CLONING OF THE RAT MDR GENE FAMILY AND REGULATION IN NORMAL AND NEOPLASTIC LIVER

大鼠 MDR 基因家族的克隆及其在正常和肿瘤肝脏中的调节

• 批准号: 3853518
• 负责人: S S THORGEIRSSON
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3853518
• 关键词: acetylaminofluorene; chemical carcinogenesis; cytochrome P450; dioxins; gene expression; genes; genetic regulation; liver cells; methylcholanthrene; molecular cloning; multidrug resistance; neoplasm /cancer genetics; neoplastic transformation; preneoplastic state; tissue /cell culture; tumor promoters

Multidrug resistance is the phenomenon by which cells become cross- resistant to a range of unrelated compounds in response to exposure to a single agent; this resistance is the result of overexpression of a 170 Kd membrane protein, p-glycoprotein, which is encoded by the mdr gene(s). Previously, we have shown that exposure of rats to xenobiotic agents such as the carcinogens aflatoxin B1, isosafrole and 2-acetylaminofluorene (2-AAF) causes increased expression of mdr mRNA in the liver. To further study the mechanism by which xenobiotics regulate mdr gene expression, we have employed a primary hepatocyte culture system. Exposure of isolated hepatocytes to methycholanthrene (MC), 2-AAF but not 2,3,7,8 - tetrachlorodibenzo-p-dioxin (TCDD) increased the expression of mdr mRNA expression; concomitant increases in cytochrome P4501A gene(s) expression were also observed with these agents. Inhibition of protein synthesis also increased the expression of mdr mRNA in these cells. The enhanced mdr expression caused by these compounds is a result of increased transcription. These data suggest that mdr expression is regulated by a protein that is distinct from the Ah receptor. To further investigate the mechanism of mdr regulation it was necessary to isolate the rat mdr genes. We have identified that the rat mdr gene family is comprised of three members; one of these genes has been isolated and characterized. Sequence analysis of a complete cDNA for a rat mdr cDNA indicated a high degree of identity to the mouse mdrlb gene (mdrl); thus, this rat cDNA was designated the mdrlb gene. Further studies on the 5' promoter and possible 3' mRNA stability regions are being performed. We observed in normal liver a slight but significant zonal difference of distribution of mdr transcripts (zone 1 > zone 3). Significant increase of mdr transcripts during regeneration was observed mainly in zone 1 hepatocytes. Our data suggest that the increased mdr expression might represent a subpopulation of preneoplastic nodules which possibly undergoes malignant transformation more specifically than GST-P positive nodules.

多药耐药是细胞交叉耐药的现象。 对一系列不相关的化合物有抗性， 单剂;这种耐药性是170 Kd的过度表达的结果。 膜蛋白，p-糖蛋白，其由MDR基因编码。 以前，我们已经表明，大鼠暴露于外源性物质， 作为致癌物的黄曲霉毒素B1、异黄樟油素和2-乙酰氨基芴 （2-AAF）引起肝脏mdr mRNA表达增加。 进一步 为了研究外源性物质调控mdr基因表达的机制， 采用了原代肝细胞培养系统。 隔离的暴露 肝细胞对甲基胆蒽（MC）、2-AAF，但对2，3，7，8- 四氯二苯并二恶英（TCDD）可增加mdr mRNA的表达 表达;细胞色素P4501 A基因表达伴随增加 也观察到了这些药物。 抑制蛋白质合成 并增加mdr mRNA的表达。 增强的 由这些化合物引起的MDR表达是由于增加的 转录。 这些数据表明，mdr的表达是由一个 与Ah受体不同的蛋白质。 为了进一步研究 MDR的调控机制有必要分离大鼠MDR 基因. 我们已经确定大鼠mdr基因家族由以下组成： 三个成员;其中一个基因已被分离和表征。 对大鼠mdr cDNA的完整cDNA序列分析表明， 与小鼠mdrlb基因（mdrl）的同一性程度;因此，该大鼠cDNA 被命名为mdrlb基因。 对5'启动子和 可能的3 ′ mRNA稳定区正在进行。 我们观察到 正常肝组织中， MDR转录物（区1 >区3）。 MDR显著增加 再生过程中的转录本主要在第1区观察到 肝细胞 我们的数据表明，mdr表达的增加可能 代表了一个肿瘤前结节的亚群， 恶性转化比GST-P阳性更特异 结节