Deubiquitinases in the regulation of NF-κB signaling

Abstract

Nuclear factor-kappa B (NF-κB) is a critical regulator of multiple biological functions including innate and adaptive immunity and cell survival. Activation of NF-κB is tightly regulated to preclude chronic signaling that may lead to persistent inflammation and cancer. Ubiquitination of key signaling molecules by E3 ubiquitin ligases has emerged as an important regulatory mechanism for NF-κB signaling. Deubiquitinases (DUBs) counteract E3 ligases and therefore play a prominent role in the downregulation of NF-κB signaling and homeostasis. Understanding the mechanisms of NF-κB downregulation by specific DUBs such as A20 and CYLD may provide therapeutic opportunities for the treatment of chronic inflammatory diseases and cancer.

Figure 1

Structural domains in A20 and CYLD. (A) A20 contains an N-terminal OTU domain responsible for the DUB activity of A20. The catalytic cysteine residue Cys103 is also important for binding to the E2 enzymes Ubc13 and UbcH5c. There is an IKKβ phosphorylation site at Ser381 downstream of the OTU domain. A20 contains seven zinc finger domains (C₂H₂) in its C-terminus. ZnF4 confers A20 E3 ligase activity and is also involved in TAX1BP1 interactions. ZnF6 and ZnF7 are important for targeting A20 to lysosomes. In addition, A20 zinc fingers have been implicated in A20 oligomerization and A20 binding to ABINs and NEMO. (B) CYLD contains a C-terminal USP domain responsible for its DUB activity. The N-terminus of CYLD contains three CAP-Gly (CAP) domains and two proline-rich (PR) motifs. The first two CAP domains mediate binding to microtubules whereas the third CAP domain regulates NEMO interactions. CYLD also contains a TRAF2 binding site (PVQES) and a phosphorylation site cluster between CAP domains 2 and 3.

Figure 2

NF-κB signaling pathways downstream of TNFR1, TLR4 and TCR/CD28. TNFR1, TLR4 and TCR/CD28 engagement trigger the K63-linked ubiquitination of RIP1, TRAF6 and MALT1, respectively. Each of these ubiquitinated signaling molecules are targeted for deubiquitination by either A20, CYLD, Cezanne or USP21, as indicated. MALT1 cleaves A20 to inactivate its function. CYLD also removes polyubiquitin chains from TAK1 in the TCR pathway.

Figure 3

Temporal regulation of NF-κB signaling downstream of TNFR1. During early stages of TNFR1 signaling (signal initiation—15-30 min after receptor engagement), RIP1 undergoes K63-linked ubiquitination, which activates IKK and NF-κB. CYLD is inactivated by IKK-mediated phosphorylation. At later times (0.5-6 h), A20 and IκBα expression is induced by NF-κB as part of a negative feedback loop. A20 interacts with TAX1BP1, Itch, RNF11 and possibly ABIN-1 and YMER to form the A20 ubiquitin-editing complex. A20 removes K63-linked ubiquitin chains from RIP1 and catalyzes the formation of K48-linked chains to trigger RIP1 degradation. A20 also disrupts cIAP1 binding to Ubc13 and promotes Ubc13 ubiquitination and degradation.