Replication, Recombination, and Symptom-Modulation Properties of the Satellite RNAs of Turnip Crinkle Virus

Satellite RNAs and satellite viruses are extraviral components that can affect either the pathogenicity, the accumulation, or both of their associated viruses while themselves being dependent on the associated viruses as helper viruses for their infection. Most of these satellite RNAs are noncoding RNAs, and in many cases, have been shown to alter the interaction of their helper viruses with their hosts. In only a few cases have the functions of these satellite RNAs in such interactions been studied in detail. In particular, work on the satellite RNAs of Cucumber mosaic virus and Turnip crinkle virus have provided novel insights into RNAs functioning as noncoding RNAs. These effects are described and potential roles for satellite RNAs in the processes involved in symptom intensification or attenuation are discussed. In most cases, models describing these roles involve some aspect of RNA silencing or its suppression, either directly or indirectly involving the particular satellite RNA.

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Structural Plasticity and Rapid Evolution in a Viral RNA Revealed by In Vivo Genetic Selection

Journal of Virology ◽

10.1128/jvi.02060-08 ◽

2008 ◽

Vol 83 (2) ◽

pp. 927-939 ◽

Cited By ~ 14

Author(s):

Rong Guo ◽

Wai Lin ◽

Jiuchun Zhang ◽

Anne E. Simon ◽

David B. Kushner

Keyword(s):

Sequence Similarity ◽

Rapid Evolution ◽

Genetic Selection ◽

Upstream Sequence ◽

Turnip Crinkle Virus ◽

In Planta ◽

High Sequence Similarity ◽

Stem Loop ◽

Satellite Rnas

ABSTRACT Satellite RNAs usually lack substantial homology with their helper viruses. The 356-nucleotide satC of Turnip crinkle virus (TCV) is unusual in that its 3′-half shares high sequence similarity with the TCV 3′ end. Computer modeling, structure probing, and/or compensatory mutagenesis identified four hairpins and three pseudoknots in this TCV region that participate in replication and/or translation. Two hairpins and two pseudoknots have been confirmed as important for satC replication. One portion of the related 3′ end of satC that remains poorly characterized corresponds to juxtaposed TCV hairpins H4a and H4b and pseudoknot ψ3, which are required for the TCV-specific requirement of translation (V. A. Stupina et al., RNA 14:2379-2393, 2008). Replacement of satC H4a with randomized sequence and scoring for fitness in plants by in vivo genetic selection (SELEX) resulted in winning sequences that contain an H4a-like stem-loop, which can have additional upstream sequence composing a portion of the stem. SELEX of the combined H4a and H4b region in satC generated three distinct groups of winning sequences. One group models into two stem-loops similar to H4a and H4b of TCV. However, the selected sequences in the other two groups model into single hairpins. Evolution of these single-hairpin SELEX winners in plants resulted in satC that can accumulate to wild-type (wt) levels in protoplasts but remain less fit in planta when competed against wt satC. These data indicate that two highly distinct RNA conformations in the H4a and H4b region can mediate satC fitness in protoplasts.

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3′-End Stem-Loops of the Subviral RNAs Associated with Turnip Crinkle Virus Are Involved in Symptom Modulation and Coat Protein Binding

Journal of Virology ◽

10.1128/jvi.74.14.6528-6537.2000 ◽

2000 ◽

Vol 74 (14) ◽

pp. 6528-6537 ◽

Cited By ~ 13

Author(s):

Jianlong Wang ◽

Anne E. Simon

Keyword(s):

Coat Protein ◽

Turnip Crinkle Virus ◽

Complementary Strand ◽

Satellite Rna ◽

Stem Loop ◽

Subviral Rnas ◽

Translation Initiation Codon ◽

Defective Interfering Rna ◽

Symptom Modulation ◽

Interfering Rna

ABSTRACT Many plant RNA viruses are associated with one or more subviral RNAs. Two subviral RNAs, satellite RNA C (satC) and defective interfering RNA G (diG) intensify the symptoms of their helper, turnip crinkle virus (TCV). However, when the coat protein (CP) of TCV was replaced with that of the related Cardamine chlorotic fleck virus (CCFV), both subviral RNAs attenuated symptoms of the hybrid virus TCV-CPCCFV. In contrast, when the translation initiation codon of the TCV CP was altered to ACG and reduced levels of CP were synthesized, satC attenuated symptoms while diG neither intensified nor attenuated symptoms. The determinants for this differential symptom modulation were previously localized to the 3′-terminal 100 bases of the subviral RNAs, which contain six positional differences (Q. Kong, J.-W. Oh, C. D. Carpenter, and A. E. Simon, Virology 238:478–485, 1997). In the current study, we have determined that certain sequences within the 3′-terminal stem-loop structures of satC and diG, which also serve as promoters for complementary strand synthesis, are critical for symptom modulation. Furthermore, the ability to attenuate symptoms was correlated with weakened binding of TCV CP to the hairpin structure.

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Fitness of a Turnip Crinkle Virus Satellite RNA Correlates with a Sequence-Nonspecific Hairpin and Flanking Sequences That Enhance Replication and Repress the Accumulation of Virions

Journal of Virology ◽

10.1128/jvi.77.14.7880-7889.2003 ◽

2003 ◽

Vol 77 (14) ◽

pp. 7880-7889 ◽

Cited By ~ 14

Author(s):

Xiaoping Sun ◽

Anne E. Simon

Keyword(s):

Systemic Infection ◽

Turnip Crinkle Virus ◽

Satellite Rna ◽

Structural Requirements ◽

Sense Orientation ◽

Flanking Sequences ◽

Satellite Rnas ◽

Systematic Evolution ◽

Base Replacement ◽

Exponential Enrichment

ABSTRACT satC, a satellite RNA associated with Turnip crinkle virus (TCV), enhances the ability of the virus to colonize plants by interfering with stable virion accumulation (F. Zhang and A. E. Simon, unpublished data). Previous results suggested that the motif1-hairpin (M1H), a replication enhancer on minus strands, forms a plus-strand hairpin flanked by CA-rich sequence that may be involved in enhancing systemic infection (G. Zhang and A. E. Simon, J. Mol. Biol. 326:35-48, 2003). In this study, sequence and structural requirements of the M1H were further assayed by replacing the 28-base M1H with 10 random bases and then subjecting the pool of satellite RNA to functional selection in plants. Unlike previous results with 28-base replacement sequences (G. Zhang and A. E. Simon, J. Mol. Biol. 326:35-48, 2003), only a few of the 10-base SELEX (systematic evolution of ligands by exponential enrichment) assay winners contained short motifs in their minus-sense orientation that were similar to TCV replication elements. However, all second- and third-round winning replacement sequences folded into hairpins flanked by CA-rich sequence predicted to be more stable on plus strands than minus strands. Plus strands of several of the most fit satellite RNAs contained insertions of CA-rich sequence at the base of their hairpins whose presence correlated with enhanced replication and reduced detection of virions. Deletion of the M1H resulted in no detectable virions despite very low satellite accumulation. These results support the hypothesis that a sequence-nonspecific plus-strand hairpin brings together flanking CA-rich sequences in the M1H region that confers fitness to satC by reducing the accumulation of stable virions.

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Molecular analysis of Greek isolates of cucumber mosaic virus from vegetables shows a low prevalence of satellite RNAs and suggests the presence of host-associated virus strains

Archives of Virology ◽

10.1007/s00705-021-05115-w ◽

2021 ◽

Author(s):

Christos A. Valachas ◽

Ioannis A. Giantsis ◽

Kyriaki Sareli ◽

Stephan Winter ◽

Eleanna Zelezniakof ◽

...

Keyword(s):

Cucumber Mosaic Virus ◽

Mosaic Virus ◽

Molecular Analysis ◽

Satellite Rnas ◽

Low Prevalence ◽

Virus Strains

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Molecular modeling and interaction between Arabidopsis sulfite oxidase and the GW motif of Turnip crinkle virus coat protein

Virology ◽

10.1016/j.virol.2020.08.013 ◽

2020 ◽

Vol 551 ◽

pp. 64-74

Author(s):

Chao Wu ◽

Srinivasaraghavan Kannan ◽

Chandra S. Verma ◽

Kunchithapadam Swaminathan ◽

Sek-Man Wong

Keyword(s):

Molecular Modeling ◽

Coat Protein ◽

Sulfite Oxidase ◽

Turnip Crinkle Virus ◽

Virus Coat Protein ◽

Virus Coat

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Characterization of cucumber mosaic virus and its satellite RNAs associated with tomato lethal necrosis in Serbia

European Journal of Plant Pathology ◽

10.1007/s10658-021-02241-8 ◽

2021 ◽

Author(s):

Ivana Stanković ◽

Ana Vučurović ◽

Katarina Zečević ◽

Branka Petrović ◽

Dušan Nikolić ◽

...

Keyword(s):

Cucumber Mosaic Virus ◽

Mosaic Virus ◽

Satellite Rnas

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Requirement for Host RNA-Silencing Components and the Virus-Silencing Suppressor when Second-Site Mutations Compensate for Structural Defects in the 3′ Untranslated Region

Journal of Virology ◽

10.1128/jvi.01566-15 ◽

2015 ◽

Vol 89 (22) ◽

pp. 11603-11618 ◽

Cited By ~ 3

Author(s):

Maitreyi Chattopadhyay ◽

Vera A. Stupina ◽

Feng Gao ◽

Christine R. Szarko ◽

Micki M. Kuhlmann ◽

...

Keyword(s):

Small Rnas ◽

Rna Silencing ◽

Untranslated Region ◽

Higher Order ◽

Order Structure ◽

Silencing Suppressor ◽

Turnip Crinkle Virus ◽

Site Mutation ◽

Primary Mutation ◽

Positive Strand Rna

ABSTRACTTurnip crinkle virus (TCV) contains a structured 3′ region with hairpins and pseudoknots that form a complex network of noncanonical RNA:RNA interactions supporting higher-order structure critical for translation and replication. We investigated several second-site mutations in the p38 coat protein open reading frame (ORF) that arose in response to a mutation in the asymmetric loop of a critical 3′ untranslated region (UTR) hairpin that disrupts local higher-order structure. All tested second-site mutations improved accumulation of TCV in conjunction with a partial reversion of the primary mutation (TCV-rev1) but had neutral or a negative effect on wild-type (wt) TCV or TCV with the primary mutation. SHAPE (selective 2′-hydroxylacylation analyzed byprimerextension) structure probing indicated that these second-site mutations reside in an RNA domain that includes most of p38 (domain 2), and evidence for RNA:RNA interactions between domain 2 and 3′UTR-containing domain 1 was found. However, second-site mutations were not compensatory in the absence of p38, which is also the TCV silencing suppressor, or indcl-2/dcl4orago1/ago2backgrounds. One second-site mutation reduced silencing suppressor activity of p38 by altering one of two GW motifs that are required for p38 binding to double-stranded RNAs (dsRNAs) and interaction with RNA-induced silencing complex (RISC)-associated AGO1/AGO2. Another second-site mutation substantially reduced accumulation of TCV-rev1 in the absence of p38 or DCL2/DCL4. We suggest that the second-site mutations in the p38 ORF exert positive effects through a similar downstream mechanism, either by enhancing accumulation of beneficial DCL-produced viral small RNAs that positively regulate the accumulation of TCV-rev1 or by affecting the susceptibility of TCV-rev1 to RISC loaded with viral small RNAs.IMPORTANCEGenomes of positive-strand RNA viruses fold into high-order RNA structures. Viruses with mutations in regions critical for translation and replication often acquire second-site mutations that exert a positive compensatory effect through reestablishment of canonical base pairing with the altered region. In this study, two distal second-site mutations that individually arose in response to a primary mutation in a critical 3′ UTR hairpin in the genomic RNA of turnip crinkle virus did not directly interact with the primary mutation. Although different second-site changes had different attributes, compensation was dependent on the production of the viral p38 silencing suppressor and on the presence of silencing-required DCL and AGO proteins. Our results provide an unexpected connection between a 3′ UTR primary-site mutation proposed to disrupt higher-order structure and the RNA-silencing machinery.

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