Journal
EMBO JOURNAL
Volume 31, Issue 2, Pages 267-278Publisher
WILEY
DOI: 10.1038/emboj.2011.395
Keywords
ARGONAUTE; cyclophilin 40 (CYP40); HSP90; posttranscriptional gene silencing (PTGS); RNA-induced silencing complex (RISC)
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Funding
- Ministry of Education, Culture, Sports, Science and Technology, Japan
- PRESTO of JST
- Grants-in-Aid for Scientific Research [21380034] Funding Source: KAKEN
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Posttranscriptional gene silencing is mediated by RNA-induced silencing complexes (RISCs) that contain AGO proteins and single-stranded small RNAs. The assembly of plant AGO1-containing RISCs depends on the molecular chaperone HSP90. Here, we demonstrate that cyclophilin 40 (CYP40), protein phosphatase 5 (PP5), and several other proteins with the tetratricopeptide repeat (TPR) domain associates with AGO1 in an HSP90-dependent manner in extracts of evacuolated tobacco protoplasts (BYL). Intriguingly, CYP40, but not the other TPR proteins, could form a complex with small RNA duplex-bound AGO1. Moreover, CYP40 that was synthesized by in-vitro translation using BYL uniquely facilitated binding of small RNA duplexes to AGO1, and as a result, increased the amount of mature RISCs that could cleave target RNAs. CYP40 was not contained in mature RISCs, indicating that the association is transient. Addition of PP5 or cyclophilin-binding drug cyclosporine A prevented the association of endogenous CYP40 with HSP90-AGO1 complex and inhibited RISC assembly. These results suggest that a complex of AGO1, HSP90, CYP40, and a small RNA duplex is a key intermediate of RISC assembly in plants. The EMBO Journal (2012) 31, 267-278. doi:10.1038/emboj.2011.395; Published online 1 November 2011
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