Abstract
Death receptor signaling is initiated by the assembly of the death-inducing signaling complex, which culminates in the activation of the initiator caspase, either caspase-8 or caspase-10. A family of viral and cellular proteins, known as FLIP, plays an essential role in the regulation of death receptor signaling. Viral FLIP (v-FLIP) and short cellular FLIP (c-FLIPS) inhibit apoptosis by interfering with death receptor signaling. The structure and mechanisms of v-FLIP and c-FLIPS remain largely unknown. Here we report a high resolution crystal structure of MC159, a v-FLIP derived from the molluscum contagiosum virus, which is a member of the human poxvirus family. Unexpectedly, the two tandem death effector domains (DEDs) of MC159 rigidly associate with each other through a hydrophobic interface. Structure-based sequence analysis suggests that this interface is conserved in the tandem DEDs from other v-FLIP, c-FLIPS, and caspase-8 and -10. Strikingly, the overall packing arrangement between the two DEDs of MC159 resembles that between the caspase recruitment domains of Apaf-1 and caspase-9. In addition, each DED of MC159 contains a highly conserved binding motif on the surface, to which loss-of-function mutations in MC159 map. These observations, in conjunction with published evidence, reveal significant insights into the function of v-FLIP and suggest a mechanism by which v-FLIP and c-FLIPS inhibit death receptor signaling.
Highlights
Apoptosis plays a central role in the development and homeostasis of metazoans [1,2,3,4,5]
One important family of such proteins is the viral FLICE-inhibitory proteins (v-FLIPs), which were initially identified in molluscum contagiosum virus and in several ␥-herpesviruses [22]. Viral FLIP (v-FLIP) have been shown to block the activation of the death receptors at the level of death-inducing signaling complex (DISC) assembly [22]
The hydrogen bonds between Asp-34 and Arg-97 are predicted to be conserved in a majority of these proteins (Fig. 2). This analysis strongly suggests that the rigid structure of MC159 and the interface determinants between the tandem death effector domains (DEDs) are representative of v-FLIPs, c-FLIP, caspase-8, and caspase-10
Summary
FADD, in turn, uses its death effector domain (DED) to interact with the N-terminal tandem DEDs of procaspase-8 or procaspase-10, thereby linking these initiator caspases to the activated death receptors within the DISC (19 –21). In addition to the tandem DEDs, c-FLIPL contains a caspase-like domain For both c-FLIPS and c-FLIPL, their tandem DEDs appear to antagonize death receptor signaling in the same manner when expressed at high levels (24, 29 –31, 33–36). The DED-containing proteins are known to control apoptosis from death receptors, the underlying mechanisms remain largely unknown. This is in part due to the technical difficulty in working with membrane-associated protein complexes. All structures conform to a common fold comprising six antiparallel ␣-helices, which is shared by caspase recruitment domain (CARD) [42]
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