High-throughput functional microRNAs profiling by recombinant AAV-based microRNA sensor arrays

microRNAs (miRNAs) are small and non-coding RNAs which play critical roles in physiological and pathological processes. A number of methods have been established to detect and quantify miRNA expression. However, method for high-throughput miRNA function detection is still lacking. We describe an ade...

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Bibliographic Details
Published in:	PloS one 2012-01, Vol.7 (1), p.e29551
Main Authors:	Tian, Wenhong, Dong, Xiaoyan, Liu, Xuerong, Wang, Gang, Dong, Zheyue, Shen, Wei, Zheng, Gang, Lu, Jianxin, Chen, Jinzhong, Wang, Yue, Wu, Zhijian, Wu, Xiaobing
Format:	Article
Language:	eng
Subjects:	12-O-Tetradecanoylphorbol-13-acetate Acetic acid Analysis Animals Apoptosis Biological activity Biology Biosensing Techniques - instrumentation Biotechnology Bone marrow Breast cancer Cell Differentiation - drug effects Cell Differentiation - genetics Cell Line Cell lines Colorectal cancer Dependovirus - drug effects Dependovirus - genetics Disease control Disease prevention DNA polymerases Efficiency Fireflies Gaussia Gene expression Gene Expression Profiling - methods Gene Expression Regulation - drug effects Genetic engineering Genetic vectors Herpes viruses High-Throughput Screening Assays - methods Humans Laboratories Life sciences Liver cancer Luciferase Medicine Methods Mice Microarray Analysis - instrumentation MicroRNA MicroRNAs MicroRNAs - genetics MicroRNAs - metabolism miRNA Physiological aspects Plasmids Polymerase chain reaction Profiling Prostate cancer Recombination, Genetic - drug effects Recombination, Genetic - genetics Ribonucleic acid RNA RNA-directed DNA polymerase Science Policy Sensor arrays Sensors Tetradecanoylphorbol Acetate - pharmacology Virology Viruses
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Summary:	microRNAs (miRNAs) are small and non-coding RNAs which play critical roles in physiological and pathological processes. A number of methods have been established to detect and quantify miRNA expression. However, method for high-throughput miRNA function detection is still lacking. We describe an adeno-associated virus (AAV) vector-based microRNA (miRNA) sensor (Asensor) array for high-throughput functional miRNA profiling. Each Asensor contains a Gaussia luciferase (Gluc) and a firefly luciferase (Fluc) expression cassette to sense functional miRNA and to serve as an internal control respectively. Using this array, we acquired functional profiles of 115 miRNAs for 12 cell lines and found "functional miRNA signatures" for several specific cell lines. The activities of specific miRNAs including the let-7 family, miR-17-92 cluster, miR-221, and miR-222 in HEK 293 cells were compared with their expression levels determined by quantitative reverse transcriptase polymerase chain reaction (QRT-PCR). We also demonstrate two other practical applications of the array, including a comparison of the miRNA activity between HEK293 and HEK293T cells and the ability to monitor miRNA activity changes in K562 cells treated with 12-O-tetradecanoylphorbol-13-acetate (TPA). Our approach has potential applications in the identification of cell types, the characterization of biological and pathological processes, and the evaluation of responses to interventions.
ISSN:	1932-6203 1932-6203