POLYCYCLIC HYDROCARBON METABOLISM AND CARCINOGENESIS

本研究的总体目标是更好地了解 细胞色素P450 IA 1的调节，即，CYPIA 1，为了影响 多环碳氢化合物致癌问题 例如苯并（a）芘（B（a）P）和3-甲基胆蒽（3 MC），因为它 CYPIA 1部分负责激活这些 物质. 这个实验室已经假设需要一个多环芳烃结合 蛋白质，并分离、纯化和表征了4S蛋白质， 满足这个要求。 我们最近发现，4S PAH- 结合蛋白与甘氨酸N-甲基转移酶（GNMT）相同。 的 本更新申请的具体目的是：a）确定 B（a）P诱导的CYPIA 1表达中的4S PAH结合蛋白，和 B）确定负调节的潜在机制， 大鼠细胞色素P450 IA 1基因，即，塞浦路斯1. 关于第一 特异性目的，转导基因对4S结合的影响 将根据CYPIA 1的活性来确定蛋白质进入啮齿动物细胞的量。 (and合适的报告基因），在施用B（a）P后， 反义寡核苷酸的施用（对4S结合 蛋白）和4S蛋白的抗体将在 细胞暴露于B（a）P后CYPIA 1的诱导，以及 在4S水平下，肝脏和肝癌细胞先前暴露于PAH 结合蛋白及其mRNA。 关于第二次 具体目标，在负调控中起作用的顺式元件 将定义CYPIA 1，与CYPIA 1相互作用的反式作用蛋白质。 这些顺式元件将被分离和表征， 负调控元件和位点之间的通讯， 将研究4S结合蛋白的相互作用。

The overall objective of this research is to better understand the regulation of cytochrome P450IA1, i.e., CYPIA1, in order to impact upon the problem of carcinogenesis induced by polycyclic hydrocarbons (PAH) such as benzo(a)pyrene (B(a)P and 3-methylcholanthrene (3MC), since it is CYPIA1 that is responsible, in part, for the activation of these substances. This laboratory has postulated the need for a PAH-binding protein and has isolated, purified, and characterized a 4S protein that fulfills this requirement. We have recently established that the 4S PAH- binding protein is identical to glycine N-methyltransferase (GNMT). The specific aims of this renewal application are to a) determine the role of the 4S PAH-binding protein in the B(a)P-induced expression of CYPIA1, and b) to ascertain the mechanisms underlying the negative regulation of the rat cytochrome P450IA1 gene, i.e., CYPIA1. With regard to the first specific aim, the effect of transduction of the gene for the 4S binding protein into rodent cells will be determined upon the activity of CYPIA1 (and of suitable reporter genes) after administration of B(a)P, the effect of administration of antisense oligonucleotides (to the 4S binding protein) and of antibodies to the 4S protein will be determined upon induction of CYPIA1 after exposure of cells to B(a)P, and the effect of prior exposure of liver and hepatoma cells to PAH upon the level of the 4S binding protein and its mRNA will be ascertained. In regard to the 2nd specific aim, the cis elements that play a role in the negative regulation of CYPIA1 will be defined, the trans-acting proteins that interact with these cis elements will be isolated and characterized, and the communication between the negative regulatory element and the sites for interaction of the 4S binding protein will be investigated.