The ability of glucocorticoids to suppress cellular immune functions, including the cytotoxic activity of natural killer cells, is well known. However, the molecular mechanism(s) of glucocorticoid-mediated suppression of cellular cytotoxicity mediated by natural killer cells is not understood. We have investigated the effects of cortisol on protein expression and cytotoxic function of natural killer cells using NK3.3, a well-characterized, cloned human natural killer cell line. Cortisol, at concentrations up to 2 microM, does not significantly alter the viability or proliferative capacity of NK3.3 cells. However, micromolar concentrations of cortisol induce the expression of a small set of proteins which are not synthesized by NK3.3 cells in the absence of cortisol, and repress the synthesis of another set of proteins including several phenotypic determinants and cytokines. In the presence of added cortisol, the synthesis of perforin mRNA was partially repressed. However, the most striking effect of cortisol on this NK clone was its repression of granzyme A synthesis. In conjunction with the downregulation of adhesion proteins, NK3.3 cells cultured in the presence of cortisol exhibit a reduced capacity to form conjugates with K562 target cells. Whereas cortisol treatment of NK3.3 cells causes an approximately 50% decrease in their ability to form conjugates with K.562 target cells, the cytotoxic function of these cells is completely abolished under the same conditions. This first report of hormonal regulation of granzyme expression and the strong correlation between granzyme A repression and cytotoxic function suggests that cortisol may regulate NK function by repression of granzyme A synthesis. In addition to demonstrating the significant influence of cortisol on natural killer cell function, these studies provide a model system for elucidation of molecular mechanism(s) whereby glucocorticoids repress cellular immune function, especially with respect to natural killer cells.