Expression of the type 1 human immunodeficiency virus Nef protein in T cells prevents antigen receptor - mediated induction of interleukin 2 mRNA ( signal

Stable transformants of the Jurkat T-cell line have been obtained that express either of two distinct forms of the type 1 human immunodeficiency virus nefgene: the nefI-encoded protein (Nef-1) contains alanine, glycine, and vpliw at positions 15, 29, and 33, respectively; the protein specified by nef-2 (Nef-2) has threonine, arginine, and alanine at the corresponding positions. When Jurkat ceUs or their Nef-2expressing transformants are treated with phorbol 12myristate 13-acetate (PMA) plus either phytohemagglutinin (PHA) or antibodies against C D ~ E , T-cell receptor B chain, or CD2, there is a prompt increase in interleukin 2 (IL-2) mRNA and intracellular calcium and in the IL-2 receptor a chain on the cell surface. Although cells expredng Nef-1 also induce calcium mobilization and the production of IL-2 receptor a chain, the formation of IL-2 mRNA is blocked in response to these stimuli. Moreover, Nef-1-expressing cells trpasfected with a plasmid in which the IL-2 promoter is fused to the chloramphenicol acetyltransferase (CAT) gene fail to induce CAT following treatment with PMA and PHA. By contrpst, the parental and Nef-2-containing cells induce CAT normally. Nef-1-expressing cells can produce IL-2 mRNA in response to a combination of PMA and ionomycin, although much less efnciently than the parental Jurkat cells or Nef-2sxpressing cells. These LIndings, and others described herein, suggest that the virally encoded Nef protein interferes with a signal emanating from the T-cell receptor complex that induces IL-2 gene transcription. activation of a variety of lymphokine genes, including the interleukin 2 (IL-2) gene. These lymphokines are required both for the clonal expansion of the antigen-reactive T cell and for the efficient recruitment of other hemopoietic cells involved in the immune response. Nef has been reported to bind GTP and to be a GTPase (14, properties it shares with the a subunits of trimeric G proteins and with Ras proteins, molecules which have known roles in signal transduction. Moreover, like members of the Src family, which are also involved in signal transduction, Nef is myristoylated at its N terminus and is associated with membranes (15, 16). These properties prompted us to determine whether Nef affects a T cell’s ability to transduce the correct signal upon antigen binding. We reasoned that interference with signal transduction in HIV-1-infected T cells could potentially alter the levels of lymphokine production and might be expected to have profound effects upon the immune response. Our results show that T cells constitutively expressing one form of Nef (Nef-1) are severely impaired in their ability to produce IL-2 mRNA in response to agents that mimic antigen binding to the antigen receptor. By contrast, cells that express Nef-2, which differs in three amino acids from Nef-1, induce IL-2 mRNA normally following the antigenic stimulus. Additional comparisons of the properties of Nef-1and Nef-2-producing cells with their parental cells are also dis-