P53 AND ESTROGEN IN PCNA EXPRESSION OSTEOSARCOMA CELLS

本研究的目的是分析人骨肉瘤细胞中肿瘤抑制蛋白p53与雌激素信号通路之间的串扰。p53基因是各种人类癌症中最常发生改变的基因。p53的失活被认为是骨肉瘤发展的重要一步。雌激素在成骨细胞谱系的发育和功能中起重要作用，雌激素刺激骨肉瘤细胞的增殖。然而，雌激素作用在功能性野生型p53存在和缺失时的结果尚不清楚。虽然已知p300/CBP是p53介导和雌激素受体介导的基因调控的辅助激活因子，但p53与雌激素功能之间的关系尚不清楚。我们拟以骨肉瘤细胞中的PCNA基因表达为模型，研究p53介导和雌激素介导的基因调控途径之间的功能相互关系及其与细胞增殖和细胞周期扰动的相关性。PCNA是细胞DNA复制和DNA修复机制的重要组成部分。骨肉瘤细胞中PCNA的表达受p53调控。我们的结果表明，PCNA基因是雌激素诱导的。此外，初步实验揭示了p300/CBP在p53介导PCNA表达调控中的重要作用。基于前期研究和初步数据，我们假设p53和雌激素信号通路在调控PCNA表达中存在显著的串扰。为了验证这一假设，我们将分析p53和雌激素受体的协同激活因子p300/CBP调控p53介导的PCNA基因表达的机制。我们拟确定p53-CBP通路在雌激素诱导的骨肉瘤细胞PCNA表达和增殖中的作用。由于PCNA的相对量对于确定其与其他对细胞周期重要的蛋白的相互作用很重要，因此也有假设认为，在细胞周期的不同阶段，PCNA的表达受到p53和雌激素协同作用的差异调节。为了验证这一假设，我们将分析细胞周期中p53和雌激素对PCNA转录的调控。这项拟议的研究将为雌激素在存在和不存在功能性p53的情况下在增殖和细胞周期检查点功能中的作用提供重要信息。它还将为分析类固醇激素和p53在正常和异常细胞过程（如肿瘤发生）中调节基因表达的串扰提供一个范例。

The objective of the proposed study is to analyze the cross-talk between the tumor suppressor protein p53 and estrogen signaling pathways in human osteosarcoma cells. The p53 gene is the most frequently altered gene in various human cancers. Inactivation of p53 is considered to be an important step in the development of osteosarcomas. Estrogens are important in the development and functioning of the osteoblast lineage and estrogens stimulate proliferation of osteosarcoma cells. However, consequence of estrogen action in the presence and in the absence of functional wild type p53 is not clear. Although it is known that p300/CBP is a coactivator for both p53-mediated and estrogen receptor-mediated gene regulation, the relationship between p53 and estrogen functions remains unknown. We propose to investigate the functional interrelationship between pathways of p53-mediated and estrogen-mediated gene regulation and its relevance to cell proliferation and perturbation of cell cycle, using PCNA gene expression in osteosarcoma cells as a model. PCNA is a critical component of the cellular DNA replication and DNA repair machineries. PCNA expression in osteosarcoma cells is regulated by p53. Our results show that the PCNA gene is estrogen-inducible. Furthermore, preliminary experiments have revealed a major role for p300/CBP in mediating regulation of PCNA expression by p53. Based on earlier studies and preliminary data, we hypothesize that there is significant cross-talk between p53 and estrogen signaling pathways in regulating PCNA expression. To test this hypothesis, we will analyze the mechanisms by which p300/CBP, a coactivator for both p53 and estrogen receptor, regulates p53-mediated PCNA gene expression. We propose to determine the role of p53-CBP pathway in estrogen-induced PCNA expression and proliferation of osteosarcoma cells. Since the relative amount of PCNA is important in specifying its interaction with other proteins important for the cell cycle, it is also hypothesized that PCNA expression is differentially regulated by coordinated action of p53 and estrogen in different stages of the cell cycle. To test this hypothesis, PCNA transcriptional regulation by p53 and estrogen during the cell cycle will be analyzed. The proposed study will provide important information on the role of estrogen in proliferation and cell cycle checkpoint function in the presence and in the absence of functional p53. It will also provide a paradigm for analyzing cross-talk between steroid hormones and p53 in regulating gene expression in both normal and abnormal cellular processes such as oncogenesis.