POSTTRANSLATIONAL RESPONSES OF STEROID HORMONES

描述（改编自申请人摘要）：类固醇激素 对翻译后蛋白运输的控制知之甚少， 但对细胞的激素调节可能是重要的机制， 功能 调查人员发现了一个以前未知的， 遗传上不同的糖皮质激素调节的翻译后反应 控制小鼠乳腺细胞表面的稳定定位 肿瘤病毒（MMTV）的糖蛋白在病毒感染的大鼠肝癌细胞。 糖皮质激素改变细胞表面相关的终浓度 MMTV通过降低转运过程中的周转率来抑制糖蛋白。 糖皮质激素对糖蛋白稳定性的这种新作用表明 存在一类新的糖皮质激素调节成分， 胞吐途径中的降解系统。 独特的不可分割MMTV 糖蛋白的类固醇调节的降解系统的目标是 通过内切蛋白水解位点的定点诱变设计。 的 研究人员列出了这项建议的四个具体目标。1)精确 不可切割的MMTV糖蛋白基因的片段将被突变或 糖皮质激素调节的周转和细胞 改变的糖蛋白的表面积累， 转染的肝癌细胞，以确定潜在的降解 糖皮质激素调节降解系统的信号。 2）。混合 糖蛋白基因将由MMTV的离散区域构建， 糖蛋白基因和人胰岛素受体基因，以测试 特异性MMTV糖蛋白结构域介导 糖皮质激素调节的稳定性反应。 3)透化的 重组MMTV糖蛋白运输的肝癌细胞系统 并允许膜不可渗透的试剂进入 细胞，将用于识别细胞位置和代谢 糖皮质激素调节的降解系统的要求。 MMTV糖蛋白运输的体外重建将用作 用于分离、表征和鉴定 糖皮质激素调节的降解系统。 最终，抗体和 将产生cDNA探针以分离降解因子以探索 其糖皮质激素依赖性表达和功能。

DESCRIPTION (Adapted from the applicant's abstract): The steroid hormone control of posttranslational protein trafficking is a poorly understood, but potentially important mechanism for hormonal regulation of cellular function. The investigator has uncovered a previously unknown and genetically distinct glucocorticoid-regulated posttranslational response that controls the stable localization of cell surface mouse mammary tumor virus (MMTV) glycoproteins in viral-infected rat hepatoma cells. Glucocorticoids alter the final concentration of cell surface-associated MMTV glycoproteins by decreasing the turnover rate during transport. This novel effect of glucocorticoids on glycoprotein stability suggests the existence of a new class of glucocorticoid-regulated components of a degradative system in the exocytic pathway. A unique uncleavable MMTV glycoprotein target for the steroid regulated degradative system was designed by site-directed mutagenesis of the endoproteolytic site. The investigator lists four Specific Aims of this proposal. 1) Precise segments of the uncleavable MMTV glycoprotein gene will be mutated or substituted in vitro and the glucocorticoid-regulated turnover and cell surface accumulation of the altered glycoproteins examined in transfected hepatoma cells in order to define potential degradative signals for the glucocorticoid-regulated degradative system. 2). Hybrid glycoprotein genes will be constructed of discrete regions of the MMTV glycoprotein gene and the human insulin receptor gene to test the ability of specific MMTV glycoprotein domains to mediate the glucocorticoid-regulated stability response. 3) A permeabilized hepatoma cell system, which reconstitutes MMTV glycoprotein trafficking in vitro and allows the entry of membrane impermeable reagents into the cell, will be used to identify the cellular location and metabolic requirements of the glucocorticoid-regulated degradative system.4) The in vitro reconstitution of MMTV glycoprotein trafficking will be used as an assay to isolate, characterize and identify components of the glucocorticoid regulated degradative system. Eventually, antibody and cDNA probes will be generated to isolate degradative factors to explore their glucocorticoid dependent expression and function.