ANDROGEN RECEPTORS IN THE TFM SYNDROME

这个建议的目标是了解生化程序， 目的：为靶器官的靶向治疗提供分子基础， 操纵这些器官的异常生长。 在过去的三年里， 我们已经分离并产生了两个非常重要的工具[全长 人/大鼠雄激素受体（AR）cDNA和单/多特异性抗体 AR]研究雄激素作用的分子机制。 我们也 在雄激素不敏感的患者中发现了几种AR突变， 睾丸女性化（Tfm）综合征。 基于这些发现，我们 我想测试四个假设： 假设1：AR缺失或缺陷可能导致Tfm综合征。 测试 基于这一假设，我们计划对AR基因编码区进行测序， 人和小鼠Tfm综合征。 这些结果可以让我们检测出 DNA和RNA水平的异常AR。 假设二：某些缺陷的AR可能阻止AR的易位， 细胞质到细胞核。 为了验证这一假设，我们计划应用 微波固定法免疫细胞学定位AR Tfm患者的成纤维细胞。 假设I和假设II的结果可能 使我们能够确定导致核反应的序列 AR的本地化过程。 假设III：Tfm小鼠的前列腺可能被重新诱导， 通过将正常AR cDNA引入尿生殖窦（UGS）而开发 的Tfm小鼠。 为了检验这一假设，将来自Tfm小鼠的UGS与来自Tfm小鼠的UGS进行比较。 用含有正常AR cDNA的逆转录病毒转染并在体内培养 检查前列腺是否被诱导。 结果可能 证明了正常AR在间质（基质）中的必要性， 前列腺发育 假设四：小鼠的Tfm综合征可以通过受精卵纠正 显微注射（转基因实验）正常AR cDNA。 为了验证这一 将正常AR cDNA克隆到表达载体中， 含有非常强的启动子（延伸因子启动子）， 显微注射到Tfm小鼠的受精卵中。 结果可以提供 Tfm综合征基因治疗的可能性。 上述研究将允许评估AR在Tfm中的作用。 综合征，并可能最终导致新的诊断 为临床治疗Tfm综合征提供了新的思路和方法。

The goals of this proposal are to understand the biochemical procedures by which at target organs and to provide a molecular basis for the clinical manipulation of abnormal growth of these organs. For the past three years, we have isolated and produced two very important tools [full-length of human/rat androgen receptor (AR) cDNAs and mono-/poly- specific antibodies to AR] for the study of molecular mechanism of androgen action. We also found several mutations of AR in patients with androgen insensitivity or testicular feminization (Tfm) syndrome. Based on these discoveries, we propose to test four hypotheses: Hypothesis I: Absent or defective AR may cause the Tfm syndrome. To test this hypothesis, we plan to sequence the coding region of AR gene from human and mice with Tfm syndrome. The results may allow us to detect the abnormal AR at DNA and RNA levels. Hypothesis II: Some defective AR may prevent the translocation of AR from cytoplasm to nuclei. To test this hypothesis, we plan to apply the immunocytology with microwave-fixed method to locate AR in cultured fibroblasts from Tfm patients. The results of hypotheses I and II may allow us to determine the sequences contributing to the nuclear localization process for the AR. Hypothesis III: The prostatic glands from Tfm mice may be reinduced and developed by introduction of normal AR cDNA into the urogenital sinus (UGS) of Tfm mice. To test this hypothesis, the UGS from Tfm mice will be transfected with retrovirus containing normal AR cDNA and cultured in vivo to be examined whether prostatic glands are induced. The results may demonstrate the necessary of normal AR in the mesenchyme (stroma) for the prostate development. Hypothesis IV: The Tfm syndrome in mice may be corrected by zygote microinjection (transgenic experiment) of normal AR cDNA. To test this hypothesis, the normal AR cDNA will be cloned into expression vector containing a very strong promoter (elongation factor promoter) and microinjected into the zygote of Tfm mice. The results may provide the possibility of gene therapy for the Tfm syndrome. The above studies will permit an evaluation of the role of AR in Tfm syndrome and may eventually lead to the development of new diagnostic methods and new ideas in clinical treatment of Tfm syndrome.