REGULATION OF ORNITHINE DECARBOXYLSE BY PHORBOL ESTERS

佛波酯对鸟氨酸脱羧基的调节

• 批准号: 2092254
• 负责人: ANDREW P BUTLER
• 金额: $ 12.72万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-03-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2092254
• 关键词: DNA binding protein; affinity chromatography; antisense nucleic acid; biological signal transduction; cell growth regulation; chemical carcinogenesis; chloramphenicol acetyltransferase; cycloheximide; gene expression; gene mutation; genetic mapping; genetic promoter element; genetic regulatory element; genetic transcription; hepatocellular carcinoma; insulin; laboratory rat; liver cells; messenger RNA; molecular cloning; molecular genetics; neoplastic cell culture for noncancer research; nucleic acid hybridization; nucleic acid sequence; oncogenes; ornithine decarboxylase; phorbols; phosphorylation; posttranslational modifications; protein kinase; reporter genes; retinoate; transcription factor; transfection /expression vector; tumor promoters; DNA binding protein; affinity chromatography; antisense nucleic acid; biological signal transduction; cell growth regulation; chemical carcinogenesis; chloramphenicol acetyltransferase; cycloheximide; gene expression; gene mutation; genetic mapping; genetic promoter element; genetic regulatory element; genetic transcription; hepatocellular carcinoma; insulin; laboratory rat; liver cells; messenger RNA; molecular cloning; molecular genetics; neoplastic cell culture for noncancer research; nucleic acid hybridization; nucleic acid sequence; oncogenes; ornithine decarboxylase; phorbols; phosphorylation; posttranslational modifications; protein kinase; reporter genes; retinoate; transcription factor; transfection /expression vector; tumor promoters

Tumor promoters, such as 12-O-tetradecanoylphorbol-13-acetate (TPA), and transforming oncogenes disrupt normal controls of gene expression and cell proliferation by acting on intracellular signal transduction pathways. One of the earliest changes in gene expression elicited by TPA, and by several oncogenes, is induction of ornithine decarboxylase (ODC). The goal of this project is to understand how the regulation of ODC is altered during the progression of normal cells to neoplasia. Towards this aim, we will compare the mechanisms by which TPA and a physiological growth factor, insulin, regulate ODC mRNA levels in a series of normal and transformed rat liver cells. We hypothesize that insulin and TPA each activate protein kinase cascades which regulate ODC by mechanisms with both common and distinct features. In order to understand these pathways, we propose to characterize the molecular events leading to ODC induction through the following specific aims. (1.) What are the DNA sequence elements required to regulate ODC transcription? The cis-acting regulatory elements in the rat ODC gene will be identified using transient expression assays. Those elements required for basal expression, induction by TPA and insulin, and inhibition by retinoic acid will be mapped. Qualitative and/or quantitative changes in ODC expression and the activity of these cis-acting elements will be studied in a series of normal, v-raf and v-raf + v-myc- transformed rat liver epithelial cells (RLE), and in the H35 hepatoma cell. (2.) What are the DNA-binding proteins required for the regulation of ODC transcription? The factors which regulate ODC transcription will be identified and characterized by protein-DNA binding assays; novel factors will be cloned. (3.) How is the activity of transcription factors required for ODC expression regulated? We will determine if transcription factors involved in ODC regulation are phosphorylated in response to TPA or insulin, and if so, whether the phosphorylation has effects on either DNA binding or transcriptional activation. In addition, we will determine whether the product of the c-raf-1 oncogene acts as an intermediate in signal transduction pathways involved in the regulation of ODC transcription or translation. The relationship of these regulatory factors to tumorigenesis will be assessed in the normal and transformed RLE. The results of these studies will provide mechanistic insight into the regulation of proliferation and gene expression by insulin and TPA in normal and transformed liver epithelial and hepatoma cells, and will contribute to an understanding of alterations in gene expression during tumorigenesis.

肿瘤启动子，例如12-O-四烷酰基-13-乙酸酯（TPA）和 转化致癌基因破坏基因表达和细胞的正常对照 通过作用于细胞内信号转导途径的增殖。 一 TPA引起的基因表达的最早变化以及几个 癌基因是鸟嘌呤脱羧酶（ODC）的诱导。 目标的目标 项目是了解ODC的调节如何改变 正常细胞向肿瘤的进展。 为了这个目标，我们将 比较TPA和生理生长因子的机制， 胰岛素，调节一系列正常和转化大鼠的ODC mRNA水平 肝细胞。 我们假设胰岛素和TPA每个都激活蛋白 激酶级联反应通过具有常见和共同的机制来调节ODC 独特的功能。 为了理解这些途径，我们建议 表征导致ODC诱导的分子事件 遵循特定目标。 （1.）需要什么DNA序列元素 调节ODC转录？ 顺式作用调节元素 大鼠ODC基因将使用瞬态表达测定识别。 那些 基础表达所需的元素，TPA和胰岛素诱导，以及 视黄酸的抑制作用将被映射。 定性和/或 ODC表达的定量变化和这些顺式作用的活性 元素将以一系列正常的V-RAF和V-RAF + V-MYC-进行研究。 转化的大鼠肝上皮细胞（RLE）和H35肝癌细胞。 （2.）ODC调节所需的DNA结合蛋白是什么 转录？ 调节ODC转录的因素将是 以蛋白-DNA结合测定为特征；新因素 将被克隆。 （3.）如何需要转录因子的活性 对于ODC表达调节？ 我们将确定转录因子是否 响应于TPA或 胰岛素，如果是这样，磷酸化是否对任何一个DNA都有影响 结合或转录激活。 此外，我们将确定 C-RAF-1癌基因的乘积是否充当中间体 ODC调节涉及的信号转导途径 转录或翻译。 这些监管因素的关系 肿瘤发生将在正常和转化的RLE中进行评估。 这 这些研究的结果将提供有关机械的洞察力 通过胰岛素和TPA在中调节增殖和基因表达 正常和转化的肝上皮和肝癌细胞，将 有助于理解基因表达的改变 肿瘤发生。