Regulation of cell surface expression of Fas (CD95) ligand and susceptibility to Fas (CD95)-mediated apoptosis in activation-induced T cell death involves calcineurin and protein kinase C, respectively

Abstract

We show that an influenza hemagglutinin-specific CD4+ murine T cell hybridoma (IP-12-7) enters the apoptotic suicide program via the Fas ligand (FasL)/Fas-mediated pathway upon T cell receptor (TCR) stimulation. These cells express Fas and FasL mRNA, cell surface Fas and intracellular FasL, but do not enter apoptosis upon Fas ligation prior to TCR stimulation. TCR stimulation additionally results in protein synthesis-dependent cell surface expression of the preformed FasL. Addition of phorbol dibutyrate (PBu2) alone was sufficient to induce susceptibility to Fas ligation induced apoptosis, while addition of both PBu2 and calcium ionophore A23187 were required to induce FasL cell surface expression. Addition of cyclosporin A completely inhibited TCR-mediated death and FasL cell surface up-regulation, but had no effect on apoptosis induced directly by Fas ligation following TCR stimulation. Inhibitors of protein kinase C (PKC) (Gö 6976 and GF 109203X) completely inhibited TCR-induced susceptibility to Fas ligation, but only partially inhibited TCR-induced cell surface expression of FasL. PKC isoenzymes alpha, beta, delta and zeta were expressed by this cell line and only the alpha and betaI isoforms translocated to the membrane fraction upon TCR stimulation. Our data suggest that in activation-induced T cell apoptosis PKC is involved in pathways that mediate the acquisition of Fas susceptibility, while calcineurin is required for cell surface expression of the preformed FasL.

Evaluation of the impact of conventional immunosuppressant on the establishment of murine transplantation tolerance – an experimental study

“…To clarify the negative impact of TAC treatment, we next tested anti‐donor antigen reactivity, as CNIs are well known to interrupt activation‐induced cells death (AICD) by down‐modulating CD95L . Splenic CD4 + CD25 − T cells were prepared from chimeras that were not treated with immunosuppressants (chimera control), TAC‐treated chimeras, or EVL‐treated chimeras on day 7 after BMT.…”

Adenosine A2A receptor‐mediated cell death of mouse thymocytes involves adenylate cyclase and Bim and is negatively regulated by Nur77

“…To determine whether the cell death receptor pathway of apoptosis plays a role in adenosine‐mediated apoptosis, the apoptosis‐inducing effect of adenosine was investigated in the presence of a FasFc chimeric molecule, which completely blocks the FasL‐mediated death of a mouse T cell hybridoma line 42, or a soluble DR5 molecule, which completely inhibits TRAIL‐mediated death of thymocytes 43. In addition, the adenosine sensitivity of thymocytes isolated from Fas‐associated death domain (FADD) dominant‐negative mice 44, which express a dominant negative mutant of FADD that blocks death receptor‐induced apoptosis, was also tested (Fig.…”

Calcineurin: Form and Function

“…In lymphocytes, calcineurin and NF-AT appear to participate in apoptosis, in part by mediating the induc-tion of Fas and Fas ligand which then interact and transduce the apopototic signal after T-cell receptor ligation (157, 226,243,421). Using a constituitive (Ca 2ϩ -and calmodulin-independent) form of calcineurin, Shibasaki and McKeon (375) demonstrated that calcineurin functions in calcium-induced apoptosis in mammalian cells deprived of growth factors and that this was a direct consequence of calcineurin's phosphatase activity.…”