Essential requirement for caspase-8/FLICE in the initiation of the Fas-induced apoptotic cascade

Abstract

This study provides the first genetic evidence that caspase-8 occupies an essential and apical position in the Fas signaling pathway and suggests that caspase-8 may participate broadly in multiple apoptotic pathways.

Caspase-8/FLICE functions as an executioner caspase in anticancer drug-induced apoptosis

“…Stimulation of the parental Jurkat A3 cells resulted in the cleavage of caspase-8 by anti-CD95, staurosporine and etoposide, whereas no detectable caspase-8 could be observed in the caspase-8 mutant cells (Figure 4). As previously shown (Juo et al, 1998), the de®ciency in caspase-8 expression rendered these cells resistant to anti-CD95 (Figure 4b), thus demonstrating its absolute requirement for CD95 signaling. Due to the lack of caspase-8 expression also no cleavage of caspase-3 and PARP could be observed upon CD95 triggering (Figure 4a).…”

“…(b) 3610 4 CD95-sensitive vector control cells (Vector; black bars) or Jurkat cells overexpressing c-FLIP (cFLIP; white bars) were stimulated with medium (Control), anti-CD95 (a-CD95; 1 mg/ml), staurosporine (Stauro; 2.5 mM), etoposide (Etopo; 25 mg/ml) or mitomycin C (Mito; 25 mg/ml) for 24 h. Induction of apoptosis was assessed by propidium iodide staining of hypodiploid apoptotic nuclei and¯ow cytometry. The mean values of triplicate cultures are shown address this point, we made use of a caspase-8-de®cient Jurkat clone (Juo et al, 1998). Stimulation of the parental Jurkat A3 cells resulted in the cleavage of caspase-8 by anti-CD95, staurosporine and etoposide, whereas no detectable caspase-8 could be observed in the caspase-8 mutant cells (Figure 4).…”

“…The CD95-resistant Jurkat subline, Jurkat-R, was generated by continuous culture in the presence of anti-CD95 mAb (IgG 3 , 1 mg/ml, BioCheck, MuÈ nster, Germany) for 6 months . Caspase-8 de®cient Jurkat cells and the parental Jurkat cell line A3 were kindly provided by J Blenis (Harvard Medical School, Boston, MA, USA) (Juo et al, 1998). Stable transfectants of Jurkat cells overexpressing c-FLIP (Kataoka et al, 1998) or Bcl-2 were a gift from J Tschopp (Lausanne, Switzerland) and H Walczak (Heidelberg, Germany), respectively.…”

Mechanisms of apoptosis by c-Myc

“…TNF family receptors use an adaptor system to directly activate caspases and they are subject to complex modulation (Ashkenazi and Dixit, 1998). Ligand binding causes receptor homotrimerization and recruitment on the cytoplasmic face of the membrane of a death-inducing signaling complex (DISC), which includes the adaptors FADD and TRADD and procaspase-8 as crucial physiological death e ectors (Juo et al, 1998;Varfolomeev et al, 1998;Yeh et al, 1998;Zhang et al, 1998). A brief outline of the CD95/ Fas and TNf receptor systems follows since c-Myc has been implied to act both upstream and downstream of each (Janicke et al, 1994;Klefstrom et al, 1994Klefstrom et al, , 1997Hueber et al, 1997;Wang et al, 1998a).…”

“…CD95/Fas also activates the Jnk pathway but there is con¯icting evidence regarding how. In one study where caspase-8 was genetically ablated, Jnk activation by CD95/Fas was defective (Juo et al, 1998). However, other studies imply activation occurs upstream or in parallel to FADD (Wajant et al, 1998), possibly through the putative CD95/Fasinteracting adaptor protein Daxx, which is proposed to activate the Jnk pathway through binding the Mekk1 kinase Ask1 (Yang et al, 1997;Chang et al, 1998).…”