INTRACELLULAR MECHANISMS OF TUMOR PROMOTION

肿瘤促进的细胞内机制

• 批准号: 6341878
• 负责人: BRUCE E. MAGUN
• 金额: $ 31.13万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-07-01 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6341878
• 关键词: DNA binding protein; DNA footprinting; calcium; cellular oncology; cycloheximide; endothelin; enzyme activity; gel mobility shift assay; gene expression; gene induction /repression; genetic enhancer element; intermolecular interaction; intracellular; messenger RNA; mitogen activated protein kinase; phosphorylation; protein biosynthesis; protein kinase; regulatory gene; thapsigargin; tissue /cell culture; transcription factor; tumor promoters

The ability of tumor promoters to stimulate clonal selection and outgrowth of initiated cell populations is generally attributed to their ability to induce a coordinated change in the expression of various immediate early genes associated with cell proliferation. We have shown that both the phorbol ester tumor promoter 12-0-tetradecanoyl-phorbol acetate and the sesquiterpene tumor promoter thapsigargin can effect the coordinate regulation of immediate early genes including c-fos and the endogenous murine retrovirus VL30. Although proximal targets have been identified for several major classes of tumor promoters, the precise pathways by which disparate tumor promoters act to inhibit the Ca2 plus -ATPase of the endoplasmic reticulum. However, it is not known how this aberration of calcium homoestasis is transduced into induction of c-fos and VL30. In our progress report, we present data indicating that thapsigargin treatment results in an inhibition of protein synthesis, which is associated with decreased MAP kinase phosphatase (MKP-1) activity and increased activation of the stress-activated protein kinases SAPK/JNK and p38/HOG1. Based on these observations, we hypothesize that treatment with thapsigargin results in the activation of protein kinase R (PKR), which phosphorylates eIF2 alpha. Phosphorylation of elF2 alpha results in a block to translation initiation, and decreased synthesis of MKP-1. Loss of MKP-1 results in increased activation of MAP kinases, and this leads to increased phosphorylation of the transcription factor Elk-1 and induction of c-fos mRNA. We propose to test the applicability of this model to calcium-dependent induction of VL30 and c-fos. We plan to identify the transcription factors responsible for induction of VL30. We will also characterize the role of CBF/A, a putative negative regulator of VL30 expression which acts via the VL30 CArG element. Since the c-fos promoter bears significant sequence similarity to the VL30 enhancer, including the presence of a CArG motif, we will determine whether CBF/A and other modulators of VL30 enhancer, including the presence of a CArG motif, we will determine whether CBF/A and other modulators of VL30 expression can also modulate c-fos expression. The role of a PKR-dependent pathway in mediating the response to thapsigargin will be determined for both VL30 and c-fos. These experiments will include determination of possible PKR-dependent changes in the phosphorylation and activation of the transcription factor(s) operating on the promoters for these two genes. The research described in this proposal will provide new insight into the molecular mechanisms by which tumor promoters cause coordinated changes in the expression of proliferation-associated genes.

肿瘤启动子刺激克隆选择和生长的能力 启动细胞群的数量通常归因于它们的能力 诱导各种即早期表达的协调变化 与细胞增殖相关的基因。 我们已经证明，两者 佛波酯肿瘤促进剂12-0-十四烷酰基-佛波醇乙酸酯和 倍半萜肿瘤促进剂毒胡萝卜素可影响协调 立即早期基因的调节，包括 c-fos 和内源性 鼠逆转录病毒VL30。 尽管已经确定了近端目标 几类主要的肿瘤促进剂，其精确途径 不同的肿瘤促进剂可抑制 Ca2+-ATP 酶 内质网。 但目前尚不清楚这种偏差是如何产生的 钙稳态被转导为 c-fos 和 VL30 的诱导。 在我们的进度报告中，我们提供的数据表明毒胡萝卜素 治疗会抑制蛋白质合成，即 与 MAP 激酶磷酸酶 (MKP-1) 活性降低相关 应激激活蛋白激酶 SAPK/JNK 的激活增加 p38/HOG1。 基于这些观察，我们假设治疗 毒胡萝卜素会激活蛋白激酶 R (PKR)，从而 磷酸化 eIF2 α。 eF2 α 的磷酸化导致 阻断翻译起始，并减少 MKP-1 的合成。 损失 MKP-1 的增加导致 MAP 激酶的激活增加，这导致 转录因子 Elk-1 磷酸化和诱导增加 c-fos mRNA。 我们建议测试该模型的适用性 VL30 和 c-fos 的钙依赖性诱导。 我们计划确定 负责诱导 VL30 的转录因子。 我们也会 描述 CBF/A 的作用，CBF/A 是 VL30 的推定负调节因子 通过 VL30 CArG 元件起作用的表达。 由于c-fos启动子 与 VL30 增强子具有显着的序列相似性，包括 存在 CArG 基序，我们将确定 CBF/A 和其他 VL30 增强子的调节剂，包括 CArG 基序的存在，我们 将确定CBF/A和VL30表达的其他调节剂是否可以 还调节 c-fos 表达。 PKR 依赖性途径的作用 将确定 VL30 和 VL30 介导对毒胡萝卜素的反应 c-fos。 这些实验将包括确定可能的 PKR 依赖性磷酸化和激活的变化 对这两个基因的启动子起作用的转录因子。 本提案中描述的研究将为以下问题提供新的见解： 肿瘤促进者引起协调变化的分子机制 增殖相关基因的表达。