Adenovirus Vector-Derived VA-RNA-Mediated Innate Immune Responses

Machitani, Mitsuhiro; Yamaguchi, Tomoko; Shimizu, Kahori; Sakurai, Fuminori; Katayama, Kazufumi; Kono, Kenji; Mizuguchi, Hiroyuki

doi:10.3390/pharmaceutics3030338

Cited by 32 publications

(36 citation statements)

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“…VA-RNAs block PKR activation, leading to inhibition of phosphorylation of eIF2α and efficient replication of Ad815. In order to examine whether an increase in the copy numbers of VA-RNA by Dicer knockdown leads to a reduction in Ad-induced phosphorylation of eIF2α, HeLa-shDicer cells were cultured in the presence or absence of Dox (100 ng ml −1 ), followed by infection with WT-Ad or Sub720.…”

Section: Resultsmentioning

confidence: 99%

“…The adenovirus (Ad) genome also encodes two noncoding small RNAs, VA-RNA I (a major species) and VA-RNA II (a minor species), which are approximately 160 nucleotide-long noncoding RNAs that are transcribed by RNA polymerase III78. VA-RNA I is rapidly transcribed after infection and accumulates to very high levels, 10 8 molecules per cell.…”

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confidence: 99%

“…This blocking leads to the inhibition of eIF2α phosphorylation, leading to the maintenance of viral protein synthesis and the enhancement of Ad amplification1213. VA-RNA II also supports Ad replication8, although VA-RNA II has not been demonstrated to inhibit PKR activation14. VA-RNA I and II have a similar stem-loop structure via an intramolecular hydrogen bond.…”

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confidence: 99%

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Dicer functions as an antiviral system against human adenoviruses via cleavage of adenovirus-encoded noncoding RNA

Machitani

Sakurai

Wakabayashi

et al. 2016

Sci Rep

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In various organisms, including nematodes and plants, RNA interference (RNAi) is a defense system against virus infection; however, it is unclear whether RNAi functions as an antivirus system in mammalian cells. Rather, a number of DNA viruses, including herpesviruses, utilize post-transcriptional silencing systems for their survival. Here we show that Dicer efficiently suppresses the replication of adenovirus (Ad) via cleavage of Ad-encoding small RNAs (VA-RNAs), which efficiently promote Ad replication via the inhibition of eIF2α phosphorylation, to viral microRNAs (mivaRNAs). The Dicer knockdown significantly increases the copy numbers of VA-RNAs, leading to the efficient inhibition of eIF2α phosphorylation and the subsequent promotion of Ad replication. Conversely, overexpression of Dicer significantly inhibits Ad replication. Transfection with mivaRNA does not affect eIF2α phosphorylation or Ad replication. These results indicate that Dicer-mediated processing of VA-RNAs leads to loss of activity of VA-RNAs for enhancement of Ad replication and that Dicer functions as a defence system against Ad in mammalian cells.

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confidence: 99%

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Dicer functions as an antiviral system against human adenoviruses via cleavage of adenovirus-encoded noncoding RNA

Machitani

Sakurai

Wakabayashi

et al. 2016

Sci Rep

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“…injection of adenovirus vectors results in hepatic tropism, making it difficult to systemically deliver adenovirus vectors for metastasis treatment . Third, the strong immunogenic nature of adenoviruses seriously hampers their efficacy and safety in vivo . Fourth, clinical studies on oncolytic adenoviruses have suggested that several modifications are necessary to enhance the oncolytic activity and improve safety in the clinical setting .…”

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Recent advances in genetic modification of adenovirus vectors for cancer treatment

Yamamoto¹,

Nagasato²,

Yoshida

et al. 2017

Cancer Science

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Adenoviruses are widely used to deliver genes to a variety of cell types and have been used in a number of clinical trials for gene therapy and oncolytic virotherapy. However, several concerns must be addressed for the clinical use of adenovirus vectors. Selective delivery of a therapeutic gene by adenovirus vectors to target cancer is precluded by the widespread distribution of the primary cellular receptors. The systemic administration of adenoviruses results in hepatic tropism independent of the primary receptors. Adenoviruses induce strong innate and acquired immunity in vivo. Furthermore, several modifications to these vectors are necessary to enhance their oncolytic activity and ensure patient safety. As such, the adenovirus genome has been engineered to overcome these problems. The first part of the present review outlines recent progress in the genetic modification of adenovirus vectors for cancer treatment. In addition, several groups have recently developed cancer‐targeting adenovirus vectors by using libraries that display random peptides on a fiber knob. Pancreatic cancer‐targeting sequences have been isolated, and these oncolytic vectors have been shown by our group to be associated with a higher gene transduction efficiency and more potent oncolytic activity in cell lines, murine models, and surgical specimens of pancreatic cancer. In the second part of this review, we explain that combining cancer‐targeting strategies can be a promising approach to increase the clinical usefulness of oncolytic adenovirus vectors.

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“…Ad expresses Ad-encoded small noncoding RNAs (VA-RNAs), which inhibit production of miRNAs (15)(16)(17)(18)(19)(20)(21)(22)(23). VA-RNAs were originally demonstrated to inhibit double-stranded RNA-dependent kinase (PKR), leading to efficient translation of viral proteins (24)(25)(26)(27); more recently, however, VA-RNAs were reported to inhibit miRNA production (15)(16)(17)(18)(19)(20)(21)(22)(23). VA-RNAs inhibit Dicer expression by inhibiting nuclear export of Dicer mRNA, leading to suppression of miRNA production (15).…”

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MicroRNA miR-27 Inhibits Adenovirus Infection by Suppressing the Expression of SNAP25 and TXN2

Machitani

Sakurai

Wakabayashi

et al. 2017

J Virol

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Recent studies have reported that host microRNAs (miRNAs) regulate infections by several types of viruses via various mechanisms and that inhibition of the miRNA processing factors enhances or prevents viral infection. However, it has not been clarified whether these effects of miRNAs extend to adenovirus (Ad) infection. Here we show that miR-27a and -b efficiently inhibit infection with an Ad via the downregulation of SNAP25 and TXN2, which are members of the SNARE proteins and the thioredoxin family, respectively. Approximately 80% reductions in Ad genomic copy number were found in cells transfected with miR-27a/b mimics, whereas there were approximately 2.5-to 5-fold larger copy numbers of the Ad genome following transfection with miR-27a/b inhibitors. Microarray gene expression analysis and in silico analysis demonstrated that SNAP25 and TXN2 are target genes of miR-27a/b. A reporter assay using plasmids containing the 3= untranslated regions of the SNAP25 and TXN2 genes showed that miR-27a/b directly suppressed SNAP25 and TXN2 expression through posttranscriptional gene silencing. Knockdown of SNAP25 led to a significant inhibition of Ad entry into cells. Knockdown of TXN2 induced cell cycle arrest at G 1 phase, leading to a reduction in Ad replication. In addition, overexpression of Ad-encoded small noncoding RNAs (VA-RNAs) restored the miR-27a/b-mediated reduction in infection level with a VA-RNA-lacking Ad mutant due to the VA-RNA-mediated inhibition of miR-27a/b expression. These results indicate that miR-27a and -b suppress SNAP25 and TXN2 expression via posttranscriptional gene silencing, leading to efficient suppression of Ad infection.IMPORTANCE Adenovirus (Ad) is widely used as a platform for replication-incompetent Ad vectors (Adv) and replication-competent oncolytic Ad (OAd) in gene therapy and virotherapy. Regulation of Ad infection is highly important for efficient gene therapies using both Adv and OAd. In this study, we demonstrate that miR-27a and -b, which are widely expressed in host cells, suppress SNAP25 and TXN2 expression through posttranscriptional gene silencing. Suppression of SNAP25 and TXN2 expression leads to inhibition of Ad entry into cells and to cell cycle arrest, respectively, leading to efficient suppression of Ad infection. Our findings provide important clues to the improvement of gene therapies using both Adv and OAd.KEYWORDS SNAP25, TXN2, adenoviruses, miR-27, posttranscriptional gene silencing

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Adenovirus Vector-Derived VA-RNA-Mediated Innate Immune Responses

Cited by 32 publications

References 100 publications

Dicer functions as an antiviral system against human adenoviruses via cleavage of adenovirus-encoded noncoding RNA

Dicer functions as an antiviral system against human adenoviruses via cleavage of adenovirus-encoded noncoding RNA

Recent advances in genetic modification of adenovirus vectors for cancer treatment

MicroRNA miR-27 Inhibits Adenovirus Infection by Suppressing the Expression of SNAP25 and TXN2

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