Cyclosporin A (CsA), a powerful immunosuppressive drug, inhibits the synthesis of lymphokines in T lymphocytes at the level of gene transcription. Using protein extracts from El4 lymphoma cells we show that the binding of lymphocyte‐specific factors interacting with the two so‐called purine boxes (Pu‐boxes) of the interleukin 2 (IL‐2) enhancer are missing in CsA‐treated cells. The CsA‐sensitive factors are newly synthesized upon induction. The most prominent factor consists of 45 kd polypeptides and contacts both Pu‐boxes at the two central G residues within the identical core sequence AAGAGGAAAA. The CsA‐mediated suppression of factor binding to the Pu‐boxes correlates well with functional studies in which the inducible, T cell‐restricted proto‐enhancer activity of Pu‐boxes was selectively repressed by CsA. These observations support the conclusion that the suppression of factor binding to the Pu‐boxes by CsA impairs the activity of IL‐2 and of further lymphokine genes, thereby inhibiting the synthesis of lymphokines in T lymphocytes.