Human immunodeficiency virus type 1 (HIV-1) can lead to a profound CD4+ T-cell deficiency. To examine the functional role of HIV-1 Nef protein on the marked loss of CD4+ cells, Nef protein was expressed in and purified from Escherichia coli as a fusion protein with T7 phage gene10 product (Nef-gene10). When peripheral blood mononuclear cells (PBMC) from healthy donors were cultivated in the presence of Nef-gene10 or the gene10 product as well as interleukin-2 (IL-2), it was found that the Nef-gene10, but not the gene10 product, induced a remarkable decline in the CD4/CD8 ratio and in the response to phytohaemagglutinin of PBMC as well as of nylon wool-passed purified T cells. Nef-gene10 inhibited the proliferation of CD4+ cells, but did not kill the cells. This suppression of the IL-2-dependent proliferation of CD4+ cells by Nef-gene10 seemed to be due to enhanced production of several lymphokines, especially of interferon-gamma. Thus, Nef protein might be partly responsible for the selective depletion of CD4+ cells in HIV-1 infection. Furthermore, the Nef-induced decline in the CD4/CD8 ratio was interrupted by anti-Nef antibodies, suggesting the possibility that a vaccine which resulted in the production of such functional Nef antibodies would be useful in the treatment of HIV-1-induced immunodysfunction.