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J Biol Chem. 2015 Mar 27;290(13):8331-47. doi: 10.1074/jbc.M114.615088. Epub 2015 Feb 5.

Roles of mRNA fate modulators Dhh1 and Pat1 in TNRC6-dependent gene silencing recapitulated in yeast.

Author information

1
From the Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai 980-8578, Japan.
2
the Institute of Molecular and Cellular Biosciences and the Department of Medical Genome Sciences, University of Tokyo, Bunkyo-ku, Tokyo 113-0032, Japan, and mishima@iam.u-tokyo.ac.jp.
3
the Graduate School of Science, Kobe University, Kobe 657-8501, Japan.
4
From the Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai 980-8578, Japan, tinada@m.tohoku.ac.jp.

Abstract

The CCR4-NOT complex, the major deadenylase in eukaryotes, plays crucial roles in gene expression at the levels of transcription, mRNA decay, and protein degradation. GW182/TNRC6 proteins, which are core components of the microRNA-induced silencing complex in animals, stimulate deadenylation and repress translation via recruitment of the CCR4-NOT complex. Here we report a heterologous experimental system that recapitulates the recruitment of CCR4-NOT complex by TNRC6 in S. cerevisiae. Using this system, we characterize conserved functions of the CCR4-NOT complex. The complex stimulates degradation of mRNA from the 5' end by Xrn1, in a manner independent of both translation and deadenylation. This degradation pathway is probably conserved in miRNA-mediated gene silencing in zebrafish. Furthermore, the mRNA fate modulators Dhh1 and Pat1 redundantly stimulate mRNA decay, but both factors are required for poly(A) tail-independent translation repression by tethered TNRC6A. Our tethering-based reconstitution system reveals that the conserved architecture of Not1/CNOT1 provides a binding surface for TNRC6, thereby connecting microRNA-induced silencing complex to the decapping machinery as well as the translation apparatus.

KEYWORDS:

Gene Silencing; Translation Regulation; Yeast; Zebrafish; mRNA Decay; miRNA Mechanism

PMID:
25657010
PMCID:
PMC4375487
DOI:
10.1074/jbc.M114.615088
[Indexed for MEDLINE]
Free PMC Article

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