Abstract
Arrays of >5,000 Saccharomyces cerevisiae proteins were screened to identify proteins that can preferentially bind a small RNA hairpin that contains a clamped adenine motif (CAM). A CAM is required for the replication of Brome Mosaic Virus (BMV), a plant-infecting RNA virus that can replicate in S. cerevisiae. Several hits were selected for further characterization in Nicotiana benthamiana. Pseudouridine Synthase 4 (Pus4) and the Actin Patch Protein 1 (App1) modestly reduced BMV genomic plus-strand RNA accumulation, but dramatically inhibited BMV systemic spread in plants. Pus4 also prevented the encapsidation of a BMV RNA in plants and the reassembly of BMV virions in vitro. These results demonstrate the feasibility of using proteome arrays to identify specific RNA-binding proteins for antiviral activities. Furthermore, the effects of Pus4 suggest that the CAM-containing RNA motif provides a regulatory link between RNA replication and encapsidation.
Publication types
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Comparative Study
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Research Support, N.I.H., Extramural
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Amino Acid Motifs / genetics
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Blotting, Northern
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Bromovirus / genetics*
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Bromovirus / physiology
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Intramolecular Transferases / genetics
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Intramolecular Transferases / metabolism
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Intramolecular Transferases / pharmacology*
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Microfilament Proteins / genetics
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Microfilament Proteins / metabolism
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Microfilament Proteins / pharmacology*
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Nicotiana / virology
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Protein Array Analysis
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RNA, Viral / metabolism*
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RNA-Binding Proteins / genetics
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RNA-Binding Proteins / metabolism
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RNA-Binding Proteins / pharmacology*
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism
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Saccharomyces cerevisiae Proteins / pharmacology*
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Virus Assembly / drug effects
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Virus Replication / drug effects*
Substances
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Microfilament Proteins
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RNA, Viral
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RNA-Binding Proteins
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Saccharomyces cerevisiae Proteins
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APP1 protein, S cerevisiae
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Intramolecular Transferases
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tRNA-pseudouridine synthase I