SIGNAL TRANSDUCTION EVENTS AND THE REGULATION OF CELL GROWTH

这个项目是为了增加我们对癌细胞的了解 生物学，重点是激素不敏感的前列腺癌，雌激素 受体阴性的乳腺癌、原始神经前体肿瘤和 通过对癌症的研究开发治疗癌症的新方法 生长调节信号转导事件。这项工作目前正在进行中 主要集中在(1)肿瘤细胞生长调控的新方面 视网膜母细胞瘤基因产物，以及(2)转录调控 一种独特形式的因子和信号转导蛋白 O-糖基化。视网膜母细胞瘤蛋白目前被认为是 仅在细胞核中起作用，且仅在细胞的胃肠道时相起作用 周而复始。相反，我们发现视网膜母细胞瘤蛋白是 在对数生长过程中显著定位于细胞质。我们的 数据显示，这种外周定位在大多数研究中都被掩盖了 Rb蛋白，因为所使用的抗体只检测核型 RB。出乎意料的是，使用共聚焦显微镜和免疫共沉淀 我们发现外周Rb与细胞相关的技术 细胞骨架，外周Rb蛋白定位于生长中 对数相细胞的尖端。这些数据表明了一种新的肿瘤模型 抑癌基因的功能，其中肿瘤抑制基因产物 注册细胞间的相互作用，并将生长停止信号发送到 原子核。 研究HMG-CoA还原酶抑制剂的抗癌作用 洛伐他汀我们发现了RB的一种新的翻译后修饰 蛋白质-O-连接N-乙酰氨基葡萄糖的加成(O-GlcNAc加成)。 我们发现洛伐他汀显著增加了O-GlcNAc的含量 修饰Rb蛋白，显著阻断Rb的磷酸化 并明显改变Rb亚细胞分布。这些研究确定了 O-GlNAc加成作为抗癌药物开发的新靶点 洛伐他汀是第一个调节O-GlcNAc的药物 新陈代谢。我们已经与英国签订了一项合作协议 生物技术公司牛津糖业系统，旨在开发新的 碳水化合物导向的抗癌药物。

This project is designed to increase our understanding of cancer cell biology, with an emphasis on hormone-refractory prostate cancer, estrogen receptor-negative breast cancer, primitive neuronal progenitor tumors, and to develop a new approach to cancer treatment through the study of growth-regulatory signal transduction events. This work is currently focused on (1) novel aspects of the regulation of cancer cell growth by the retinoblastoma gene product, and (2) the regulation of transcription factors and signal transducting proteins by a unique form of O-glycosylation. The retinoblastoma protein is currently thought to function only in the cell nucleus and only in the GI phase of the cell cycle. In contrast, we found that the retinoblastoma protein is prominently localized in the cell cytoplasm during logarithmic growth. Our data show that this peripheral localization is masked in most studies of the Rb protein because the antibody used only detects the nuclear form of Rb. Unexpectedly, using confocal microscopy and coimmuno-precipitation techniques we found that peripheral Rb is associated with the cell cytoskeleton, and that peripheral Rb protein is localized in the growing tip of logarithmic phase cells. These data suggest a new model of tumor suppressor gene function, in which the tumor suppressor gene product registers cell-cell interaction and sends a growth arrest signal to the nucleus. While studying the anticancer action of the HMG-CoA reductase inhibitor lovastatin we identified a new posttranslational modification of the Rb protein- the addition of O-linked N-acetylglucosamine (O-GlcNAc addition). We found that lovastatin greatly increased the amount of O-GlcNAc modification of the Rb protein, dramatically blocked Rb phosphorylation and markedly changed Rb subcellular distribution. These studies identify O-GlNAc addition as a new target in anticancer drug development and lovastatin as the first example of a drug that regulates O-GlcNAc metabolism. We have established a cooperative agreement with the British biotechnology company Oxford GlycoSystems, aimed at developing new carbohydrate-directed anticancer drugs.