POLYCYCLIC HYDROCARBON METABOLISM AND CARCINOGENESIS

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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.