title: Analysis of ligand-specific co-repressor binding to the androgen receptor.
abstract: The recruitment of co-repressors to the androgen receptor is an important mechanism for reducing androgen-mediated gene activation. Importantly, co-repressors play a major role in the treatment of hormone-dependent growing tissue, such as prostate cancer and breast cancer. In line with this, co-repressor dysfunction seems to be a major player for development of castration-resistant prostate cancer or therapy-resistant breast cancer. The molecular basis of hormone therapy by particular antihormones (antagonists) for the androgen receptor (AR) is mediated by enhanced recruitment and activity of co-repressors that cause repression of AR target genes that regulate proliferation and alteration of cancer cells. Therefore co-repressor recruitment is a crucial molecular mechanism of gene repression as well as inhibition of cancer growth. Here we describe different strategies to investigate co-repressor recruitment to the AR. First, we developed a modified mammalian two-hybrid system to investigate the recruitment of co-repressors to the AR within mammalian cells. This assay is very useful for the identification of the molecular mechanism of new AR antagonists and for molecular analysis of castration-resistant prostate cancer expressing the AR. Second, we describe a technique to analyze the interaction of AR isolated from human prostate cancer cells with a newly generated AR-specific co-repressor peptide, which is bacterially expressed and affinity purified by glutathione-S-transferase affinity precipitation assays in vitro. In summary, these methods can greatly facilitate the study of AR-co-repressor interactions.
authors.affiliation: Institute of Human Genetics, Jena University Hospital, Jena, Germany, claudia.gerlach@mti.uni-jena.de
authors.forename: Claudia
authors.initials: C
authors.lastname: Gerlach
authors.forename: Daniela
authors.initials: D
authors.lastname: Roell
authors.forename: Aria
authors.initials: A
authors.lastname: Baniahmad
mesh_terms: D018929:Cell Culture Techniques; D056970:Co-Repressor Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D047468:Immunoprecipitation; D008024:Ligands; D011944:Receptors, Androgen / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020798:Two-Hybrid System Techniques
chemical_list: D056970:Co-Repressor Proteins; D008024:Ligands; D011944:Receptors, Androgen; D011993:Recombinant Fusion Proteins