相扑蛋白
雄激素受体
转染
赖氨酸
免疫沉淀
化学
细胞生物学
受体
恩扎鲁胺
转录因子
兴奋剂
泛素
生物
分子生物学
生物化学
基因
前列腺癌
遗传学
氨基酸
癌症
作者
Miia M. Rytinki,Sanna Kaikkonen,Päivi Sutinen,Jorma J. Palvimo
出处
期刊:Methods in molecular biology
日期:2011-01-01
卷期号:: 183-197
被引量:20
标识
DOI:10.1007/978-1-61779-243-4_12
摘要
Androgen receptor (AR) is a ligand-controlled transcription factor that is deregulated and therefore targeted in prostate cancer. In addition to androgens, AR is regulated by post-translational modifications (PTMs). SUMOylation, conjugation of small ubiquitin-related modifier (SUMO) protein 1, 2, or 3, is a bulky PTM regulating several important physiological processes. We have shown that AR is modified by SUMO-1 at two conserved lysine residues in its N-terminal domain. This agonist-enhanced modification represses the transcriptional activity of the receptor in a reversible and target gene-selective fashion. Acceptor sites for SUMOs are also found in several other nuclear receptors. Since the cellular steady-state level of SUMO modifications of most substrates, including AR, is very low, transfection- and SUMO overexpression-based protocols are often needed to render the modifications clearly detectable. This chapter describes protocols for analyzing AR SUMOylation in cultured cells by immunoblotting, gel mobility shift assays, and immunoprecipitation. These methodologies are generally applicable for determining whether a particular protein is SUMOylated and for identifying the lysine residue(s) modified.
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