The objective of this study is to investigate the activation of the H-ras oncogene in mouse skin which has been treated with N-heterocyclic aromatic compounds dibenz[a,j]acridine (DBajA) and dibenzo[c,g]carbazole (DBC), both of which are significant occupational and environmental pollutants produced from the incomplete combustion of organic materials.
The specific aims will involve the following: 1. Determine the relative transforming abilities of DNA extracted from tumors induced by topical application of DBC and DBajA. This will be accomplished by transfecting the DNA into NIH3T3 cells and scoring transformed foci. DNA extracted tumors induced by 7, 12-dimethylbenz[a]anthracene will be used as a positive control. 2. Examine the NIH3T3 foci for the presence of an altered p21 protein, the protein encoded by the H-ras oncogene. This will be done by analyzing the electrophoretic mobility of p21 for changes indicative of mutations in the 12th and 61st codons, the two """"""""hot spots"""""""" for activation of the H-ras oncogene. 3. Examine DNA extracted from NIH3T3 foci for the presence of an activated H-ras oncogene and determine the nature of the mutation responsible for this activation. Analysis of restriction fragment length polymorphisms will be performed to determine which specific nucleotides have been mutated. These studies will investigate the activation of the H-ras oncogene in mouse skin by DBC and DBaja. The H-ras oncogene has been shown to be activated in 10-20% of all human tumors as well as in many chemically- induced laboratory animal cancers. The activation of this oncogene is believed to play a significant role in the development of cancer. The elucidation of the mechanism of oncogene activation is important in understanding the mechanism of carcinogenesis by N-heterocyclic compounds.
