title: In Vitro Transcription to Study WT1 Function.
abstract: In vitro transcription methods using mammalian nuclear extracts have been available for over 30 years and have allowed sophisticated biochemical analyses of the transcription process. This method has been extensively used to study the basic mechanisms of transcription, allowing the identification of the general transcription factors and elucidation of their mechanisms of action. Gene-specific transcriptional regulators have also been studied using in vitro transcription. This has facilitated the identification of their cofactors and provided information on their function that is invaluable to facilitate their study in a more physiological setting. Here we describe the application of in vitro transcription methods to study the mechanism of action of WT1. Coupling transcription assays with methods to purify transcription complexes, and protein affinity chromatography, has provided insights into how WT1 can both positively and negatively regulate transcription.
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Office G50, Biomedical Sciences Building, University Walk, Clifton, Bristol, BS8 1TD, UK. Stefan.Roberts@bristol.ac.uk.
authors.forename: Stefan G E
authors.initials: SG
authors.lastname: Roberts
mesh_terms: D002467:Cell Nucleus / Q000235:genetics; D002846:Chromatography, Affinity; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D011994:Recombinant Proteins / Q000235:genetics; D014158:Transcription, Genetic; D025721:WT1 Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D006657:Histones; D011994:Recombinant Proteins; D025721:WT1 Proteins; C583751:WT1 protein, human
keywords: Affinity chromatography; BASP1; Histone; Nuclear; Promoter; Transcription; WT1