Diverse ubiquitin linkages regulate RIP kinases-mediated inflammatory and cell death signaling

Abstract

Members of the RIP kinase family are key regulators of inflammation and cell death signaling implicated in maintaining immune responses and proper tissue homeostasis. Increasing evidence points to post-translational modifications of RIP1, RIP2 and RIP3 as being critical for regulating their function. Ubiquitination and the E3 ligases, such as inhibitors of apoptosis (IAP) proteins and LUBAC, that direct substrate selectivity as well as the deubiquitinating enzymes, such as A20 and OTULIN, that reverse these modifications dictate the outcome of RIP kinase signaling. Perturbation of the tightly regulated RIP1, RIP2 and RIP3 ubiquitination can lead to signaling disbalance in TNF, TLR and NOD1/2-controlled pathways and result in severe human pathologies. In this review, we focus on the biological function of ubiquitin-modifying enzymes in the context of RIP1, RIP2 and RIP3 signaling. We also discuss the impact of deregulated ubiquitin networks in RIP1, RIP2 and RIP3 signaling pathways on human health.

Figure 1

The domain structure of RIP1, RIP2 and RIP3 with RIP-interacting proteins. RIP1, RIP2 and RIP3 consist of amino-terminal kinase domains that harbor phosphosites important for necroptotic signaling (RIP1/S166 and RIP3/S199/277) or NF-κB activation (RIP2/S176). The RHIM domain, located within the intermediate domain of RIP1 and the unique C terminus of RIP3, is essential for interaction of RHIM-containing proteins and necroptotic signaling. The C-terminal death domain of RIP1 mediates death receptor signaling and apoptosis while the C-terminal CARD domain of RIP2 mediates signaling via NOD1/2 binding. Distinct proteins interact with ubiquitinated RIPs and important ubiquitin sites have been identified for complex I signaling (RIP1/Lys377), NOD signaling (RIP2/Lys209) and necroptosis (RIP1/Lys115 and RIP3/Lys5). In addition, a large number of proteins, like A20, CYLD and LUBAC, use ubiquitin chains as a bridge for interaction with RIPs. Interacting proteins listed in parentheses are indirect binders (TRAF2 and c-IAPs)

Figure 2

The regulatory role of RIP1 ubiquitination in TNFR1-mediated complex I. TNF stimulation leads to the recruitment of TRADD, TRAF2, c-IAP1/2 and RIP1 to TNFR1. c-IAP proteins polyubiquitinate (K11 and K63) RIP1 and themselves, creating as a scaffold for further recruitment of LUBAC. LUBAC mediates linear polyubiquitin, leading to gene activation via IKK complex. Several DUBs have been implicated in the regulation of RIP1-dependent complex I by removing linear (CYLD and OTULIN) and K63-linked polyubiquitin chains (A20 and CYLD)

Figure 3

The regulatory role of ubiquitin on RIP1 and RIP3-dependent cell death. Inhibition of transcription or translation can divert TNF-mediated signaling to the formation of an intracellular complex II consisting of TRADD, FADD and caspase-8 in a RIP1-independent apoptotic cell death. This cell death pathway can be promoted by A20 and CYLD that deubiquitinate complex I components and thereby support the switch to complex II. A TNF-induced RIP1-dependent apoptotic cell death occurs in IAP-depleted (by IAP antagonist or KO) or LUBAC-deficient conditions leading to the assembly complex II consisting of RIP1, FADD and caspase-8. Activation of RIP1-dependent cell death under caspase-8 inhibited or deficient conditions can lead to a necroptotic form of cell death that is mediated by RIP1 and RIP3 and results in the activation of MLKL. Recent studies showed various polyubiquitin chains on components of necrosome that are potentially needed for proper formation and function of necrosome

Figure 4

The role of RIP1 and RIP3 polyubiquitination in TLR3 and TLR4 signaling. LPS stimulates TLR4 while dsRNA (or PolyI:C) stimulates TLR3. This activation of TLRs allows TIR (Toll/IL-1 receptor) domain to interact with RHIM-containing TRIF that mediates interaction with RIP1 or RIP3. RIP1-dependent activation of NF-κB is mediated by polyubiquitination by Pellino1, leading to subsequent recruitment and activation of TAB–TAK1 and IKK complexes for NF-κB gene activation. TLR3/4 can induce cell death in a TRIF–RIP1–RIP3-dependent manner, which involves RIP1, RIP3 and MLKL ubiquitination in necroptotic signaling

Figure 5

The role of polyubiquitination of RIP2 in NOD1 and NOD2 signaling. Bacterial infection involves secretion of peptidoglycan fragments DAP and muramyl dipeptide to stimulate NOD1- and NOD2-mediated signaling, which leads to RIP2 polyubiquitination and activation of NF-κB or MAPK pathways. A large number of E3 ligases and DUBs are associated with the regulation of RIP2. XIAP promotes K63-linked polyubiquitination on RIP2, leading to generation of LUBAC-mediated linear chains. This allows the activation of the TAB2/3-TAK1 and IKK complexes. Positive regulators, like Pellino3 and c-IAP1/2, also mediate K63-linked chains on RIP2 while removal of polyubiquitin chains by the DUBs OTULIN, CYLD and A20 inhibits NOD1/2-RIP2 signaling