ANETTE SUNDSTEDT*, MIKAEL SIGVARDSSON*t, TOMAS LEANDERSONt, GUNNAR HEDLUND*C, TERJE KALLAND*t,

Anergy is a major mechanism to ensure antigen-specific tolerance in T lymphocytes in the adult. In vivo, anergy has mainly been studied at the cellular level. In this study, we used the T-cell-activating superantigen staph- ylococcal enterotoxin A (SEA) to investigate molecular mech- anisms of T-lymphocyte anergy in vivo. Injection of SEA to adult mice activates CD4+ T cells expressing certain T-cell receptor (TCR) variable region 13-chain families and induces strong and rapid production of interleukin 2 (IL-2). In contrast, repeated injections of SEA cause CD4+ T-cell dele- tion and anergy in the remaining CD4+ T cells, characterized by reduced expression of IL-2 at mRNA and protein levels. We analyzed expression of AP-1, NF-cB, NF-AT, and octamer binding transcription factors, which are known to be involved in the regulation of IL-2 gene promoter activity. Large amounts of AP-1 and NF-cB and significant quantities of NF-AT were induced in SEA-activated CD4+ spleen T cells, whereas Oct-i and Oct-2 DNA binding activity was similar in both resting and activated T cells. In contrast, anergic CD4+ T cells contained severely reduced levels of AP-1 and Fos/Jun- containing NF-AT complexes but expressed significant amounts of NF-KB and Oct binding proteins after SEA stimulation. Resolution of the NF-KB complex demonstrated predominant expression of p50-p65 heterodimers in activated CD4+ T cells, while anergic cells mainly expressed the tran- scriptionally inactive p50 homodimer. These alterations of transcription factors are likely to be responsible for repression of IL-2 in anergic T cells.