

Internal Ribosomal Entry Site ◽

Enterovirus 71 ◽

Entry Site ◽

Viral Translation ◽

Element Binding Protein ◽

Upstream Element ◽

Ribosomal Entry

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Engineered Oncolytic Poliovirus PVSRIPO Subverts MDA5-Dependent Innate Immune Responses in Cancer Cells

Journal of Virology ◽

10.1128/jvi.00879-18 ◽

2018 ◽

Vol 92 (19) ◽

Cited By ~ 6

Author(s):

Ross W. Walton ◽

Michael C. Brown ◽

Matthew T. Sacco ◽

Matthias Gromeier

Keyword(s):

Cancer Immunotherapy ◽

Internal Ribosomal Entry Site ◽

Type I ◽

Cell Infection ◽

Entry Site ◽

Neoplastic Cells ◽

Who Grade ◽

Antiviral Responses ◽

Viral Spread ◽

Dismal Prognosis

ABSTRACTWe are pursuing cancer immunotherapy with a neuro-attenuated recombinant poliovirus, PVSRIPO. PVSRIPO is the live attenuated type 1 (Sabin) poliovirus vaccine carrying a heterologous internal ribosomal entry site (IRES) of human rhinovirus type 2 (HRV2). Intratumoral infusion of PVSRIPO is showing promise in the therapy of recurrent WHO grade IV malignant glioma (glioblastoma), a notoriously treatment-refractory cancer with dismal prognosis. PVSRIPO exhibits profound cytotoxicity in infected neoplastic cells expressing the poliovirus receptor CD155. In addition, it elicits intriguing persistent translation and replication, giving rise to sustained type I interferon (IFN)-dominant proinflammatory stimulation of antigen-presenting cells. A key determinant of the inflammatory footprint generated by neoplastic cell infection and its role in shaping the adaptive response after PVSRIPO tumor infection is the virus's inherent relationship to the host's innate antiviral response. In this report, we define subversion of innate host immunity by PVSRIPO, enabling productive viral translation and cytopathogenicity with extremely low multiplicities of infection in the presence of an active innate antiviral IFN response.IMPORTANCEEngaging innate antiviral responses is considered key for instigating tumor-antigen-specific antitumor immunity with cancer immunotherapy approaches. However, they are a double-edged sword for attempts to enlist viruses in such approaches. In addition to their role in the transition from innate to adaptive immunity, innate antiviral IFN responses may intercept the viral life cycle in cancerous cells, prevent viral cytopathogenicity, and restrict viral spread. This has been shown to reduce overall antitumor efficacy of several proposed oncolytic virus prototypes, presumably by limiting direct cell killing and the ensuing inflammatory profile within the infected tumor. In this report, we outline how an unusual recalcitrance of polioviruses toward innate antiviral responses permits viral cytotoxicity and propagation in neoplastic cells, combined with engaging active innate antiviral IFN responses.

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Flavivirus RNA methylation

Journal of General Virology ◽

10.1099/vir.0.062208-0 ◽

2014 ◽

Vol 95 (4) ◽

pp. 763-778 ◽

Cited By ~ 66

Author(s):

Hongping Dong ◽

Katja Fink ◽

Roland Züst ◽

Siew Pheng Lim ◽

Cheng-Feng Qin ◽

...

Keyword(s):

Immune Response ◽

Rna Structure ◽

Innate Immune ◽

Viral Rna ◽

Entry Site ◽

Internal Ribosome Entry ◽

Viral Translation ◽

Rna Capping

The 5′ end of eukaryotic mRNA contains the type-1 (m7GpppNm) or type-2 (m7GpppNmNm) cap structure. Many viruses have evolved various mechanisms to develop their own capping enzymes (e.g. flavivirus and coronavirus) or to ‘steal’ caps from host mRNAs (e.g. influenza virus). Other viruses have developed ‘cap-mimicking’ mechanisms by attaching a peptide to the 5′ end of viral RNA (e.g. picornavirus and calicivirus) or by having a complex 5′ RNA structure (internal ribosome entry site) for translation initiation (e.g. picornavirus, pestivirus and hepacivirus). Here we review the diverse viral RNA capping mechanisms. Using flavivirus as a model, we summarize how a single methyltransferase catalyses two distinct N-7 and 2′-O methylations of viral RNA cap in a sequential manner. For antiviral development, a structural feature unique to the flavivirus methyltransferase was successfully used to design selective inhibitors that block viral methyltransferase without affecting host methyltransferases. Functionally, capping is essential for prevention of triphosphate-triggered innate immune activation; N-7 methylation is critical for enhancement of viral translation; and 2′-O methylation is important for subversion of innate immune response during viral infection. Flaviviruses defective in 2′-O methyltransferase are replicative, but their viral RNAs lack 2′-O methylation and are recognized and eliminated by the host immune response. Such mutant viruses could be rationally designed as live attenuated vaccines. This concept has recently been proved with Japanese encephalitis virus and dengue virus. The findings obtained with flavivirus should be applicable to other RNA viruses.

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IRES-targeting small molecule inhibits enterovirus 71 replication via allosteric stabilization of a ternary complex

Nature Communications ◽

10.1038/s41467-020-18594-3 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 1

Author(s):

Jesse Davila-Calderon ◽

Neeraj N. Patwardhan ◽

Liang-Yuan Chiu ◽

Andrew Sugarman ◽

Zhengguo Cai ◽

...

Keyword(s):

Rna Structure ◽

Drug Targets ◽

Ternary Complex ◽

Biochemical Characterization ◽

Internal Ribosomal Entry Site ◽

Enterovirus 71 ◽

Entry Site ◽

Stem Loop ◽

Viral Translation ◽

Significant Toxicity

Abstract Enterovirus 71 (EV71) poses serious threats to human health, particularly in Southeast Asia, and no drugs or vaccines are available. Previous work identified the stem loop II structure of the EV71 internal ribosomal entry site as vital to viral translation and a potential target. After screening an RNA-biased library using a peptide-displacement assay, we identify DMA-135 as a dose-dependent inhibitor of viral translation and replication with no significant toxicity in cell-based studies. Structural, biophysical, and biochemical characterization support an allosteric mechanism in which DMA-135 induces a conformational change in the RNA structure that stabilizes a ternary complex with the AUF1 protein, thus repressing translation. This mechanism is supported by pull-down experiments in cell culture. These detailed studies establish enterovirus RNA structures as promising drug targets while revealing an approach and mechanism of action that should be broadly applicable to functional RNA targeting.

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