Epithelial cells, growing in tissue-like colonies, have marked and characteristic morphological and biochemical responses to both tumor promoters and to transformation that are similar in many ways. The highly differentiated MDCK line is extremely sensitive to tumor promoting agents. A characteristic morphological signature results from exposure to promoters that is visualized with fluorescent anti-cytokeratin antibodies and by video-enhanced phase microscopy. Within 15 minutes, the stationary epithelial colonies are converted to clusters of independent and highly motile cells. Quantitative morphometric analysis of the cell response to various promoters will measure the rate of change of: a) total inter-nuclear spacing, b) nuclear shape and c) integrated curvature of cellular processes within the colonies as a function of promoter dose. Analysis will use a simple graphics tablet and a small computer. We will correlate the in vitro response with promoter efficacy, as observed in vivo, to determine if our measurements constitute a short-term test for complete and second stage promoters. Permanent transformation by a single dose of a complete carcinogen or by infection with Moloney sarcoma virus results in a morphological signature and an altered metabolic response similar to that induced by tumor promoters. The metabolic response to an altered cell shape is also markedly changed. A transfection system will be established with MDCK in order to insert several activated oncogenes in addition to v-mos, some of which should give similar morphological and biochemical responses. Complete carcinogens give a prompt response which in some ways resembles that of promoters and the possibility of the promotion-like activity resulting from protein alkylation by these agents will be tested using ultimate carcinogens. Phosphorylation changes, including those specific to tyrosine, in the presence of TPA have been noted by others and we find these to be located principally in the nuclear matrix. We will examine phosphorylation in the presence of other promoters and in cells transformed by specific oncogenes. The activity of protein kinase C will be determined and the possible association of this activity with skeletal elements in promoted and transformed cells will be measured.
