Genetic deletion of faim reveals its role in modulating c-FLIP expression during CD95-mediated apoptosis of lymphocytes and hepatocytes

Abstract

Fas-apoptosis inhibitory molecule (FAIM) is inducibly expressed in B lymphocytes and had been shown to antagonize Fas-mediated killing of B-cell lines in vitro. However, its mechanism and role in vivo are unknown. We have generated faim À/À mice and found these mutants to be viable. In contrast to fas À/À mice, faim À/À mice have normal B-and T-cell populations. However, faim À/À B cells and thymocytes show increased sensitivity to Fas-triggered apoptosis in vitro, and faim À/À mice suffer greater mortality and exhibit exacerbated liver damage in response to Fas (CD95) engagement in vivo. The lack of FAIM results in greater activation of caspase-8 and -3 in Fas-stimulated thymocytes. CD95 or Fas (APO-1/TNFRSF6) is a member of the tumor necrosis factor (TNF) receptor superfamily and is capable of inducing apoptosis in a variety of cell-types. 1,2 The physiological ligand for Fas is CD178 (FasL), which is expressed mainly by activated T cells. 3 In mice, mutation in fas (lpr mice) or fasl (gld mice) leads to lymphoproliferation, the accumulation of abnormal lymphocytes, autoantibody production, and this ultimately results in systemic autoimmunity. 3,4 Thus, Fasmediated apoptosis is critical for regulating the function and homeostasis of lymphocytes.Fas-signaling is triggered upon the binding of FasL, which leads to the trimerization of Fas and the formation of the downstream death-inducing signaling complex (DISC) that comprises the cytosolic adapter protein FADD and procaspase-8. 5 Procaspase-8 undergoes auto-cleavage to generate active caspase-8, which in turn leads to the activation of caspase-3 and execution of apoptosis. As apoptosis plays an important role in physiology, its activation process must be tightly regulated. Hence, other than activators, negative regulators of Fas-signaling also exist. One of these inhibitors of Fas-mediated apoptosis is the cellular FLICE-inhibitory protein (c-FLIP) which is known to antagonize Fas-signaling by interfering with the recruitment of procaspase-8 to the DISC. 6 Fas-apoptosis inhibitory molecule (FAIM) was also cloned as an inducibly expressed, anti-apoptotic protein that antagonized Fas-triggered cell death of B-cell lines in vitro. 7 However, its mode of action is unknown. FAIM is highly conserved in evolution and widely expressed in all tissues although it bears no homology to other known proteins. 7 Although most tissues express the short isoform of the protein (FAIM-S), neuronal cells express both FAIM-S and a long isoform, FAIM-L. 8 FAIM-S was shown to promote neurite outgrowth through a mechanism involving NF-kB and Ras-ERK activation, 8 whereas FAIM-L was demonstrated to protect neurons from cell death in response to Fas and TNFR1 engagement. 9 Apart from its role in protecting B-lymphocytes and neuronal cells from death receptor-induced apoptosis, 7,9 FAIM was also shown to improve the survival of HEK-293 cells in serum-free media 10 and CHO cells in fed-batch cultures. 11 This suggests a potential application of FAIM in biotechnology through its role in e...

Fas Apoptosis Inhibitory Molecule Regulates T Cell Receptor-mediated Apoptosis of Thymocytes by Modulating Akt Activation and Nur77 Expression

“…Subsequently, it was also shown to play a role in NF-B activation during neurite outgrowth (25). We also recently demonstrated that it played a role in Fas-triggered apoptosis of lymphocytes and hepatocytes (27). In this report, we further demonstrate a role for FAIM in TCR-mediated apoptosis of thymocytes using cells with loss and gain of function of FAIM.…”

“…Induced by TCR Stimulation and Inhibits TCR-mediated Apoptosis of Thymocytes-As FAIM is induced by antigen receptor stimulation in B cells (24,27), we examined whether TCR cross-linking could up-regulate FAIM expression in thymocytes. WT thymocytes expressed a basal level of FAIM ex vivo, and we noticed an increase in FAIM protein levels in these cells following an 18-h treatment with a high dose (10 g/ml) of anti-CD3 and anti-CD28 (1 g/ml) antibodies (Fig.…”

“…Apoptosis Assay-Cells with DNA fragmentation was determined by propidium iodide staining as described (27). Briefly, thymocytes were treated with plate-bound anti-CD3 plus anti-CD28 antibodies for the indicated time and fixed in ethanol.…”

“…We had previously inactivated faim in mice and found that FAIM regulates Fas-triggered apoptosis of lymphocytes and hepatocytes by modulating cellular FLICE-like inhibitory protein expression, hence affecting the binding of caspase-8 to Fas (27). Here, we examined the role of FAIM in TCR-triggered apoptosis of thymocytes and demonstrated that FAIM was upregulated by TCR engagement and that it modulated TCR-induced apoptosis of thymocytes.…”

Pharmacological Modulation of Caspase-8 in Thymus-Related Medical Conditions

“…Caspase-8 is active in medullary, semimature heat-stable antigen hi SP thymocytes as a consequence of strong TCR stimulation. The death receptors-Fas, FADD, Fas apoptotic inhibitory molecule (Huo et al, 2009), and FADD-like IL1b-converting enzyme inhibitory protein (FLIP)-are involved upstream of caspase-8 activation, whereas BH3 interacting domain death agonist (Bid) and HDAC7 are activated downstream. Finally, caspase-8 has been linked to the protein glucocorticoid-induced leucine zipper (Gilz) in mediating glucocorticoid-induced thymocyte apoptosis (Pozzesi et al, 2014).…”