RIP1's function in NF-κB activation: from master actor to onlooker

Abstract

The receptor interacting protein kinase 1 (RIP1) is a crucial component of the TNFR1 response. In this issue of Cell Death and Differentiation, Wong et al. has set a cat among the pigeons by challenging the commonly accepted model in which RIP1 is essential for TNFR1-induced NF-kB activation. Their new data will force the scientific community to adapt and refine the model of NF-kB activation. Tumor necrosis factor (TNF) is a multifunctional cytokine. Upon binding to TNFR1, it activates distinct pathways with diametrically opposed consequences: killing cells or promoting survival. Its protective effect is achieved mainly by activating the NF-kB pathway, which induces the transcription of a set of pro-survival genes. In most cells, exposure to TNF is lethal only if the NF-kB signaling pathway is inhibited. Because NF-kB has a crucial role in the pathological consequences of TNF action, the mechanism of its activation has attracted the attention of scientists over many years. The data collected so far indicate an important role for RIP1 as a signaling node that contributes to TNFR1’s life and death decisions and have led to the development of a commonly accepted model (reviewed by Wertz and Dixit and Skaug et al.). According to this model, exposure to TNF results in recruitment of a complex consisting of TRADD, RIP1, TRAF2, cIAP1 and cIAP2 at the receptor, allowing cIAP1 and cIAP2 to conjugate RIP1 with K-linked polyubiquitin chains. The addition of these chains to RIP1 has two major consequences. First, it prevents RIP1 from activating cell death signaling pathways, which is dependent on FADD and caspase-8 in apoptosis and on RIP3 in necroptosis. Second, it creates a platform for the recruitment of the protein kinase TAK1 (which acts in concert with the regulatory proteins TAB2 and TAB3) and the IkB kinase complex IKKa–IKKb–NEMO. Both TAB and NEMO were shown to dock at K-linked polyubiquitin chains, and it is believed that the close proximity of TAK1 to the IKK complex on RIP1’s K-polyubiquitin chains is sufficient for TAK1 to activate IKKb by phosphorylation. Once activated, IKKb phosphorylates IkBa, a signal for the K-ubiquitination and proteasomal degradation of IkBa. Releasing the inhibitory effect of IkBa then permits NF-kB dimers to translocate to the nucleus and transactivate pro-survival genes. The de-ubiquitinating enzymes A20 and CYLD have been identified as negative regulators that edit RIP1 K-polyubiquitin chains, thereby limiting the duration of the NF-kB response. In the absence of cIAP1 and cIAP2, RIP1 does not get K-ubiquitinated and TNF exposure induces RIP1-dependent cell death. Therefore, in the present model, RIP1 is crucial for cell survival through activation of the NF-kB pathway. In the current issue from Cell Death and Differentiation, Wong et al. re-examined RIP1’s functions downstream of the TNFR1 using wt and ripk1 / primary and SV40 large T immortalized mouse embryonic fibroblasts (MEFs). When the authors treated the cells with a combination of TNF and the IAP antagonist compound A, they observed that wt MEFs succumbed to the treatment but ripk1 / MEFs did not. These results are consistent with several recently published studies and confirm that RIP1 has a pro-cell death function in the absence of cIAP1 and cIAP2 activity. More interestingly, the authors report that TNF alone has a minor impact on the viability of ripk1 / MEFs, and that a significant difference between the survival of wt and ripk1 / MEFs is observed only when NF-kB action is blocked by the translation inhibitor cycloheximide. These results are remarkable because TNF-induced activation of NF-kB had been shown to protect cells from death. Therefore, although these new data confirm a pro-survival function of RIP1, they seriously question the obligate role of RIP1 in TNFR1-dependent NF-kB activation. These doubts were confirmed when Wong et al. demonstrated that TNF-induced IkBa degradation and recovery, as well as RelA nuclear translocation, occurs normally in both primary and transformed ripk1 / MEFs. The MEF results provided by Wong et al. are at odds with previous studies that report a crucial role for RIP1 in NF-kB activation. For example, Kelliher et al. reported that the nuclear extract from TNF-treated Ripk1 / Abelson virustransformed pre-B cells failed to bind to an NF-kB probe in electrophoretic mobility shift assay. Ea et al. showed that the K-polyubiquitination on lysine 377 of RIP1 is required for NF-kB activation in human Jurkat T cells by serving as a docking site for the recruitment of TAK1 and the IKK complex. These studies might be reconciled by considering a cell-typespecific role for RIP1 in NF-kB activation, perhaps highlighting the pitfalls of not integrating cell specificity in many of the established signaling models. Differences in responses