Presentation/Session Information

Session Information

Session Title: RNA Interference, Noncoding RNAs, and Genetic Technologies Session Type: Parallel
Session Location: Carnesale Palisades Ballroom Session Time: Sat, Jun 27 8:30AM - 11:30AM

Presentation Information

Program Number: 139 Presentation Time: 8:42AM - 8:54AM

Presentation Content

Toward an understanding of cooperative miRNA-mediated silencing.M.N. Flamand 1, H.H. Gan 2, E. Wu 1, A. Vashisht 3, G. Jannot 4, J. Wohlschlegel 3, M.J. Simard 4, T.F. Duchaine 1. 1)Goodman Cancer Research Centre/Biochemistry, McGill University, Montreal, Quebec; 2)Department of Chemistry, New York University, New York, NY; 3)Department of Biological Chemistry David Geffen School of Medicine at UCLA, Los Angeles, CA; 4)Laval University Cancer Research Centre, Hôtel-Dieu de Québec (Centre Hospitalier Universitaire de Québec), Quebec City, QC

MicroRNAs (miRNAs), small RNA molecules derived from gene-encoded hairpins, play roles in various biological phenomena ranging from development to environmental responses. From within the miRNA-Induced Silencing Complex (miRISC), miRNAs base-pair with 3’ un-translated regions (3’UTRs) of mRNAs, and direct a series of gene silencing mechanisms. At the core of the miRISC are specialized Argonautes; ALG-1 and ALG-2 bind to miRNAs in C. elegans. While several lines of evidence point to the importance of cooperating interactions between multiple miRNA-binding sites on individual target mRNAs, they are still largely investigated as functionally independent regulatory units. We have previously shown that mRNA deadenylation, a prominent mechanism of miRNA-mediated silencing, strictly requires target site cooperation.

Investigation of the mechanistic basis for cooperative silencing by miRNAs in C. elegans unveiled its multi-faceted nature. Firstly, miRNA-binding sites cooperate in miRISC recruitment; binding to a canonical site enables the subsequent recruitment of miRISC to weaker, and even non-canonical sites on mRNAs, a process we coined trans-seeding. Secondly, we find that interactions between miRISC units are required to trigger miRNA-mediated silencing. At least part of such interactions may occur through homo- or hetero-dimerization of the ALG-1/2 Argonautes. Structural and thermodynamic modeling of ALG-1 proteins associated to cooperating miRNA-binding sites identified residues that may be required for cooperation. We are systematically validating these candidates in vivo and using embryonic cell-free assays by exploiting CRISPR/Cas-9 mediated genome-editing strategies. Furthermore, we found that two interactors of cooperative miRISC, the poly(A) binding proteins PAB-1 and PAB-2, are required for the full functions of miRNAs in vivo. Surprisingly, while they are dispensable for miRNA-mediated deadenylation, they are required for all miRNA-mediated translation repression.

Insight in the mechanistic nature of cooperative miRNA-mediated silencing will enable more reliable prediction of the identity of targets, the extent of their silencing, and their biological impact.

Please note: Abstract shown here should NOT be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

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