The choline deficient-methionine low diet (CMD) is unique among hepatocarcinogenic treatments, producing hepatocellular carcinomas due to the absence of a nutrient rather than the addition of a chemical carcinogen. This diet can damage DNA, an initiation effect, and may do so through lipid peroxidation; derangement in cell control mechanisms due to repeated necrogenic damage; or aberrant methylation. Promotional effects of the CMD diet on many other carcinogens has been repeatedly demonstrated. We postulate that an initiation period can be separated from promotion, i.e., a period early in the process which causes irreversible damage to DNA but which alone will not lead to tumor development; if followed by a promoter, however, tumors will develop. This is a necessary first step to elucidate mechanisms and properly interpret findings at different stages in the lengthy process. We also propose to characterize the sequential development of preneoplastic lesions in the CMD diet alone, to elucidate the role of these lesions. This will include relevant markers such as hypomethylation, peroxidation, xenobiotic metabolism and cytoskeletal damage. We will also examine DNA damage in lesions and at critical time points in the process. It is the working hypothesis that CMD causes initial DNA damage due to a combination of aberrant methylation and peroxidation-generated effects, "fixes" that damage due to its necrogenic stimulation of cell proliferation much as PH does, and then promotes the damage due to ongoing cell proliferation. Examining these questions will clarify the role of an important dietary constituent in carcinogenesis.