Multi-Pole Approach to Structural Science

May 10 - 13, 2015

Structural insights to the role of the CCR4-NOT complex and GW182 and DDX6 proteins in miRNA-mediated repression
Witold Filipowicz

Structural insights to the role of the CCR4-NOT complex and GW182 and DDX6 proteins in microRNA-mediated repression Witold Filipowicz Friedrich Miescher Institute for Biomedical Research, 4002 Basel, Switzerland MicroRNAs (miRNAs) are ~20-nt-long regulatory RNAs expressed in eukaryotes. They regulate gene expression post-transcriptionally, by imperfectly base-pairing to 3’UTRs of mRNAs what results in translational repression and mRNA deadenylation and degradation. Most of mammalian genes are predicted to be subject to miRNA regulation. Clearly, discovery of miRNAs added a new dimension to the complexity and regulation of eukaryotic genomes. We will discuss current knowledge about the mechanism of miRNA-mediated repression, focusing on a role of GW182 proteins and the multi-subunit CCR4-NOT complex in translational repression and mRNA deadenylation. Our recent work performed in collaboration with Elena Conti’s group at MPI in Martinsried and Marcin Nowotny and Andrzej Dziembowski in Warsaw, revealed the mechanism of the CCR4/NOT recruitment by GW182 proteins and a role of Trp-containing motifs (W-motifs) of GW182s in this process. Structural studies showed that the C-terminal repressive domain of GW182 associates with CCR4/NOT by directly interacting with the complex subunit CNOT9, which contains two Trp-binding pockets. We have also demonstrated that, further downstream, the CCR4-NOT complex recruits the DEAD-box RNA helicase/ATPase DDX6 to mRNA. Structural analyses of the apo form of DDX6 and its complex with the CCR4/NOT scaffold subunit CNOT1 revealed that the DDX6 interaction with CCR4/NOT is accompanied by a major conformational change in DDX6 resulting in activation of its ATPase activity. The DDX6 activation is required for its function in miRNA-mediated repression of translation.