T cell precursors or prothymocytes present in the spleen and bone marrow of mice lack the differentiation antigens found on mature thymocytes. Incubation of prothymocytes with the hormone products of thymic epithelial cells can trigger induction of antigens like Thy 1.2. We have recently found that interleukin-2 will also induce Thy 1.2. We have found that a 4 day incubation of prothymocytes (derived from the spleens of nu/nu mice) with natural and recombinant interleukin-2 in the presence of human thymic epithelial cells results in their synergistic effect on prothymocyte differentiation as measured by Thy 1.2 induction using a fluorescence activated cell sorter (FACS). This coculture condition also caused a synergistic enhancement of proliferation of the prothymocytes and the appearance of a population of large lymphoblasts. Analysis of the presence of Thy 1.2 surface antigen indicated that induction of this surface antigen occurs on both medium sized and large lymphoblast populations. Under these conditions induction of Ly 2 surface antigen was not detected and prothymocytes at the end of the coculture did not manifest IL-1 or mitogen responsiveness. Incubation of prothymocytes in the presence of thymic epithelial cells alone resulted in a lesser degree of Thy 1.2 induction and proliferation and in the consistent induction of low levels of interleukin-2 (0.5-1 unit/ml). Thymosin fraction V did not have a synergistic effect with IL-2 on the cell proliferation. The enhanced response of prothymocytes to IL-2 in the presence of thymic epithelial cells presumably results from the enhanced induction of IL-2 receptors and/or responsiveness. The observation offers further support for a role of interleukin-2 in the regulation of T cell ontogeny.