Identification of five human novel genes associated with cell proliferation by cell-based screening from an expressed cDNA ORF library
Identification of five human novel genes associated with cell proliferation by cell-based screening from an expressed cDNA ORF library
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DOI：
10.1016/j.lfs.2007.08.006
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发表时间：
2007-09-15
期刊：
影响因子：
6.1
通讯作者：
Ma, Dalong
中科院分区：
文献类型：
--
作者：
Ma, Xi;Wang, Xinyu;Ma, Dalong
功能谱分析技术的发展为高通量功能基因组学研究提供了契机。我们描述了一个基于细胞的筛选系统，以确定新的人类基因与细胞增殖。该方法集成了荧光素酶报告基因活性、荧光染色、自动显微镜和细胞表型测定。利用该系统对本实验室克隆的409个人类新基因进行了筛选，发现其中27个基因的启动子-海肾荧光素酶报告质粒（pRL）活性显著上调。通过形态学观察、钙黄绿素AM荧光染色、MTT法和细胞周期分析，确定TRAF 3 IP 3、ZNF 306、ZNF 250、SGOL 1和ZNF 434 5个基因与细胞增殖相关。此外，我们发现，TRAF 3 IP 3基因，最初被确定为与TRAF 3特异性相互作用，通过调节c-Jun N-末端激酶（INK）途径刺激细胞生长，并且TRAF 3 IP 3的RNAi证实了这种作用是生理性的和必要的。综上所述，我们建立了一个快速、高效的生长调控基因筛选系统。使用新的筛选系统，我们首次确定了与细胞增殖相关的五个基因。(C)2007爱思唯尔公司All rights reserved.
The development of functional profiling technologies provides opportunity for high-throughput functional genomics studies. We describe a cell-based screening system to identify novel human genes associated with cell proliferation. The method integrates luciferase reporter gene activity, fluorescence stain, automated microscopy and cellular phenotype assays. We successfully used the system to screen 409 novel human genes cloned by our lab and found that 27 genes significantly up-regulated promoter-Renilla luciferase reporter plasmid (pRL) activity. Among them, five genes, TRAF3IP3, ZNF306, ZNF250, SGOL1, and ZNF434, were determined through morphological observation, calcein AM fluorescence stain, MTT assay and cell cycle analysis to be associated with cell proliferation. Furthermore, we showed that the gene TRAF3IP3, which initially was identified to specifically interact with TRAF3, stimulated cell growth by modulating the c-Jun N-terminal kinase (INK) pathway, and RNAi of TRAF3IP3 confirmed that the effect was physiological and necessary. In summary, we integrated a rapid and efficient system for screening novel growth regulatory genes. Using the new screening system we identified five genes associated with cell proliferation for the first time. (C) 2007 Elsevier Inc. All rights reserved.