This project will study the mechanism of cellular differentiation and of tumor promotion in the human epidermal keratinocyte culture system. In order to meet these objectives, a system has been developed in which human keratinocytes can be reversibly maintained in an undifferentiated, basal state by adaptation to growth in 10 to the minus 8 M phorbal myristate acetate (PMA). These cells retain an undifferentiated morphology and resist cornification even after the cultures become confluent. However, after growth in the absence of PMA for one week, the adapted cultures regain their ability to differentiate. The observation that some cells can grow in the presence of PMA without undergoing terminal differentiation allows selection of a homogeneous population. The phenotype of the cells in the PMA-adapted cultures will be determined by measuring their growth characteristics, dependence on feeder cells and hormones, production of plasminogen activator, degree of cornification, and keratin patterns. The conversion from the undifferentiated state to terminal differentiation in these synchronized cultures will then be studied. Growth requirements necessary to allow differentiation will be determined. An understanding of the control of differentiation in his system may lead to strategies for intervention in tumors of epithelial origin.