Suppression of RNA silencing by Flock house virus B2 protein is mediated through its interaction with the PAZ domain of Dicer.
FASEB J
; 23(6): 1845-57, 2009 Jun.
Article
em En
| MEDLINE
| ID: mdl-19193719
ABSTRACT
RNA silencing is a conserved pathway that functions as an antiviral mechanism. The majority of viruses encode silencing suppressors that interfere with siRNA- and miRNA-guided silencing pathways. The insect flock house virus B2 protein (FHVB2) functions as an RNAi silencing suppressor that inhibits siRNA biogenesis. Here, we describe the generation of a GFP silent sensor line (Sf21) and a GFP sensor line expressing FHVB2 to study RNAi suppression mechanisms. Overexpression of FHVB2 resulted in suppression of GFP-RNAi and resumption of GFP expression. Protein fractionation studies with FHVB2-transfected cells showed that FHVB2 associates with a high-molecular-weight complex of Dicer and dsRNA/siRNAs. Yeast two-hybrid and pulldown assays revealed an interaction between FHVB2 and Drosophila Dicer proteins that appeared to involve PAZ domains. To map the FHVB2 domains interacting with Dicer, we used a 17-residue C-terminal deletion mutant. RNAi suppression was reversed in cells transfected with the FHVB2 mutant as revealed by loss of GFP. Additional yeast two-hybrid and in vitro pulldown assays confirmed that the C-terminal region of FHVB2 was involved in the interaction with the PAZ domains of Dicers. These results thus reveal a novel interaction between FHVB2 and Dicer that leads to suppression of siRNA biogenesis.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Virais
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Nodaviridae
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Proteínas de Drosophila
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Interferência de RNA
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Ribonuclease III
Limite:
Animals
Idioma:
En
Ano de publicação:
2009
Tipo de documento:
Article