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Proc Natl Acad Sci U S A. 2017 Apr 11;114(15):3927-3932. doi: 10.1073/pnas.1620019114. Epub 2017 Mar 29.

Identification of targets of tumor suppressor microRNA-34a using a reporter library system.

Author information

1
Department of Systems BioMedicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University (TMDU), Bunkyo, Tokyo 113-8519, Japan.
2
Department of Systems BioMedicine, National Research Institute for Child Health and Development, Setagaya, Tokyo 157-8535, Japan.
3
Imperial College London, South Kensington, London SW7 2AZ, United Kingdom.
4
Department of Breast Surgery, Nagoya City University Graduate School of Medical Sciences, Nagoya, Aichi 467-8601, Japan.
5
Division of Biology, California Institute of Technology, Pasadena, CA 91125.
6
Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA 91010.
7
Department of Systems BioMedicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University (TMDU), Bunkyo, Tokyo 113-8519, Japan; asahara.syst@tmd.ac.jp.
8
Core Research for Evolutional Science and Technology (CREST), Japan Agency for Medical Research and Development (AMED), Tokyo 100-0004, Japan.
9
Department of Experimental Medicine, The Scripps Research Institute, San Diego, CA 92131.

Abstract

miRNAs play critical roles in various biological processes by targeting specific mRNAs. Current approaches to identifying miRNA targets are insufficient for elucidation of a miRNA regulatory network. Here, we created a cell-based screening system using a luciferase reporter library composed of 4,891 full-length cDNAs, each of which was integrated into the 3' UTR of a luciferase gene. Using this reporter library system, we conducted a screening for targets of miR-34a, a tumor-suppressor miRNA. We identified both previously characterized and previously uncharacterized targets. miR-34a overexpression in MDA-MB-231 breast cancer cells repressed the expression of these previously unrecognized targets. Among these targets, GFRA3 is crucial for MDA-MB-231 cell growth, and its expression correlated with the overall survival of patients with breast cancer. Furthermore, GFRA3 was found to be directly regulated by miR-34a via its coding region. These data show that this system is useful for elucidating miRNA functions and networks.

KEYWORDS:

breast cancer; miR-34a; microRNA target screening; reporter library system

PMID:
28356515
PMCID:
PMC5393199
DOI:
10.1073/pnas.1620019114
[Indexed for MEDLINE]
Free PMC Article

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