MicroRNA Targeting of Neurotropic Flavivirus: Effective Control of Virus Escape and Reversion to Neurovirulent Phenotype

Heiss, Brian; Maximova, Olga A.; Thach, Dzung C.; Speicher, James M.; Pletnev, Alexander G.

doi:10.1128/jvi.07125-11

Cited by 56 publications

(93 citation statements)

References 37 publications

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“…Here, our results showed that synonymous mutations in the target sites of hsa-miR-296-5p in EV71 strain BrCr could facilitate virus infection and impair the inhibitory effects of hsa-miR-296-5p on the virus. This finding also suggests that virus escape from miRNA-mediated suppression can occur through mutations within the seed region of the miRNA target sequence, which is in line with data obtained for other viruses (53)(54)(55)(56). Insertions in the miR-9, miR-124a, miR-128a, miR-218, or let-7c target sites of the neurovirulent chimeric tickborne encephalitis/dengue virus (TBEV/DEN4) genome were found to be sufficient to prevent the development of otherwise lethal encephalitis in mice infected with a high dose of virus.…”

Section: Discussionsupporting

confidence: 78%

Human MicroRNA hsa-miR-296-5p Suppresses Enterovirus 71 Replication by Targeting the Viral Genome

Zheng

Wang

et al. 2013

J Virol

164

155

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Enterovirus 71 (EV71) has emerged as a major cause of neurological disease following the near eradication of poliovirus. Accumulating evidence suggests that mammalian microRNAs (miRNAs), a class of noncoding RNAs of 18 to 23 nucleotides (nt) with important regulatory roles in many cellular processes, participate in host antiviral defenses. However, the roles of miRNAs in EV71 infection and pathogenesis are still unclear. Here, hsa-miR-296-5p expression was significantly increased in EV71-infected human cells. As determined by virus titration, quantitative real-time PCR (qRT-PCR), and Western blotting, overexpression of hsa-miR-296-5p inhibited, while inhibition of endogenous hsa-miR-296-5p facilitated, EV71 infection. Additionally, two potential hsa-miR-296-5p targets (nt 2115 to 2135 and nt 2896 to 2920) located in the EV71 genome (strain BrCr) were bioinformatically predicted and validated by luciferase reporter assays and Western blotting. Genomic alignment of various EV71 strains revealed synonymous mutations in hsa-miR-296-5p target sequences. Furthermore, the introduction of synonymous mutations into the EV71 BrCr genome by site-directed mutagenesis impaired the viral inhibitory effects of hsa-miR-296-5p and facilitated mutant virus infection. Meanwhile, compensatory mutations in corresponding hsa-miR-296-5p target sequences of the EV71 HeN strain (GenBank accession number JN256064) restored the inhibitory effects of the miRNA. These results indicate that hsa-miR-296-5p inhibits EV71 replication by targeting the viral genome. Our findings support the notion that cellular miRNAs can inhibit virus infection and that the virus mutates to escape suppression by cellular miRNAs.

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Section: Discussionsupporting

confidence: 78%

Human MicroRNA hsa-miR-296-5p Suppresses Enterovirus 71 Replication by Targeting the Viral Genome

Zheng

Wang

et al. 2013

J Virol

164

155

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“…Previous reports indicate that increasing the number of miRNA targets within a genome is the most efficacious way to silence gene expression (22,23). In the present study, an enhanced effect on attenuating CVB3 virulence in vitro was not evident when inserting three target sequence copies (miR-206 plus miR-133) rather than just a single copy of miR-206.…”

Section: Discussionmentioning

confidence: 41%

Coxsackievirus B3 Engineered To Contain MicroRNA Targets for Muscle-Specific MicroRNAs Displays Attenuated Cardiotropic Virulence in Mice

Yao

Wang

et al. 2015

J Virol

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Coxsackievirus B3 (CVB3) is trophic for cardiac tissue and is a major causative agent for viral myocarditis, where local viral replication in the heart may lead to heart failure or even death. Recent studies show that inserting microRNA target sequences into the genomes of certain viruses can eradicate these viruses within local host tissues that specifically express the cognate microRNA. Here, we demonstrated both in vitro and in vivo that incorporating target sequences for miRNA-133 and -206 into the 5= untranslated region of the CVB3 genome ameliorated CVB3 virulence in skeletal muscle and myocardial cells that specifically expressed the cognate cellular microRNAs. Compared to wild-type CVB3, viral replication of the engineered CVB3 was attenuated in human TE671 (rhabdomyosarcoma) and L6 (skeletal muscle) cell lines in vitro that expressed high levels of miRNA-206. In the in vivo murine CVB3-infection model, viral replication of the engineered CVB3 was attenuated specifically in the heart that expressed high levels of both miRNAs, but not in certain tissues, which allowed the host to retain the ability to induce a strong and protective humoral immune response against CVB3. The results of this study suggest that a microRNA-targeting strategy to control CVB3 tissue tropism and pathogenesis may be useful for viral attenuation and vaccine development. IMPORTANCECoxsackievirus B3 (CVB3) is a major causative agent for viral myocarditis, and viral replication in the heart may lead to heart failure or even death. Limiting CVB3 replication within the heart may be a promising strategy to decrease CVB3 pathogenicity. miRNAs are ϳ21-nucleotide-long, tissue-specific endogenous small RNA molecules that posttranscriptionally regulate gene expression by imperfectly binding to the 3= untranslated region (UTR), the 5= UTR, or the coding region within a gene. In our study, muscle-specific miRNA targets (miRT) were incorporated into the CVB3 genome. Replication of the engineered viruses was restricted in the important heart tissue of infected mice, which reduced cardiac pathology and increased mouse survival. Meanwhile, replication ability was retained in other tissues, thus inducing a strong humoral immune response and providing long-term protection against CVB3 rechallenge. This study suggests that a microRNA-targeting strategy can potentially control CVB3 tissue tropism and pathogenesis and may be useful for viral attenuation and vaccine development. C oxsackievirus B3 (CVB3) is a clinically relevant infectious agent that causes viral myocarditis, pancreatitis, and aseptic meningitis. CVB3 is cardiotropic and efficiently replicates in the heart. We and others have shown in various animal models that CVB3 replication induces a direct cytopathic effect on infected cells, leading to tissue damage (1, 2). At present, there is no effective therapy against CVB3, and strategies to develop such a therapy are greatly desired (3, 4). Limiting CVB3 replication within the heart may be a promising strategy to decrease CVB3 pathoge...

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“…The diverse expression profiles of miRNAs provide a unique tool by which virus transcription can be controlled through exploitation of the small RNA silencing potential. Previous work from our group and others have demonstrated that insertion of complementary miRNA target sequences into the genome of a virus transforms that miRNA into an effective short interfering RNA (siRNA) (28)(29)(30)(31)(32)(33)(34). For example, incorporating miR-93 target sites into the nucleoprotein (NP) gene of IAV demonstrated that virus replication was inversely proportional to miR-93 levels both in vitro and in vivo (28).…”

mentioning

confidence: 99%

Hematopoietic-specific targeting of influenza A virus reveals replication requirements for induction of antiviral immune responses

Langlois¹,

Varble²,

Chua³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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A coordinated innate and adaptive immune response, orchestrated by antigen presenting cells (APCs), is required for effective clearance of influenza A virus (IAV). Although IAV primarily infects epithelial cells of the upper respiratory tract, APCs are also susceptible. To determine if virus transcription in these cells is required to generate protective innate and adaptive immune responses, we engineered IAV to be selectively attenuated in cells of hematopoietic origin. Incorporation of hematopoietic-specific miR-142 target sites into the nucleoprotein of IAV effectively silenced virus transcription in APCs, but had no significant impact in lung epithelial cells. Here we demonstrate that inhibiting IAV replication in APCs in vivo did not alter clearance, or the generation of IAV-specific CD8 T cells, suggesting that cross-presentation is sufficient for cytotoxic T lymphocyte activation. In contrast, loss of in vivo virus infection, selectively in APCs, resulted in a significant reduction of retinoic acid-inducible gene I-dependent type I IFN (IFN-I). These data implicate the formation of virus replication intermediates in APCs as the predominant trigger of IFN-I in vivo. Taking these data together, this research describes a unique platform to study the host response to IAV and provides insights into the mechanism of antigen presentation and the induction of IFN-I.

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MicroRNA Targeting of Neurotropic Flavivirus: Effective Control of Virus Escape and Reversion to Neurovirulent Phenotype

Cited by 56 publications

References 37 publications

Human MicroRNA hsa-miR-296-5p Suppresses Enterovirus 71 Replication by Targeting the Viral Genome

Human MicroRNA hsa-miR-296-5p Suppresses Enterovirus 71 Replication by Targeting the Viral Genome

Coxsackievirus B3 Engineered To Contain MicroRNA Targets for Muscle-Specific MicroRNAs Displays Attenuated Cardiotropic Virulence in Mice

Hematopoietic-specific targeting of influenza A virus reveals replication requirements for induction of antiviral immune responses

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