title: Use of genome-wide RNAi screens to identify regulators of embryonic stem cell pluripotency and self-renewal.
abstract: Embryonic stem cells (ESCs) are characterized by two defining features: pluripotency and self-renewal. They hold tremendous promise for both basic research and regenerative medicine. To fully realize their potentials, it is important to understand the molecular mechanisms regulating ESC pluripotency and self-renewal. The development of RNA interference (RNAi) technology has revolutionized functional genetic studies in mammalian cells. In recent years, genome-wide RNAi screens have been adopted to systematically study ESC biology, and have uncovered many previously unknown regulators, including transcription factors, chromatin remodelers, and posttranscriptional modulators. Here, we describe a method for the identification of regulators of ESC pluripotency and self-renewal using RNAi screens, as well as assays for further validation and characterization of the identified candidates. With modifications, this method can also be adapted to study the fate specification events during ESC differentiation.
authors.affiliation: Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, Research Triangle Park, NC, 27709, USA.
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Zheng
authors.forename: Guang
authors.initials: G
authors.lastname: Hu
mesh_terms: D018929:Cell Culture Techniques; D002454:Cell Differentiation; D019070:Cell Lineage; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism*; D005434:Flow Cytometry; D023281:Genomics / Q000379:methods*; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D013194:Staining and Labeling; D014162:Transfection
chemical_list: D034741:RNA, Small Interfering
references.citation: Nature. 2010 Nov 11;468(7321):316-20
references.pmid: 20953172
references.citation: Stem Cells. 2012 May;30(5):910-22
references.pmid: 22367759
references.citation: Nature. 2010 Sep 23;467(7314):430-5
references.pmid: 20720539
references.citation: Cell Stem Cell. 2009 May 8;4(5):403-15
references.pmid: 19345177
references.citation: J Vis Exp. 2012 May 30;(63):
references.pmid: 22688536
references.citation: Cell. 2008 Jul 11;134(1):162-74
references.pmid: 18614019
references.citation: Nat Rev Mol Cell Biol. 2011 Sep 23;12(10):680-6
references.pmid: 21941277
references.citation: Nature. 2006 Aug 3;442(7102):533-8
references.pmid: 16767105
references.citation: Genes Dev. 2009 Apr 1;23(7):837-48
references.pmid: 19339689
references.citation: Annu Rev Cell Dev Biol. 2001;17:435-62
references.pmid: 11687496
references.citation: Cell. 2008 Feb 22;132(4):661-80
references.pmid: 18295582
references.citation: Cell. 2011 Mar 18;144(6):940-54
references.pmid: 21414485