TOXICOLOGICAL CONTROL MECHANISMS OF HUMAN CYP1A2 GENE

人CYP1A2基因的毒理学控制机制

• 批准号: 2193835
• 负责人: LINDA C QUATTROCHI
• 金额: $ 16.69万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-30 至 1999-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2193835
• 关键词: DNA binding protein; DNA footprinting; RNase protection assay; antisense nucleic acid; carbopolycyclic compound; cytochrome P450; cytotoxicity; dioxins; environmental toxicology; gene expression; genetic enhancer element; genetic promoter element; genetic regulatory element; human tissue; liver function; northern blottings; nucleic acid sequence; polymerase chain reaction; site directed mutagenesis; tissue /cell culture; toxicant interaction; toxin metabolism; transcription factor; transfection; western blottings

Toxicity of the chemical class of xenobiotics that include polycyclic aromatic hydrocarbons (PAH) and halogenated hydrocarbons, such as polychlorinated biphenyls and dibenzodioxins, most likely occurs through control of gene expression. The molecular mechanism is largely unknown, but is currently believed to involve, In part, a cytosolic receptor, the aryl hydrocarbon or Ah-receptor. If the mechanism responsible for the many toxic effects associated with these classes of chemicals is to be unraveled, it will be necessary to study other genes that may be regulated through additional cellular mediators. This grant proposal focuses on the human cytochrome P4501A2 gene (CYP1A2), a member of the PAH-inducible CYP1A gene family that is prominent in human liver, and metabolizes drugs, such as acetaminophen, caffeine, environmental agents such as arylamines and dietary constituents, such as heterocyclic amines and aflatoxins. The molecular mechanism for the regulation of the human CYP1A2 gene will be studied through the characterization of cis-acting elements and identification of trans-acting factors utilizing transient transfection assays and in vitro DNA binding assays, such as DNase I footprinting. In vivo footprinting studies in human primary hepatocytes will be conducted to examine the temporal relationship among transcription factors in regulating CYP1A2 gene expression. Several model systems will be utilized for the proposed research, including human hepatoma cell lines, human liver and non-proliferating cultures of human hepatocytes. A Cell Culture Core and Human Tissue Bank will provide the necessary support for these studies. It is believed that a combination of these molecular and cell culture approaches will elucidate the mechanism by which this important gene is regulated.

包括多环在内的化学类外源物质的毒性 芳香烃 (PAH) 和卤代烃，例如 多氯联苯和二苯并二恶英，最有可能通过 基因表达的控制。分子机制很大程度上未知， 但目前认为部分涉及胞质受体 芳基烃或Ah-受体。如果机制负责很多 与这些类别化学品相关的毒性作用应 解开后，有必要研究其他可能受到调控的基因 通过额外的细胞介质。该拨款提案的重点是 人类细胞色素 P4501A2 基因 (CYP1A2)，PAH 诱导基因的成员 CYP1A基因家族在人类肝脏中很重要，并且代谢药物， 例如对乙酰氨基酚、咖啡因、环境剂（例如芳基胺） 以及膳食成分，例如杂环胺和黄曲霉毒素。这 人类CYP1A2基因调控的分子机制将是 通过顺式作用元件的表征进行研究 利用瞬时转染鉴定反式作用因子 分析和体外 DNA 结合分析，例如 DNase I 足迹分析。在 将在人类原代肝细胞中进行体内足迹研究 检查转录因子之间的时间关系 调节 CYP1A2 基因表达。将使用多个模型系统 对于拟议的研究，包括人类肝癌细胞系、人类 肝脏和人肝细胞的非增殖培养物。细胞培养 核心和人体组织库将为这些提供必要的支持 研究。人们相信这些分子和细胞的结合 文化方法将阐明这一重要的机制 基因受到调控。