MOLECULAR MECHANISMS OF CYTOCHROME P4501A1 EXPRESSION

细胞色素 P4501A1 表达的分子机制

• 批准号: 6043519
• 负责人: BHAGAVATULA MOORTHY
• 金额: $ 18.86万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 2003-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6043519
• 关键词: NAD(P)H dehydrogenase; animal genetic material tag; aromatic hydrocarbon receptor; breast neoplasms; carcinogen testing; chemical carcinogenesis; cytochrome P450; enzyme activity; gene induction /repression; gene mutation; genetic promoter element; glucuronosyltransferase; glutathione transferase; laboratory rat; lung neoplasms; methylcholanthrene; mutagen testing; neoplasm /cancer genetics; nucleic acid sequence; receptor binding

DESCRIPTION: Environmental chemicals such as polycyclic aromatic hydrocarbons (PAHs), which are present in cigarette smoke and charcoal-broiled meats, cause cancer in laboratory animals and are suspected human carcinogens. The stated long-term goal of this application is to prevent chemically-induced malignancies in humans. The proposed work is based on studies indicating that some PAHs cause long-term induction in rats of cytochrome P450 1A1 (P450 1A1), an enzyme that can play important roles in carcinogenesis. Specific Aim 1 will investigate the mechanisms of 3-methylcholanthrene (3MC)-mediated persistent induction of P450 1A1 in rats. Rats will be challenged with 3MC, and nuclear A receptor levels (gel mobility shift assays), P450 1A1 gene transcription rates (run-on transcription), mRNA levels (hybridization), and P450 1A1 activities/contents will be determined at selected time points. Specific Aim 2 will test the hypothesis that persistent MC metabolites contribute to the sustained P450 1A1 induction. Animals will be treated with [3H]-3MC, the nucle will be isolated, and it will be determined whether 3MC metabolites interact with the Ah receptor. Specific Aim 3 will test the hypothesis that 3MC causes sequence-specific DNA adduction on the regulatory regions of the P450 1A1 promoter, leading to novel mechanisms by which DNA adducts mimic the 3MC-Ah receptor complex and upregulate P450 1A1 gene expression. Plasmids carrying P450 1A1 promoter will be used to generate in vivo or in vitro adducts with 3MC. The adducted plasmids will be transiently transfected into Hepa-1 cells t determine whether 3MC-DNA adducts on the promoter will induce P450 1A1 gene expression in the absence of the inducer. Specific Aim 4 will measure P450 1A1 activities and DNA adduct levels (32P-postlabeling) in rat lung and mammary tissues to test the hypothesis that 3MC elicits sustained induction of P450 1A and persistent DNA damage in target organs for carcinogenesis. In Specific Aim 5, the activities/levels of P450 1B1, glutathione S-transferase-a, NAD(P)H quinone reductase, and UDP-glucuronosyl transferases will be determined to tes whether 3MC persistently induces other enzymes of the Ah gene battery.

描述：环境化学物质，例如多环芳烃 碳氢化合物（PAHS），存在于香烟烟和 木炭肉肉，导致实验室动物癌，可疑 人类致癌物。 该应用程序的长期既定目标是 防止人类化学引起的恶性肿瘤。 拟议的工作是 基于表明某些PAH导致大鼠长期诱导的研究 细胞色素P450 1A1（P450 1A1），一种可以发挥重要作用的酶 在癌变中。 具体目标1将调查 3-甲基胆碱（3MC）介导的P450 1A1持续诱导 老鼠。 大鼠将受到3MC和核A受体水平的挑战（凝胶 移动性转移测定），P450 1A1基因转录率（跑步 转录），mRNA水平（杂交）和P450 1A1 活动/内容将在选定的时间点确定。 具体的 AIM 2将检验以下假设，即持续的MC代谢产物有助于 持续的P450 1A1诱导。 动物将用[3H] -3MC处理， 将隔离核，并确定是否3MC 代谢物与AH受体相互作用。 特定目标3将测试 假设3MC会引起调节性序列特异性DNA的内收。 P450 1A1启动子的区域，导致了新的机制 加合物模拟3MC-AH受体复合物和上调P450 1A1基因 表达。 携带P450 1A1启动子的质粒将用于生成 带有3MC的体内或体外加合物。 加合的质粒将是 瞬时转染Hepa-1细胞t确定3MC-DNA是否是 启动子上的加合物将在不存在的情况下诱导P450 1A1基因表达 诱导者。 特定目标4将测量P450 1A1活动和DNA 大鼠肺和乳腺组织中的加合物水平（32p固定标签）以测试 假设3MC引起P450 1A的持续诱导和持续的诱导 靶器官的DNA损伤用于致癌。 在特定目标5中 P450 1B1的活动/水平 还原酶和UDP-葡萄糖基转移酶将确定为TES 3MC是否持续诱导AH基因电池的其他酶。