Autoproteolysis of PIDD marks the bifurcation between pro-death caspase-2 and pro-survival NF-κB pathway

Abstract

Upon DNA damage, a complex called the PIDDosome is formed and either signals NF-jB activation and thus cell survival or alternatively triggers caspase-2 activation and apoptosis. PIDD (p53-induced protein with a death domain) is constitutively processed giving rise to a 48-kDa N-terminal fragment containing the leucine-rich repeats (LRRs, PIDD-N) and a 51-kDa C-terminal fragment containing the death domain (DD, PIDD-C). The latter undergoes further cleavage resulting in a 37-kDa fragment (PIDD-CC). Here we show that processing occurs at S446 (generating PIDD-C) and S588 (generating PIDD-CC) by an auto-processing mechanism similar to that found in the nuclear pore protein Nup98/96 and inteins. Auto-cleavage of PIDD determines the outcome of the downstream signaling events. Whereas initially formed PIDD-C mediates the activation of NF-jB via the recruitment of RIP1 and NEMO, subsequent formation of PIDD-CC causes caspase-2 activation and thus cell death. A non-cleavable PIDD mutant is unable to translocate from the cytoplasm to the nucleus and loses both activities. In this way, autoproteolysis of PIDD might participate in the orchestration of the DNA damage-induced life and death signaling pathways.

p53-induced protein with a death domain (PIDD) isoforms differentially activate nuclear factor-kappaB and caspase-2 in response to genotoxic stress

“…We have recently shown that PIDD isoform 1 is autoprocessed at S446 and S588 to generate a PIDD-C and a PIDD-CC fragment (Tinel et al, 2007), in agreement with previous reports (Telliez et al, 2000;Pick et al, 2006). The 11 amino-acid deletion in isoform 2 encompasses the second cleavage site used for PIDD The two C terminal fragments generated by PIDD autoprocessing have distinct functions (Tinel et al, 2007).…”

“…Two levels of regulation had already been reported: at the transcriptional level, as PIDD is a known p53 target gene , and at the post-translational level, as autoprocessing of PIDD determines its partners of interaction and, therefore, the downstream signaling events (Tinel et al, 2007). The alternative splicing of PIDD that we report here could represent a third level of regulation, acting at the post-transcriptional level.…”

“…The 11 amino-acid deletion in isoform 2 encompasses the second cleavage site used for PIDD The two C terminal fragments generated by PIDD autoprocessing have distinct functions (Tinel et al, 2007). PIDD-C is able to interact with RIP1 and NEMO to activate NF-kB in response to DNA damage (Janssens et al, 2005).…”

The death domain-containing protein Unc5CL is a novel MyD88-independent activator of the pro-inflammatory IRAK signaling cascade

“…21 Cleavage at an HFS site is also involved in the biogenesis of the nuclear envelope protein Nup98. 22 Surprisingly, by sequence comparison, we identified a bona fide HFS site in Unc5CL (aa 227-229) but not in any other ZU5-UPA-DDcontaining proteins, suggesting that Unc5CL might also undergo such autoproteolytic cleavage (Figures 1a and b and Supplementary Figure S1).…”

“…Previous studies on Nup98 and PIDD have shown that this site can be cleaved involving formation of a thioester intermediate via an N-S acyl shift. 21,22 However, cleavage of the thioester intermediate requires addition of the nucleophilic agent hydroxylamine. To test whether Unc5CL HFC also forms a thioester intermediate, we analyzed the sensitivity to hydroxylamine-induced cleavage ( Figure 1e).…”