ANDROGEN RECEPTOR DEGRADATION

描述（由申请人提供）：人雄激素受体（AR）是核受体超家族的成员，也是用于控制前列腺癌的药物的主要靶点。这种靶向是基于AR是前列腺生长和分化的决定因素。因此，了解AR在细胞内是如何调节的是很重要的。先前的研究表明，AR在无配体状态下非常不稳定，这表明蛋白质水解对调节AR稳态水平的重要性。我们的研究主要集中在AR降解的机制，以及这个过程本身是如何被调节的。在初步研究中，我们发现AR是通过泛素/蛋白酶体途径降解的。研究进一步表明，这一过程受到一种名为Chip的蛋白质的影响，这种蛋白质与hsp70和hsp90等分子伴侣蛋白相关。我们还表明，转染Chip可以抑制同样表达AR的前列腺癌细胞的生长。该项目的长期目标是全面表征AR降解的机制，以及Chip如何导致前列腺癌细胞生长抑制。在aim 1中，我们将确定靶向AR降解的信号。

DESCRIPTION (Provided by applicant): The human androgen receptor (AR) is a member of the nuclear receptor superfamily and a major target of drugs used to control prostate cancer. This targeting is based on the AR being a determinant of prostate gland growth and differentiation. For this reason, it is important to understand how the AR is regulated inside cells. Previous studies showed that the AR is very unstable in its unliganded state, pointing to the importance of proteolysis to regulating AR steady state levels. Our studies are focused on the mechanisms underlying AR degradation, and how this process may itself be regulated. In preliminary studies, we show that the AR is degraded by the ubiquitin/proteasome pathway. This process is further shown to be affected by a protein called Chip that associates with molecular chaperones such as Hsp7O and Hsp9O. We also show that transfection of Chip inhibits growth of prostate cancer cells that also express AR. The long term goals of this project are to fully characterize the mechanisms governing AR degradation, and how Chip leads to prostate cancer cell growth inhibition In aim 1, we will determine the signals that target AR for degradation. Our hypothesis is that the signals for this event are based partly on AR conformation and partly on cis-acting sequences. We will first determine the role of protein folding in AR degradation, and then examine the role of a putative cis-acting signal, called a PEST sequence, in AR degradation. We will also test for the role of AR phosphorylation as a signal for its degradation. In aim 2, we will identify the sites of AR ubiquitinylation and the molecular components that catalyze this reaction. We will determine whether there is a functional relationship between ubiquitinylation, sumoylation, and the PEST sequence. We will also identify the E2 and E3 enzymes that conjugate ubiquitin to AR. Further studies will confirm that these enzymes are expressed in human prostate glandular epithelium. In aim 3, we will detrermine the mechanism by which Chip leads to arrest of prostate cancer cell growth. We will test whether there is a correlation between Chip effects on cell growth and decreasing AR levels, and we will compare the mechanisms of growth arrest in LNCaP cells and their androgen-independent derivative (LNCaPAI) in terms of apoptosis, cell cycle arrest or necrotic cell death.