PART III. REQUIREMENTS FOR GENERATION AND DOWN- REGULATION OF CYTOTOXIC T LYMPHOCYTES zyxw Cross-linking of T-cell Receptors Is Insufficient to Induce IL-2 Responsiveness (Activation) in Resting Lyt-2+ T Cells IL-4 or RIF Are Essential as Second Signal" zy HERMANN WAGNER, KLAUS HEEG, AND CONNY HARDT zyxw Department of Medical Microbiology und Inimirnology University of Ulm 0-7900 Ulm, West Germany INTRODUCTION Two signals are thought to control activation of resting Lyt-2' murine T cells. Cross-linking of T-cell receptor structures by antigen recognition is believed to induce expression of functional (high affine) interleukin-2 (IL-2) receptors (i.e., IL-2 responsiveness), whereas autocrine- or paracrine-provided IL-2 is assumed to promote their growth and maturation into cytotoxic effector cells.' This view falls short of explaining why zyxwv iri uivo and in vitro the immunogenicity of constitu- tively expressed allogeneic MHC antigens is restricted to certain accessory cells such as dendritic cells.? In what sense then differ functional effective (immuno- genic) stimulator cells from cells that present MHC antigens in a nonimmunogenic form? Is immunogenicity governed by a "costimulator activity" provided as addi- tional signal by antigen-presenting cells (APC)? In attempts to search for the postulated costimulator activity, we devised an in vitro bioassay. This bioassay is based on the experimental finding that high- density (resting) murine Lyt-2' T cells exposed to the mitogen concanavalin A (Con A) remain refractory to the growth-promoting effect of IL-2, that is, the ligand Con A lacks the ability to induce functional IL-2 receptors3Subsequently, we have extended this finding to high-density Lyt-2 zyxw ' T cells, the antigen receptors of which were cross-linked by anti-F23 monoclonal antibodies (mAb) covalently bound to sepharose beads, or by exposure to nonimmunogenic allogeneic stimula- tor cells. Here we describe that in all three systems the costimulator activity provided by the T-helper cell product interleukin-4 (IL-4) or by the macrophage product IL-2 receptor-inducing factor (RIF)' restricts the induction of IL-2 re- sponsiveness. This work zyxwvuts was supported by the SFB 377 and the BMFT. zyx 1u1