Supplementary MaterialsSupplementary Information Supplementary Information srep07691-s1. for its cell cycle inhibitor properties; it regulates early G1-S transition by inhibiting cyclin-dependent kinases in complex with cyclins A and E or D1. It was initially assumed that p21 deletion would lead to extensive tumor development but p21-deficient mice are essentially cancer-free2,3. Deficiency in p21 combined with mild autoreactive backgrounds such as 129/Sv C57BL/64 or the Gadd45a-deficient mice show severe lupus-like autoimmunity glomerulonephritis, which leads to death5,6. p21?/? mice on the autoimmunity-resistant C57BL/6 (B6) background exhibited mild autoimmune manifestations7 and it was suggested that p21 acts as a suppressor of autoimmunity. In one report, lack of p21 appeared to reduce disease in autoimmune BXSB male background8, and it was considered that this controversy was probably due to the atypical BXSB background7,9. The p21 autoimmunity-suppressing activity was reinforced by analysis of Egr-2 deficient autoreactivity-developing mice, which downmodulate p21 expression in T cells9. Data from p21?/? mice suggested a possible role for p21 in the expansion of activated but not of na?ve T CH5132799 cells7. In a different system, increased p21 expression by CD4+ T cells from elite (infection-free) HIV-exposed individuals, appeared critical for evasion of HIV infection10. In addition to regulating adaptive immune CH5132799 responses, p21 controls innate immunity, modulating macrophage activation through the NF-B activation pathway11 and inflammatory cytokine production11,12,13. p21 thus emerges as an important regulator of immunity that controls innate and adaptive responses, and maintains autoimmunity development at bay14,15,16. (lymphoproliferation spontaneous mutation) mice deficient in Fas (CD95), show defective activation-induced cell death (AICD) of restimulated T cells17. mice develop lymphadenopathy due to accumulation of double negative T cells (DN; TCR+CD4?CD8?B220+), and lupus-like autoimmune disease, probably due to CD4+ T cell hyperactivation18. One of the unexplained symptoms caused by Fas deficiency is massive hyperproliferation of DN T cells, CD4+ effector (CD44hi/CD62Lhi), memory (CD44hi/CD62Llo), and CD8+ effector/memory T CH5132799 cells in lymphoid organs. Accumulation of effector/memory T cells is critical for development of autoimmunity, as they secrete large amounts of IFN-, a cytokine necessary for lupus development in and other spontaneous or induced murine lupus models19,20,21,22. C57BL/6/(B6/mice on the autoimmune-prone MRL background (MRL/and MRL/mice. We found that p21 overexpression inhibited B6/DN T cell lymphadenopathy and decreased effector/memory T cell expansion and autoimmune symptoms. Further analysis revealed an unanticipated p21 capacity to decrease the activation of effector/memory B6/T cells and their IFN- production. p21 is a potent autoimmunity suppressor, since when overexpressed in MRL/mice, efficiently reduced death rates. Exogenous p21 effects were evident in but not in control B6 mice, indicating that autoimmune but not normal T cells require p21 to control activation and IFN- production. Therefore, therapeutic approaches that target autoimmunity but not normal responses are feasible. Results T cell-directed p21 expression inhibits CH5132799 effector/memory T cell accumulation in B6/but not in B6 mice By two months of age, B6/mice show a predisposition to autoimmunity and begin to accumulate memory and DN T cells in lymphoid organs, with development of autoimmune characteristics and lymphadenopathy17. As lack of p21 leads to increased expansion of repeatedly stimulated T cells without affecting primary T cell responses7, we hypothesized that directed transgenic p21 expression in B6/mouse T cells would Rabbit Polyclonal to PLD1 (phospho-Thr147) reduce spontaneous accumulation of effector/memory T cells and ameliorate lupus characteristics in these mice. We generated B6 and B6/mice that specifically express a human p21 transgene in T cells under the proximal Lck promoter (B6-p21tg and B6/characteristics of peripheral T cells. T cell development and differentiation in B6-p21tg mice is normal26, as confirmed by similarity of thymic populations in B6 and B6-p21tg and in B6/and B6/and accumulate with age in B6 mice, possibly due to exposure to endogenous and environmental antigens. In B6/mice, however, effector/memory CD4+ T cells accumulate to a much greater extent, as confirmed at two and four months of age (Fig. 1A, B, left). Enhanced p21 expression in T cells greatly reduced effector and memory CD4+ T cell accumulation in B6/mice (Fig. 1A). The transgene nonetheless did.