Highly complementary target RNAs promote release of guide RNAs from human Argonaute2

Argonaute proteins use small RNAs to guide the silencing of complementary target RNAs in many eukaryotes. Although small RNA biogenesis pathways are well studied, mechanisms for removal of guide RNAs from Argonaute are poorly understood. Here we show that the Argonaute2 (Ago2) guide RNA complex is extremely stable, with a half-life on the order of days. However, highly complementary target RNAs destabilize the complex and significantly accelerate release of the guide RNA from Ago2. This "unloading" activity can 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 leads to more potent silencing of abundant mRNAs in mammalian cells. These findings help to explain why the 3' ends of mammalian microRNAs (miRNAs) rarely match their targets, suggest a mechanism for sequence-specific small RNA turnover, and offer insights for controlling small RNAs in mammalian cells.