2013 Fiscal Year Final Research Report

Application of sensitive EMT/MET reporter to identification of target genes for breast cancer stem cells.

Research Project

Project/Area Number	24650620
Research Category	Grant-in-Aid for Challenging Exploratory Research
Allocation Type	Single-year Grants
Research Field	Tumor biology
Research Institution	Kyoto University
Principal Investigator	YOSHIKAWA Kiyotsugu 京都大学, 医学(系)研究科(研究院), 准教授 (40333562)
Project Period (FY)	2012-04-01 – 2014-03-31
Keywords	EMT / 乳癌幹細胞 / MET / レポーター / 発現クローニング / トリプルネガティブ乳癌
Research Abstract	In order to identify cDNA/shRNA that can induce EMT or MET, we established a reporter system where vimentin or E-cadherin promoter drives blasticidine-resistant gene and GFP in HMLE epithelial or MDA-MB-231 mesenchymal breast cancer cell line. The cells were infected with cDNA or shRNA library and subjected to blasticidine selection. In EMT screening, Snail and Twist2 were identified as EMT inducer, and two genes, members of mitochondrial complex I, and a hub gene in signal transduction were identified to support soft aga-colonogenecity of mesenchymal cells. In MET screen, a potent MET-inducing shRNA (called shP1) was identified. The shP1 was tested for soft agar colony formation using monoclonal dual reporter cell clones from SUM159 mesenchymal breast cancer cell line. Interestingly, there are two sets of reporter clones with different response to shP1. Colony formation did not decreased in one set of reporter clones, while in the other clone shP1 reduced colony formation.