The focus of this project is to investigate the molecular mechanism involved in a novel function of Daxx, which can be explored in developing strategies for autoimmune diseases. The multifunctional protein Daxx was originally identified as a potential mediator of Fas-induced apoptosis. Mutations in the Fas gene lead to an autoimmune-lymphoproliferative syndrome (ALPS), due to impaired apoptosis in lymphocytes and other immune cells. While Fas-induced cell death is initiated at the membrane, Daxx is present not only in the cytoplasm but also in the nucleus. Many potential Daxx-binding proteins have been identified, including the tumor suppressor p53 family members as well as the oncoprotein MDM2. However, the in vivo function of Daxx is poorly defined, as deletion of Daxx in germ cells leads to early embryonic lethality. During lymphocyte development and function, cell proliferation, survival and death are tightly regulated by various genes. Dysregulation in these processes may lead to autoimmune diseases. We and others identified the Fas- associated death domain (FADD) protein as a critical adaptor for Fas to recruit and activate the initiator caspase 8 during apoptotic signaling. Interestingly, FADD deficiency also has an embryonic lethal effect. Whereas lymphocytes in lymphocyte-specific FADD-deficient mice are resistant to Fas-induced apoptosis, these conditional FADD knockout mice do not develop autoimmune diseases, unlike that in Fas mutant mice. This paradox is explained by our recent study which reveals a potent, RIP1-dependent necrotic pathway unleashed in the absence of FADD. Our preliminary data show that deletion of RIP3, a member of the RIP1 family, restores normal development in FADD-deficient mice. Importantly, the resulting RIP3-/- FADD-/- double knockout (DKO) mice develop progressive autoimmune diseases, similar to that in Fas mutant mice. These data contrast a model suggesting that Daxx promotes FADD-independent apoptosis in the Fas pathway. Indeed, our preliminary data shows that Daxx-deficient T cells from conditional Daxx knockout mice have no defect in Fas-induced apoptosis. Unexpectedly, Daxx-/- T cells are hypersensitive to activation-induced cell death (AICD) when stimulated through the T cell antigen receptor (TCR). We hypothesize that in primary lymphocytes, there is an additional death pathway which is independent of the Fas-FADD-Caspase 8 axis, and is suppressed by Daxx. Therefore, targeting Daxx would promote cell death and serves as a strategy for autoimmune disease therapy. However, the molecular mechanism of this novel function of Daxx in the immune system is unclear. Therefore, we propose: 1) to elucidate the cell death pathway(s) in primary lymphocytes that is inhibited by Daxx;2) to test for the effect o genetic and pharmacological ablation of Daxx function on suppressing autoimmune diseases. Successful completion of these specific aims will provide new insight into the mechanisms of the action by Daxx and lead to better design of measures for prevention and treatment of autoimmune and related diseases.
Our study reveals a novel function for Daxx as a negative regulator of activation induced cell death (AICD), which is critical for suppressing autoimmune diseases. In this project, we will elucidate the molecular mechanism by which Daxx exerts its inhibitory effect on death responses in lymphocytes. Importantly, we will explore genetic and pharmacological means that target Daxx and promote apoptosis in lymphocytes as potential therapeutics for autoimmune diseases and related pathological conditions.
|Dowling, John P; Cai, Yubo; Bertin, John et al. (2016) Kinase-independent function of RIP1, critical for mature T-cell survival and proliferation. Cell Death Dis 7:e2379|