The role of CAP3 in CD95 signaling: new insights into the mechanism of procaspase-8 activation

Golks, Alexander; Brenner, Dirk; Schmitz, Ingo; Watzl, Carsten; Krueger, Andreas; Krammer, Peter H.; Lavrik, Inna N.

doi:10.1038/sj.cdd.4401766

Cited by 33 publications

(21 citation statements)

References 30 publications

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“…This also fits to our previous studies of caspase-8 prodomain generation at the DISC. 13 The analysis of procaspase-8 and -10 processing in the lysates at later time points after CD95 stimulation further supported our findings ( Figure 2). Indeed, the intermediate procaspase-10 cleavage product increased up to 40% of the total procaspase-10 amounts over the first 30 min following CD95 stimulation (Figure 2a), while the p43/p41 levels of procaspase-8 always remained below 20% (Figure 2b).…”

Section: Resultssupporting

confidence: 87%

Molecular architecture of the DED chains at the DISC: regulation of procaspase-8 activation by short DED proteins c-FLIP and procaspase-8 prodomain

et al. 2015

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The CD95/Fas/APO-1 death-inducing signaling complex (DISC), comprising CD95, FADD, procaspase-8, procaspase-10, and c-FLIP, has a key role in apoptosis induction. Recently, it was demonstrated that procaspase-8 activation is driven by death effector domain (DED) chains at the DISC. Here, we analyzed the molecular architecture of the chains and the role of the short DED proteins in regulating procaspase-8 activation in the chain model. We demonstrate that the DED chains are largely composed of procaspase-8 cleavage products and, in particular, of its prodomain. The DED chain also comprises c-FLIP and procaspase-10 that are present in 10 times lower amounts compared with procaspase-8. We show that short c-FLIP isoforms can inhibit CD95-induced cell death upon overexpression, likely by forming inactive heterodimers with procaspase-8. Furthermore, we have addressed mechanisms of the termination of chain elongation using experimental and mathematical modeling approaches. We show that neither c-FLIP nor procaspase-8 prodomain terminates the DED chain, but rather the dissociation/association rates of procaspase-8 define the stability of the chain and thereby its length. In addition, we provide evidence that procaspase-8 prodomain generated at the DISC constitutes a negative feedback loop in procaspase-8 activation. Overall, these findings provide new insights into caspase-8 activation in DED chains and apoptosis initiation.

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Section: Resultssupporting

confidence: 87%

Molecular architecture of the DED chains at the DISC: regulation of procaspase-8 activation by short DED proteins c-FLIP and procaspase-8 prodomain

et al. 2015

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“…Furthermore, it has been shown that CD95 DISC formation at the membrane peaks minutes after stimulation and that the amount of the active CD95 DISC is markedly reduced 1 h after stimulation (21). The kinetics of DISC formation contradict our results (Fig.…”

Section: Cd95 Stimulation With Different Concentrations Of Lz-cd95l Rcontrasting

confidence: 56%

CD95 Stimulation Results in the Formation of a Novel Death Effector Domain Protein-containing Complex

Lavrik

Möck

Golks³

et al. 2008

Journal of Biological Chemistry

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Stimulation of CD95 (APO-1/Fas) by its natural ligand CD95L (APO-1L/FasL) leads to the formation of the death-inducing signaling complex. Here we report that upon CD95 stimulation in several T and B cell lines, a novel signaling complex is formed, which we term complex II. Complex II is composed of the death effector domain proteins as follows: procaspase-8a/b, three isoforms of c-FLIP (c-FLIP L , c-FLIP S , c-FLIP R ), and FADD. Notably, complex II does not contain CD95. Based on our findings we suggest that CD95 signaling includes two steps. The first step involves formation of the death-inducing signaling complex at the cell membrane. The second step involves formation of the cytosolic death effector domain protein-containing complex that may play an important role in amplification of caspase activation.Apoptotic cell death is common in multicellular organisms. Apoptosis can be triggered by a number of factors, including UV-or ␥-irradiation, chemotherapeutic drugs, and signaling from death receptor (1). CD95 (APO-1/Fas) is a member of the death receptor family, a subfamily of the tumor necrosis factor receptor superfamily (1-3). Cross-linking of CD95 with its natural ligand CD95L (CD178) (4) or with agonistic antibodies such as anti-APO-1 (5) induces apoptosis in sensitive cells.The death-inducing signaling complex (DISC) 3 is formed within seconds after CD95 stimulation. The DISC consists of oligomerized, probably trimerized CD95, the adaptor molecule FADD, two isoforms of procaspase-8 (procaspase-8/a and procaspase-8/b), procaspase-10, and c-FLIP L/S/R (6 -10). The interactions between the molecules at the DISC are based on homotypic contacts. The death domain of the receptor interacts with the death domain of FADD, whereas the death effector domain (DED) of FADD interacts with the N-terminal tandem DEDs of procaspases-8, -10, and c-FLIP L/S/R . The binding of procaspase-8 to the DISC results in processing of the zymogene, for which a two-step mechanism has been described (11). The first cleavage step generates the two subunits p43/p41 and p12. In a second cleavage step, the active enzyme subunits p18, p10, and the prodomains p26/p24 are produced. As a result the active caspase-8 heterotetramer p10 2 -p18 2 is released into the cytosol to propagate the apoptotic signal.Two CD95 signaling pathways were established (12). Type I cells are characterized by high levels of CD95 DISC formation and increased amounts of active caspase-8. Activated caspase-8 directly leads to the activation of downstream effector caspases-3 and -7. Type II cells are characterized by lower levels of CD95 DISC formation and thus lower levels of active caspase-8. In this case, signaling requires an additional amplification loop that involves the cleavage by caspase-8 of the Bcl-2 family protein Bid to generate truncated Bid and subsequent truncated Bid-mediated release of cytochrome c from mitochondria (10). The release of cytochrome c from mitochondria results in apoptosome formation followed by activation of procaspase-9, which in tur...

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“…This processing occurs at Asp residues located between the prodomain and the large and the small catalytic subunits (Figure 2). 30,34 We have shown recently that cleavage between the prodomain and the large and the small catalytic subunits occurs simultaneously, though there is a preference towards a cleavage between the large and small catalytic subunits. 35 This processing results in the generation of a number of the cleavage products, including p43/p41, p26/ p24, CAP3, p30, p18 and p10 (Figure 2).…”

Section: Procaspase-8 and Its Activation At The Discmentioning

confidence: 99%

“…35 This processing results in the generation of a number of the cleavage products, including p43/p41, p26/ p24, CAP3, p30, p18 and p10 (Figure 2). [34][35][36][37] At the first cleavage step, the N-terminal p43/p41 and the C-terminal p30 cleavage products are generated. Importantly, these cleavage products already possess catalytic activity.…”

Section: Procaspase-8 and Its Activation At The Discmentioning

confidence: 99%

Regulation of CD95/Fas signaling at the DISC

2011

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CD95 (APO-1/Fas) is a member of the death receptor (DR) family. Stimulation of CD95 leads to induction of apoptotic and nonapoptotic signaling pathways. The formation of the CD95 death-inducing signaling complex (DISC) is the initial step of CD95 signaling. Activation of procaspase-8 at the DISC leads to the induction of DR-mediated apoptosis. The activation of procaspase-8 is blocked by cellular FLICE-inhibitory proteins (c-FLIP). This review is focused on the role in the CD95-mediated signaling of the death effector domain-containing proteins procaspase-8 and c-FLIP. We discuss how dynamic cross-talk between procaspase-8 and c-FLIP at the DISC regulates life/death decisions at CD95. Open QuestionsThe exact mechanism of CD95-mediated non-apoptotic signaling is not established. The stoichiometry of the CD95 DISC is a question of future studies. New molecules may be found to be associated with the DISC. CD95Signaling CD95 (also called APO-1; Fas; fas antigen; tumor necrosis factor receptor superfamily member 6, TNFRSF6) is a member of the DR family, a subfamily of the tumor necrosis factor receptor superfamily. 1 All members of the DR family are characterized by a cytoplasmic region termed death domain (DD). 2,3 DD are 80-100 amino-acid long motifs involved in the transduction of the apoptotic signal. Crosslinking of CD95 with its natural ligand (L), CD95L (CD178) 4 or with agonistic antibodies such as anti-APO-1 5 induces apoptosis in sensitive cells.Stimulation of CD95 has been also reported to trigger nonapoptotic pathways. [6][7][8][9][10][11][12] However, details of CD95-mediated non-apoptotic pathways remain largely unknown. Importantly, it has been shown that membrane-bound CD95L is essential for the cytotoxic activity, whereas soluble CD95L appears to promote autoimmunity and tumorigenesis via induction of non-apoptotic pathways, in particular NF-kB. 13 Future studies should elucidate more details on the mechanism of nonapoptotic action of CD95L.Binding of CD95L or agonistic antibodies to CD95 leads to formation of a receptor complex at the cellular membrane, which was named DISC. 14 The DISC consists of oligomerized receptors, the DD-containing adaptor molecule FADD/MORT1 (Fas-associated DD), procaspase-8 (FLICE, MACHa, Mch5), procaspase-10 and the c-FLIP (Figure 1). [15][16][17] The interactions between the molecules at the DISC are based on homotypic contacts. The DD of the receptor interacts with the DD of FADD, whereas the death effector domain (DED) of FADD interacts with the N-terminal tandem DEDs of procaspases-8, -10 and c-FLIP. As a result of DISC formation procaspase-8 is activated at the DISC resulting in the formation of the active caspase-8, which leads to apoptosis.The initial events of DISC formation have not been clarified yet. Pre-oligomerization of CD95 via the pre-ligand assembly domain has been reported to have an important role in apoptosis initiation. 18 Recently, there have been several new reports on the X-ray structure of the complex formed by isolated CD95 and FADD DDs. 19,20 ...

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The role of CAP3 in CD95 signaling: new insights into the mechanism of procaspase-8 activation

Cited by 33 publications

References 30 publications

Molecular architecture of the DED chains at the DISC: regulation of procaspase-8 activation by short DED proteins c-FLIP and procaspase-8 prodomain

Molecular architecture of the DED chains at the DISC: regulation of procaspase-8 activation by short DED proteins c-FLIP and procaspase-8 prodomain

CD95 Stimulation Results in the Formation of a Novel Death Effector Domain Protein-containing Complex

Regulation of CD95/Fas signaling at the DISC

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