雄激素受体与核受体辅助抑制因子的关系

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雄激素受体与核受体辅助抑制因子的关系

廖国庆3,汤恢焕,吕新生

(中南大学湘雅医院外科,长沙410008)

[摘要]　目的:探讨雄激素受体(AR )与核受体辅助抑制因子(SMRT )是否能相互作用及其作用部位。方法:重

组构建AR ,SMRT 基因或其基因片段的质粒,体外转录,合成35S 标记融合蛋白,采用转染试验及哺乳动物细胞双杂交实验(transient transfection ,mammalian two 2hybrid test ),GST 沉淀试验(GST pull 2down assay ),间接免疫荧光观察

(indirect immunofluorescence staining )的方法,观察AR 与SMRT 的关系。结果:AR 具有内在的转录抑制活性,可与SMRT 直接作用,其作用部位:在AR 分子上位于配体结合区(LBD ),而DNA 结合区能增强这种作用;在SMRT 分子

上则位于羧基端核受体作用区(IDs ),在这个区域的L XXXIXXXI/L 功能基团突变后将会影响AR 与SMRT 的结合。结论:AR 通过其LBD 区与SMRT 分子中的ID2相互作用。

[关键词]　受体;　雄激素;　SMRT ;　相互作用

[中图分类号]　Q781 [文献标识码]　A [文章编号]　167227347(2004)022*******

Androgen receptor and SMRT

L IAO Guo 2qing 3,TAN G Hui 2huan ,L U ・・

Xin 2sheng

(Depart ment of S urgery ,Xiangya Hospital ,Cent ral South U niversity ,Changsha 410008,Chi na )

Abstract :　Objective To explore the interaction between androgen receptor (AR )and silencing

mediator for retinoid and thyroid hormone receptor (SMR T )and their interaction site.Methods We recombined and constructed AR ,SMR T gene and gene fragments ,in vitro translated 35S fusion proteins to investigate the relationship between AR and SMR T using transient transfection ,mammalian two 2hy 2brid test ,GST pull 2down assay ,and indirect immunofluorescence staining.R esults AR possessed an intrinsic transcriptional repression activity and AR interacted directly with SMR T.One interactive sur 2face on AR was mapped to the ligand 2binding domain (LBD ),and the presence of DNA binding domain enhanced this interaction.The binding surface on SMR T was mapped to the carboxyl 2terminal nuclear receptor interacting domain (ID ),and mutation of the L XXXIXXXI/L corepressor motif within this do 2main interferred with the interaction.Conclusion LBD domain on the AR can interact with ID2motif on the SMR T.

K ey w ords :　receptor ;　androgen ;　SMR T ;　interaction

[J Cent South Univ (Med Sci ),2004,29(2):0157206]

AR (androgen receptor )是核受体的成员之一,它不但与男性成熟有关,而且在前列腺癌进展中有重要作用[1]。SMR T (silencing mediator for retinoid and thyroid hormone receptor )是核受体辅助抑制因子,目前大量研究结果表明,SMR T 对许多核受体的活性,如ER ,PR ,等具有抑制作用,对平衡体内激素的消涨具有重要作用[2,3]。但AR 与SMR T 的关系如何,目前研究不多。本研究旨在探讨AR 能否与SMR T 作用及其作用部位。1　材料与方法

1.1　质粒和试剂 AR cDNA ,G al4D BD 与AR 的融

合基因片段(G 42AR A/B ,G 42AR DE ,G 42AR )分别由Dr.Chang (R ochester 大学医学中心,R ochester ,NY )和Dr.Lirim Shemshedini (T oled o 大学,T oled o ,Ohio )惠赠。G 4s 2AR ,G 42AR 是由本实验室构建,用全长人类AR cDNA 插入到pCMX 2G al Xho1和Asp718位点之间,PCR 扩增,由此构建1～500,1～660,501～660,501～919,和661～919AR 各片段,并插入到pCMX 2HA 载体

中,构建重组新的AR 及AR 片段质粒。人类SMR T e [4]

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51①收稿日期:2003211220 作者简介:廖国庆(19622),男,湖南衡东人,博士,副教授,主要从事肿瘤外科的基础和临床研究。3通讯作

者,E 2mail :liaoguoqing @

中南大学学报(医学版)

J Cent South Univ (Med Sci )

　2004,29(2)