The ability of two HLA-DR-expressing human T cell clones to function as antigen-presenting cells (APC) was investigated using highly purified T cells. The results demonstrated that these T cell clones are unable to act as autonomous APC, and that recognition of nominal or alloantigens on the surface of T cells leads to a state of nonresponsiveness. The first observation was that a T cell clone with specificity for the 306-324 peptide of influenza hemagglutinin (HA), and raised from a DR1 responder, exhibited apparent degeneracy of major histocompatibility complex restriction when cultured with peptide in the presence of peripheral blood mononuclear cells (PBMC) expressing a wide variety of structurally unrelated DR types. However, when the PBMC were pulsed with peptide and washed before coculture with the clone, peptide was exclusively recognized with DR1Dw1. This implied that in the presence of soluble peptide the T cells were displaying ligand to each other, and that the third-party APC were providing costimulatory signals. To test the ability of T cells to act as autonomous APC, accessory cell-free preparations of two DR1-restricted clones were cultured with peptide in the presence or the absence of added B cell APC. T cell purity was established by the absence of proliferation in response to the mitogen phytohemagglutinin (PHA). PHA-nonresponsive T cells were completely unable to proliferate in response to peptide alone; furthermore, preculture of the HA-specific clone, in the complete absence of accessory cells, with the same concentration of peptide (1 microgram/ml) that induced optimal proliferation when presented by conventional APC, led to profound nonresponsiveness. The same phenomenon was also observed when two of three anti-DR1 alloreactive T cell clones were precultured with a DR1-expressing T cell clone. The ability of the DR1-expressing clone to induce nonresponsiveness in anti-DR1 clones correlated with recognition of the DR1 alloantigen on the DR1-expressing clone.