Crystal Structure of the Caenorhabditis elegans Apoptosome Reveals an Octameric Assembly of CED-4

Abstract

The CED-4 homo-oligomer or apoptosome is required for initiation of programmed cell death in Caenorhabditis elegans by facilitating autocatalytic activation of the CED-3 caspase zymogen. How the CED-4 apoptosome assembles and activates CED-3 remains enigmatic. Here we report the crystal structure of the complete CED-4 apoptosome and show that it consists of eight CED-4 molecules, organized as a tetramer of an asymmetric dimer via a previously unreported interface among AAA(+) ATPases. These eight CED-4 molecules form a funnel-shaped structure. The mature CED-3 protease is monomeric in solution and forms an active holoenzyme with the CED-4 apoptosome, within which the protease activity of CED-3 is markedly stimulated. Unexpectedly, the octameric CED-4 apoptosome appears to bind only two, not eight, molecules of mature CED-3. The structure of the CED-4 apoptosome reveals shared principles for the NB-ARC family of AAA(+) ATPases and suggests a mechanism for the activation of CED-3.

Cellular Mechanisms Controlling Caspase Activation and Function

“…Although alterations in mitochondrial dynamics have been reported to modulate apoptosis in C. elegans, there is no evidence that the apoptosis in this organism involves mitochondrial permeabilization, and the worm apoptosome is active once EGL-1 relieves apoptosome inhibition. Similar to the mammalian apoptosome, the C. elegans apoptosome appears to contain two CED-3 molecules, but with a ratio of eight CED-4 to two CED-3 (Qi et al 2010).…”

Apaf1 apoptotic function critically limits Sonic hedgehog signaling during craniofacial development

“…Third, based on the CED4 structure of the apoptosome, the WHD is predicted to encircle the 'hutch' space into which caspase 9 fits, 13 suggesting the domain could be critical for caspase 9 placement or function. Finally, several proteins have been reported to bind Apaf1 in either inhibitory or potentiating manners; thus, it is plausible that the L375P mutation alters (either stabilizing or disrupting) such an interaction, resulting in a loss-of-function phenotype (Figure 6a).…”

“…11 Whereas the Apaf1 apoptosome in mammals exhibits sevenfold symmetry and forms a wheel-like particle, 12 the CED-4 apoptosome in C. elegans is an octamer: a symmetric conical shape of four asymmetric dimers. 13 Inside the cone is a space termed the 'hutch' that can fit two CED-3 molecules. 13 For both CED-3 and caspase 9, the processed form associates with the apoptosome and triggers the cleavage of a variety of targets associated with apoptotic cell death.…”

“…13 Inside the cone is a space termed the 'hutch' that can fit two CED-3 molecules. 13 For both CED-3 and caspase 9, the processed form associates with the apoptosome and triggers the cleavage of a variety of targets associated with apoptotic cell death. 14 Apaf1 is composed of six domains: an N-terminal caspase recruitment domain (CARD), a nucleotide-binding domain, a winged-helix domain (WHD), two helical domains and a domain at the C-terminus of the protein containing 12-13 WD-40 repeats.…”

“…The CARD domain of Apaf1 interacts with the prodomain of caspase 9, 4,12 whereas the WD-40 repeats bind to cytochrome c. 12,[15][16] Once associated with cytochrome c, Apaf1 undergoes a conformational change 11 such that ATP binds at the interface between the nucleotide-binding domain, the first helical domain and the WHD, 17 allowing further intermolecular interactions that form the apoptosome, where the WHD is predicted to be an essential mediator of oligomerization. 13 Mice with targeted deletions of Apaf1 exhibit embryonic lethality and morphological defects, highlighting the pivotal role of this protein in apoptosis during mammalian embryogenesis. [18][19] Loss of Apaf1 causes decreased apoptosis, best documented in the embryonic neuroectoderm, which results in failed neural tube closure and cleft palate, among other morphological defects.…”