The mechanism of growth enhancement of 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors by high fat diet was investigated in 169 female Sprague-Dawley rats fed a semisynthetic diet containing 0, 8 or 40% corn oil by weight. The animals, most of which were intubated with 10 mg of DMBA twice during the 8th week of age, were fed an unsupplemented or fat-supplemented diet for 2 months from the 12th to the 21st week of age and finally killed during the 21st week of age. Compared to the rats fed a fat-free diet, a significantly increased yield of mammary adenocarcinoma with a shorter latency was observed in the rats fed a 40% corn oil diet. DNA synthesis of the biopsied mammary tumors more than 1 month after diet switch-over was estimated in terms of the rate of [3H]thymidine incorporation into the tissue. Significantly higher DNA synthesis of the mammary tumors biopsied from proestrous hosts was found, as compared with the tumors in similar hosts at other stages of the estrous cycle. The greatest enhancement of DNA synthesis in small and rapidly growing tumors was observed in the 40% corn oil diet group. Feeding with 40% corn oil diet always resulted in elevation of the weight percentage of linoleic acid in both simple lipids and phospholipids of the mammary tumors. These data indicate that the effects of dietary fat on mammary tumor growth might be mediated by the enhancement of tumor response to certain hormones, and that the enhanced responsiveness of tumors was associated with increased linoleic acid in tumor lipids.