Kawate Hisaya, Wu Yin, Ohnaka Keizo, Tao Rong-Hua, Nakamura Kei-Ichiro, Okabe Taijiro, Yanase Toshihiko, Nawata Hajime, Takayanagi Ryoichi
Department of Geriatric Medicine, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
J Clin Endocrinol Metab. 2005 Nov;90(11):6162-9. doi: 10.1210/jc.2005-0179. Epub 2005 Aug 23.
Recent imaging studies revealed that androgen receptor (AR) is ligand-dependently translocated from the cytoplasm into the nucleus and forms intranuclear fine foci. In this study, we examined whether intracellular dynamics of mutant ARs detected in two androgen insensitivity syndrome (AIS) patients was impaired.
ARs with mutations in the DNA-binding domain were functionally characterized and compared with the wild-type AR.
In a complete AIS patient (subject 1), cysteine residue 579 in the first zinc finger motif of AR was substituted for phenylalanine (AR-C579F). Another mutation (AR-F582Y) was found in a partial AIS patient (subject 2).
AR-F582Y retained less than 10% of the transactivation activity of the wild-type AR, whereas no ligand-dependent transactivation was detected for AR-C579F. Image analyses of the receptors fused to green fluorescent protein showed that the wild-type AR was ligand-dependently translocated into the nucleus in which it formed fine subnuclear foci. Surprisingly, after the addition of dihydrotestosterone, the two mutant ARs initially formed large cytoplasmic dots, many of which were found to be close to mitochondria by electron microscopy. Subsequently, a part of the ligand-bound mutant ARs gradually entered the nucleus to form a smaller number of larger dots, compared with the wild-type AR. Fluorescence recovery after photobleaching analysis revealed that the intranuclear mobility of the mutant ARs decreased, compared with that of the wild-type AR.
These results suggest that the abnormal translocation, localization, and mobility of the mutant ARs may be the cause of AIS in these subjects.
最近的影像学研究表明，雄激素受体（AR）在配体的作用下从细胞质转移至细胞核，并在核内形成精细的核内病灶。在本研究中，我们检测了在两名雄激素不敏感综合征（AIS）患者中检测到的突变型AR的细胞内动力学是否受损。
对DNA结合域发生突变的AR进行功能表征，并与野生型AR进行比较。
在一名完全性AIS患者（受试者1）中，AR第一个锌指基序中的半胱氨酸残基579被苯丙氨酸取代（AR-C579F）。在一名部分性AIS患者（受试者2）中发现了另一种突变（AR-F582Y）。
AR-F582Y保留的野生型AR反式激活活性不到10%，而未检测到AR-C579F的配体依赖性反式激活。对与绿色荧光蛋白融合的受体进行图像分析表明，野生型AR在配体的作用下转移至细胞核，在核内形成精细的核内病灶。令人惊讶的是，添加二氢睾酮后，两种突变型AR最初形成大的细胞质点，通过电子显微镜发现其中许多靠近线粒体。随后，与野生型AR相比，一部分与配体结合的突变型AR逐渐进入细胞核，形成数量较少的较大的点。光漂白后荧光恢复分析表明，与野生型AR相比，突变型AR在核内的移动性降低。
这些结果表明，突变型AR的异常转运、定位和移动性可能是这些受试者发生AIS的原因。
