Gene Silencing Mediated by Long-Lived Argonaute Protein-RNA Complexes

The activity of the Argonaute class of proteins stands out as a cellular control mechanism that by binding RNA regulates protein synthesis through "remote-control" gene silencing.

Argonaute proteins are the catalytic components of the RNA-induced silencing complex (RISC), the protein complex responsible for RNA interference (RNAi) gene silencing. Argonaute proteins bind different classes of small noncoding RNAs, including microRNAs (miRNAs), small interfering RNAs (siRNAs), and Piwi-interacting RNAs (piRNAs). Small RNAs guide Argonaute proteins to their specific targets through sequence complementarity, which typically leads to silencing of the target. Some of the Argonaute proteins have endonuclease activity directed against messenger RNA (mRNA) strands that display extensive complementarity to their bound small RNA, and this is known as Slicer activity. These proteins are also partially responsible for selection of the guide strand and destruction of the passenger strand of the siRNA substrate.

Investigators at The Scripps Research Institute (La Jolla, CA, USA) had previously employed X-ray crystallography to determine the high-resolution atomic structure of an Argonaute 2-miRNA complex. In the current work, they focused on the mechanism that causes the Argonaute protein to separate from its RNA guide strand.

Results published in the May 9, 2013, issue of the journal Molecular Cell revealed that the Argonaute2 (Ago2)-guide RNA complex was extremely stable, with a half-life on the order of days. However, highly complementary target RNAs destabilized the complex and significantly accelerated release of the guide RNA from Ago2. This “unloading” activity could be enhanced by mismatches between the target and the guide 5′ end and attenuated by mismatches to the guide 3′ end. The introduction of 3′ mismatches led to more potent silencing of abundant mRNAs in mammalian cells.

These findings help to explain why the 3′ ends of mammalian miRNAs rarely match their targets, suggest a mechanism for sequence-specific small RNA turnover, and offer insights for controlling small RNAs in mammalian cells.

“Learning to control natural gene silencing processes will allow an entirely new approach to treating human disease,” said senior author Dr. Ian J. MacRae, assistant professor of integrative structural and computational biology at the Scripps Research Institute. “I can think of many applications for these. One of the most obvious would be against hepatitis C virus, which requires a certain miRNA in liver cells for efficient replication; an RNA-based drug that speeds up the unloading of this virus-enhancing miRNA would be a powerful approach for shutting down the virus.”

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