Clarifying mammalian RISC assembly in vitro.
Publication/Presentation Date
4-29-2011
Abstract
BACKGROUND: Argonaute, the core component of the RNA induced silencing complex (RISC), binds to mature miRNAs and regulates gene expression at transcriptional or post-transcriptional level. We recently reported that Argonaute 2 (Ago2) also assembles into complexes with miRNA precursors (pre-miRNAs). These Ago2:pre-miRNA complexes are catalytically active in vitro and constitute non-canonical RISCs.
RESULTS: The use of pre-miRNAs as guides by Ago2 bypasses Dicer activity and complicates in vitro RISC reconstitution. In this work, we characterized Ago2:pre-miRNA complexes and identified RNAs that are targeted by miRNAs but not their corresponding pre-miRNAs. Using these target RNAs we were able to recapitulate in vitro pre-miRNA processing and canonical RISC loading, and define the minimal factors required for these processes.
CONCLUSIONS: Our results indicate that Ago2 and Dicer are sufficient for processing and loading of miRNAs into RISC. Furthermore, our studies suggest that Ago2 binds primarily to the 5'- and alternatively, to the 3'-end of select pre-miRNAs.
Volume
12
First Page
19
Last Page
19
ISSN
1471-2199
Published In/Presented At
Tan, G. S., Garchow, B. G., Liu, X., Metzler, D., & Kiriakidou, M. (2011). Clarifying mammalian RISC assembly in vitro. BMC molecular biology, 12, 19. https://doi.org/10.1186/1471-2199-12-19
Disciplines
Business Administration, Management, and Operations | Health and Medical Administration | Management Sciences and Quantitative Methods
PubMedID
21529364
Department(s)
Administration and Leadership
Document Type
Article