Downregulation of Bamboo mosaic virus replication requires the 5′ apical hairpin stem loop structure and sequence of satellite RNA

ABSTRACT The replication of porcine circovirus type 1 (PCV1) is thought to occur by rolling-circle replication (RCR), whereby the introduction of a single-strand break generates a free 3′-hydroxyl group serving as a primer for subsequent DNA synthesis. The covalently closed, single-stranded genome of PCV1 replicates via a double-stranded replicative intermediate, and the two virus-encoded replication-associated proteins Rep and Rep′ have been demonstrated to be necessary for virus replication. However, although postulated to be involved in RCR-based virus replication, the mechanism of action of Rep and Rep′ is as yet unknown. In this study, the ability of PCV1 Rep and Rep′ to “nick” and “join” strand discontinuities within synthetic oligonucleotides corresponding to the origin of replication of PCV1 was investigated in vitro. Both proteins were demonstrated to be able to cleave the viral strand between nucleotides 7 and 8 within the conserved nonanucleotide motif (5′-TAGTATTAC-3′) located at the apex of a putative stem-loop structure. In addition, the Rep and Rep′ proteins of PCV1 were demonstrated to be capable of joining viral single-stranded DNA fragments, suggesting that these proteins also play roles in the termination of virus DNA replication. This joining activity was demonstrated to be strictly dependent on preceding substrate cleavage and the close proximity of origin fragments accomplished by base pairing in the stem-loop structure. The dual “nicking/joining” activities associated with PCV1 Rep and Rep′ are pivotal events underlying the RCR-based replication of porcine circoviruses in mammalian cells.

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Interferon-inducible ribonuclease ISG20 inhibits hepatitis B virus replication through directly binding to the epsilon stem-loop structure of viral RNA

PLoS Pathogens ◽

10.1371/journal.ppat.1006296 ◽

2017 ◽

Vol 13 (4) ◽

pp. e1006296 ◽

Cited By ~ 41

Author(s):

Yuanjie Liu ◽

Hui Nie ◽

Richeng Mao ◽

Bidisha Mitra ◽

Dawei Cai ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Virus Replication ◽

Viral Rna ◽

Loop Structure ◽

Stem Loop ◽

Stem Loop Structure ◽

B Virus

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The 5′ Untranslated Region of Alfalfa Mosaic Virus RNA 1 Is Involved in Negative-Strand RNA Synthesis

Journal of Virology ◽

10.1128/jvi.77.20.11284-11289.2003 ◽

2003 ◽

Vol 77 (20) ◽

pp. 11284-11289 ◽

Cited By ~ 12

Author(s):

A. Corina Vlot ◽

John F. Bol

Keyword(s):

Mosaic Virus ◽

Untranslated Region ◽

Rna Synthesis ◽

Alfalfa Mosaic Virus ◽

Loop Structure ◽

Stem Loop ◽

Genomic Rnas ◽

Negative Strand ◽

Stem Loop Structure ◽

Virus Rna

ABSTRACT The three genomic RNAs of alfalfa mosaic virus each contain a unique 5′ untranslated region (5′ UTR). Replacement of the 5′ UTR of RNA 1 by that of RNA 2 or 3 yielded infectious replicons. The sequence of a putative 5′ stem-loop structure in RNA 1 was found to be required for negative-strand RNA synthesis. A similar putative 5′ stem-loop structure is present in RNA 2 but not in RNA 3.

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RNA signals in the 3′ terminus of the genome of Equine arteritis virus are required for viral RNA synthesis

Journal of General Virology ◽

10.1099/vir.0.81750-0 ◽

2006 ◽

Vol 87 (7) ◽

pp. 1977-1983 ◽

Cited By ~ 18

Author(s):

Nancy Beerens ◽

Eric J. Snijder

Keyword(s):

Secondary Structure ◽

Virus Replication ◽

Rna Secondary Structure ◽

Rna Synthesis ◽

Full Length ◽

Equine Arteritis Virus ◽

Viral Rna ◽

Loop Structure ◽

Stem Loop ◽

Stem Loop Structure

RNA virus genomes contain cis-acting sequences and structural elements involved in virus replication. Both full-length and subgenomic negative-strand RNA synthesis are initiated at the 3′ terminus of the positive-strand genomic RNA of Equine arteritis virus (EAV). To investigate the molecular mechanism of EAV RNA synthesis, the RNA secondary structure of the 3′-proximal region of the genome was analysed by chemical and enzymic probing. Based on the RNA secondary structure model derived from this analysis, several deletions were engineered in a full-length cDNA copy of the viral genome. Two RNA domains were identified that are essential for virus replication and most likely play a key role in viral RNA synthesis. The first domain, located directly upstream of the 3′ untranslated region (UTR) (nt 12610–12654 of the genome), is mainly single-stranded but contains one small stem–loop structure. The second domain is located within the 3′ UTR (nt 12661–12690) and folds into a prominent stem–loop structure with a large loop region. The location of this stem–loop structure near the 3′ terminus of the genome suggests that it may act as a recognition signal during the initiation of minus-strand RNA synthesis.

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Construction of an Infectious cDNA Clone of Aichi Virus (a New Member of the Family Picornaviridae) and Mutational Analysis of a Stem-Loop Structure at the 5′ End of the Genome

Journal of Virology ◽

10.1128/jvi.75.17.8021-8030.2001 ◽

2001 ◽

Vol 75 (17) ◽

pp. 8021-8030 ◽

Cited By ~ 55

Author(s):

Jun Sasaki ◽

Yasuhiro Kusuhara ◽

Yoshimasa Maeno ◽

Nobumichi Kobayashi ◽

Teruo Yamashita ◽

...

Keyword(s):

Virus Replication ◽

Cdna Clone ◽

Rna Replication ◽

Infectious Cdna Clone ◽

Viral Rna ◽

Aichi Virus ◽

Loop Structure ◽

Stem Loop ◽

Stem Loop Structure ◽

The Family

ABSTRACT Aichi virus is the type species of a new genus,Kobuvirus, of the family Picornaviridae. In this study, we constructed a full-length cDNA clone of Aichi virus whose in vitro transcripts were infectious to Vero cells. During construction of the infectious cDNA clone, a novel sequence of 32 nucleotides was identified at the 5′ end of the genome. Computer-assisted prediction of the secondary structure of the 5′ end of the genome, including the novel sequence, suggested the formation of a stable stem-loop structure consisting of 42 nucleotides. The function of this stem-loop in virus replication was investigated using various site-directed mutants derived from the infectious cDNA clone. Our data indicated that correct folding of the stem-loop at the 5′ end of the positive strand, but not at the 3′ end of the negative strand, is critical for viral RNA replication. The primary sequence in the lower part of the stem was also suggested to be crucial for RNA replication. In contrast, nucleotide changes in the loop segment did not so severely reduce the efficiency of virus replication. A double mutant, in which both nucleotide stretches of the middle part of the stem were replaced by their complementary nucleotides, had efficient RNA replication and translation abilities but was unable to produce viruses. These results indicate that the stem-loop at the 5′ end of the Aichi virus genome is an element involved in both viral RNA replication and production of infectious virus particles.

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In Vitro Synthesis of Minus-Strand RNA by an Isolated Cereal Yellow Dwarf Virus RNA-Dependent RNA Polymerase Requires VPg and a Stem-Loop Structure at the 3′ End of the Virus RNA

Journal of Virology ◽

10.1128/jvi.01050-06 ◽

2006 ◽

Vol 80 (21) ◽

pp. 10743-10751 ◽

Cited By ~ 6

Author(s):

Toba A. M. Osman ◽

Robert H. A. Coutts ◽

Kenneth W. Buck

Keyword(s):

Rna Polymerase ◽

Mosaic Virus ◽

Rna Synthesis ◽

Loop Structure ◽

Minus Strand ◽

Stem Loop ◽

Stem Loop Structure ◽

Virus Rna ◽

Yellow Dwarf Virus

ABSTRACT Cereal yellow dwarf virus (CYDV) RNA has a 5′-terminal genome-linked protein (VPg). We have expressed the VPg region of the CYDV genome in bacteria and used the purified protein (bVPg) to raise an antiserum which was able to detect free VPg in extracts of CYDV-infected oat plants. A template-dependent RNA-dependent RNA polymerase (RdRp) has been produced from a CYDV membrane-bound RNA polymerase by treatment with BAL 31 nuclease. The RdRp was template specific, being able to utilize templates from CYDV plus- and minus-strand RNAs but not those of three unrelated viruses, Red clover necrotic mosaic virus, Cucumber mosaic virus, and Tobacco mosaic virus. RNA synthesis catalyzed by the RdRp required a 3′-terminal GU sequence and the presence of bVPg. Additionally, synthesis of minus-strand RNA on a plus-strand RNA template required the presence of a putative stem-loop structure near the 3′ terminus of CYDV RNA. The base-paired stem, a single-nucleotide (A) bulge in the stem, and the sequence of a tetraloop were all required for the template activity. Evidence was produced showing that minus-strand synthesis in vitro was initiated by priming by bVPg at the 3′ end of the template. The data are consistent with a model in which the RdRp binds to the stem-loop structure which positions the active site to recognize the 3′-terminal GU sequence for initiation of RNA synthesis by the addition of an A residue to VPg.

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A predicted stem-loop in coat protein-coding sequencing of tobacco vein banding mosaic virus is required for efficient replication

Phytopathology ◽

10.1094/phyto-10-20-0463-r ◽

2021 ◽

Author(s):

Zhi-Yong Yan ◽

Le Fang ◽

Xiao-Jie Xu ◽

De-Jie Cheng ◽

Cheng-Ming Yu ◽

...

Keyword(s):

Coat Protein ◽

Mosaic Virus ◽

Spontaneous Mutant ◽

Loop Structure ◽

Base Pairs ◽

Stem Loop ◽

Synonymous Substitutions ◽

Coding Sequence ◽

Stem Loop Structure ◽

Viral Accumulation

Potyviral Coat protein (CP) is involved in the replication and movement of potyviruses. However, little information is available on the roles of CP-coding sequence in potyviral infection. Here, we introduced synonymous substitutions to the codon c574g575c576 coding conserved residue arginine at position 192 (R192) of tobacco vein banding mosaic virus (TVBMV) CP. Substitution of the codon c574g575c576 to a574g575a576 or a574g575g576, but not c574g575a576, c574g575t576, or c574g575g576, reduced the replication, cell-to-cell movement, and accumulation of TVBMV in Nicotiana benthamiana plants, suggesting that c574 was critical for replication of TVBMV. Nucleotides 531 to 576 of the TVBMV CP-coding sequence were predicted to form a stem-loop structure, in which four consecutive c-g base pairs (C576-G531, c532-g575, c574-g533, and C534-G573) were located at the stem. Synonymous substitutions of R178-codon c532g533c534 to A532G533A534 and A532G533G534, but not c532g533a534, c532g533t534, or c532g533g534, reduced the replication levels, cell-to-cell, and systemic movement of TVBMV, suggesting that c532 was critical for TVBMV replication. Synonymous substitutions disrupting base pairs C576-G531 and C534-G573 did not affect viral accumulation. After three serial passage inoculation, the accumulation of spontaneous mutant viruses was restored and codons A532G533A534, A532G533G534, a574g575a576, or a574g575g576 of mutants was separately changed to C532G533A534, C532G533G534, C574g575a576, or C574g575g576. Synonymous mutation of R178 and R192 also reduced viral accumulation in N. tabacum plants. Therefore, we concluded that the two consecutive c532-g575 and c574-g533 base pairs played critical roles in TVBMV replication via maintaining the stability of stem-loop structure formed by nucleotides 531 to 576 of CP-coding sequence.

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