High-Resolution In Vivo Identification of miRNA Targets by Halo-Enhanced Ago2 Pull-Down. Academic Article uri icon

Overview

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.

publication date

  • June 3, 2020

Research

keywords

  • Argonaute Proteins
  • Embryonic Stem Cells
  • Glioma
  • MicroRNAs
  • RNA, Messenger
  • Recombinant Fusion Proteins

Identity

PubMed Central ID

  • PMC7446397

Scopus Document Identifier

  • 85087530351

Digital Object Identifier (DOI)

  • 10.1016/j.molcel.2020.05.009

PubMed ID

  • 32497496

Additional Document Info

volume

  • 79

issue

  • 1