High-Resolution In Vivo Identification of MiRNA Targets by Halo-Enhanced Ago2 Pull-Down

Overview

Journal Mol Cell

Publisher Cell Press

Specialty Cell Biology

Date 2020 Jun 5

PMID 32497496

Citations 24

Authors

Xiaoyi Li

Yuri Pritykin

Carla P Concepcion

Yuheng Lu

Gaspare La Rocca

Minsi Zhang

Bryan King

Peter J Cook

Yu Wah Au

Olesja Popow

Joao A Paulo

Hannah G Otis

Chiara Mastroleo

Paul Ogrodowski

Ryan Schreiner

Kevin M Haigis

Doron Betel

Christina S Leslie

Andrea Ventura

Affiliations

Soon will be listed here.

Abstract

The identification of microRNA (miRNA) targets by Ago2 crosslinking-immunoprecipitation (CLIP) methods has provided major insights into the biology of this important class of non-coding RNAs. However, these methods are technically challenging and not easily applicable to an in vivo setting. To overcome these limitations and facilitate the investigation of miRNA functions in vivo, we have developed a method based on a genetically engineered mouse harboring a conditional Halo-Ago2 allele expressed from the endogenous Ago2 locus. By using a resin conjugated to the HaloTag ligand, Ago2-miRNA-mRNA complexes can be purified from cells and tissues expressing the endogenous Halo-Ago2 allele. We demonstrate the reproducibility and sensitivity of this method in mouse embryonic stem cells, developing embryos, adult tissues, and autochthonous mouse models of human brain and lung cancers. This method and the datasets we have generated will facilitate the characterization of miRNA-mRNA networks in vivo under physiological and pathological conditions.

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