記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/s41586-020-2231-y

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担当区分：最終著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.21769/BioProtoc.2673

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担当区分：筆頭著者,　責任著者   記述言語：英語  

DOI： 10.1038/s41477-017-0028-2

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

Viruses often encode viral silencing suppressors (VSSs) to counteract the hosts' RNA silencing activity. The cricket paralysis virus 1A protein (CrPV-1A) is a unique VSS that binds to a specific Argonaute protein (Ago)-the core of the RNA-induced silencing complex (RISC)-in insects to suppress its target cleavage reaction. However, the precise molecular mechanism of CrPV-1A action remains unclear. Here we utilized biochemical and single-molecule imaging approaches to analyze the effect of CrPV-1A during target recognition and cleavage by Drosophila Ago2-RISC. Our results suggest that CrPV-1A obstructs the initial target searching by Ago2-RISC via base pairing in the seed region. The combination of biochemistry and single-molecule imaging may help to pave the way for mechanistic understanding of VSSs with diverse functions.

DOI： 10.1093/nar/gkx748

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE  

The bipartite genomic RNAs of red clover necrotic mosaic virus (RCNMV) lack a 5' cap and a 3' poly(A) tail. RNA1 encodes viral replication proteins, and RNA2 encodes a movement protein (MP). These proteins are translated in a cap-independent manner. We previously identified two cis-acting RNA elements that cooperatively recruit eukaryotic translation initiation factor (eIF) complex eIF4F or eIFiso4F to RNA1. Such cis-acting RNA elements and host factors have not been identified in RNA2. Here we found that translation of RNA1 was significantly compromised in Arabidopsis thaliana carrying eif4f mutation. RNA1 replicated efficiently in eifiso4fl mutants, suggesting vigorous translation of the replication proteins from RNA1 in the plants. In contrast, MP accumulation was decreased in eifiso4f1 mutants but not in eif4f mutants. Collectively, these results suggest that RCNMV uses different eIF complexes for translation of its bipartite genomic RNAs, which may contribute to fine-tuning viral gene expression during infection.

DOI： 10.1016/j.virol.2017.06.015

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担当区分：最終著者,　責任著者   記述言語：英語   掲載種別：論文集(書籍)内論文  

DOI： 10.1007/978-1-4939-7165-7_4

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担当区分：責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATURE PUBLISHING GROUP  

It remains unclear how post-transcriptional gene silencing (PTGS) in plants discriminates aberrant RNAs from canonical messenger RNAs (mRNAs). The key step of plant PTGS is the conversion of aberrant RNAs into double-stranded RNAs by RNA-DEPENDENT RNA POLYMERASE6 (RDR6). Here, we show that RDR6 itself selects aberrant poly(A)-less mRNAs over canonical polyadenylated mRNAs as templates at the initiation step of complementary strand synthesis. This mechanism can be viewed as an innate safeguard against 'self-attack' by PTGS.

DOI： 10.1038/nplants.2017.36

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担当区分：筆頭著者   記述言語：英語  

DOI： 10.1016/j.tcb.2015.07.011

PubMed

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：CELL PRESS  

miRNAs silence their complementary target mRNAs by translational repression as well as by poly(A) shortening and mRNA decay. In Drosophila, miRNAs are typically incorporated into Argonaute1 (Ago1) to form the effector complex called RNA-induced silencing complex (RISC). Ago1-RISC associates with a scaffold protein GW182, which recruits additional silencing factors. We have previously shown that miRNAs repress translation initiation by blocking formation of the 48S and 80S ribosomal complexes. However, it remains unclear how ribosome recruitment is impeded. Here, we examined the assembly of translation initiation factors on the target mRNA under repression. We show that Ago1-RISC induces dissociation of eIF4A, a DEAD-box RNA helicase, from the target mRNA without affecting 50 cap recognition by eIF4E in a manner independent of GW182. In contrast, direct tethering of GW182 promotes dissociation of both eIF4E and eIF4A. We propose that miRNAs act to block the assembly of the eIF4F complex during translation initiation.

DOI： 10.1016/j.molcel.2014.09.004

PubMed

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担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   出版者・発行元：Elsevier BV  

DOI： 10.1016/j.molcel.2013.10.033

PubMed

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担当区分：責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/embor.2013.73

PubMed

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

Viruses employ an alternative translation mechanism to exploit cellular resources at the expense of host mRNAs and to allow preferential translation. Plant RNA viruses often lack both a 5' cap and a 3' poly(A) tail in their genomic RNAs. Instead, cap-independent translation enhancer elements (CITEs) located in the 3' untranslated region (UTR) mediate their translation. Although eukaryotic translation initiation factors (eIFs) or ribosomes have been shown to bind to the 3' CITEs, our knowledge is still limited for the mechanism, especially for cellular factors. Here, we searched for cellular factors that stimulate the 3' CITE-mediated translation of Red clover necrotic mosaic virus (RCNMV) RNA1 using RNA aptamer-based one-step affinity chromatography, followed by mass spectrometry analysis. We identified the poly(A)-binding protein (PABP) as one of the key players in the 3' CITE-mediated translation of RCNMV RNA1. We found that PABP binds to an A-rich sequence (ARS) in the viral 3' UTR. The ARS is conserved among dianthoviruses. Mutagenesis and a tethering assay revealed that the PABP-ARS interaction stimulates 3' CITE-mediated translation of RCNMV RNAl. We also found that both the ARS and 3' CITE are important for the recruitment of the plant eIF4F and eIFiso4F factors to the 3 ' UTR and of the 40S ribosomal subunit to the viral mRNA. Our results suggest that dianthoviruses have evolved the ARS and 3' CITE as substitutes for the 3' poly(A) tail and the 5' cap of eukaryotic mRNAs for the efficient recruitment of eIFs, PABP, and ribosomes to the uncapped/nonpolyadenylated viral mRNA.

DOI： 10.1128/JVI.00538-12

PubMed

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE  

Programmed - 1 ribosomal frameshifting (- 1 PRF) is one viral translation strategy to express overlapping genes in positive-strand RNA viruses. Red clover necrotic mosaic virus ( RCNMV) uses this strategy to express its replicase component protein p88. In this study, we used a cell-free translation system to map cis-acting RNA elements required for 1 PRF. Our results show that a small stem-loop structure adjacent to the cap-independent translation element in the 3' untranslated region (UTR) of RCNMV RNA1 is required for - 1 PRF. Site-directed mutagenesis experiments suggested that this stem-loop regulates - 1 PRF via base-pairing with complementary sequences in a bulged stem-loop adjacent to the shifty site. The existence of RNA elements responsible for - 1 PRF and the cap-independent translation of replicase proteins in the 3' UTR of RNA1 might be important for switching translation to replication and for regulating the ratio of p88 to p27. (C) 2011 Published by Elsevier

DOI： 10.1016/j.virol.2011.05.012

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DOI： 10.1016/j.virol.2011.02.017

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

Recognition of RNA templates by viral replicase proteins is one of the key steps in the replication process of all RNA viruses. However, the mechanisms underlying this phenomenon, including primary RNA elements that are recognized by the viral replicase proteins, are not well understood. Here, we used aptamer pulldown assays with membrane fractionation and protein-RNA coimmunoprecipitation in a cell-free viral translation/replication system to investigate how viral replicase proteins recognize the bipartite genomic RNAs of the Red clover necrotic mosaic virus (RCNMV). RCNMV replicase proteins bound specifically to a Y-shaped RNA element (YRE) located in the 3' untranslated region (UTR) of RNA2, which also interacted with the 480-kDa replicase complexes that contain viral and host proteins. The replicase-YRE interaction recruited RNA2 to the membrane fraction. Conversely, RNA1 fragments failed to interact with the replicase proteins supplied in trans. The results of protein-RNA coimmunoprecipitation assays suggest that RNA1 interacts with the replicase proteins coupled with their translation. Thus, the initial template recognition mechanisms employed by the replicase differ between RCNMV bipartite genomic RNAs and RNA elements are primary determinants of the differential replication mechanism.

DOI： 10.1128/JVI.01754-10

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virol.2010.05.022

PubMed

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virol.2009.05.037

PubMed

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1128/JVI.01027-08

PubMed

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virol.2008.02.004

PubMed

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virol.2007.07.016

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1128/JVI.80.8.3781-3791.2006

PubMed

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DOI： 10.2222/jsv.59.179

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