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Mammalian miRNA RISC recruits CAF1 and PABP to affect PABP-dependent deadenylation

Marc R Fabian1, Géraldine Mathonnet, Thomas Sundermeier

  • 1Department of Biochemistry, McGill University, Montreal, QC H3G 1Y6, Canada.

Molecular Cell
|September 1, 2009
PubMed

Insights

MicroRNAs (miRNAs) repress gene expression through a two-step process. First, translation is inhibited, followed by mRNA deadenylation mediated by the GW182 protein interacting with PABP.

Area of Science:

  • Molecular Biology
  • Gene Regulation
  • Biochemistry

Background:

  • MicroRNAs (miRNAs) are key regulators of gene expression.
  • miRNAs typically function by binding to the 3' untranslated region (3'UTR) of target mRNAs.
  • This binding leads to translational inhibition and/or mRNA deadenylation and destabilization.

Purpose of the Study:

  • To investigate the mechanism and kinetics of miRNA-mediated mRNA deadenylation.
  • To elucidate the role of specific proteins in the miRNA-mediated deadenylation pathway.
  • To understand the sequential steps involved in miRNA-mediated gene silencing.

Main Methods:

  • Experiments were conducted using mouse Krebs-2 ascites extract.
  • Analysis of miRNA-mediated deadenylation kinetics.
  • Co-immunoprecipitation assays to study protein interactions within the RNA-induced silencing complex (miRISC).

Main Results:

  • miRNA-mediated mRNA deadenylation occurs after initial translational inhibition, confirming a two-step repression mechanism.
  • A let-7 miRNA-loaded miRISC interacts with poly(A)-binding protein (PABP) and the deadenylases CAF1 and CCR4.
  • miRNA-mediated deadenylation requires CAF1 activity and PABP, identifying PABP as a crucial miRNA coactivator.
  • The GW182 protein directly interacts with PABP, and this interaction is essential for miRNA-mediated deadenylation.

Conclusions:

  • miRNA-mediated gene silencing involves a coordinated two-step process: translational inhibition followed by deadenylation.
  • The GW182 protein plays a critical role in recruiting PABP to the miRISC, facilitating deadenylation.
  • PABP acts as a coactivator in miRNA-mediated deadenylation, highlighting a novel aspect of gene regulation by miRNAs.

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