Coordinated Regulation of Toll-Like Receptor and NOD2 Signaling by K63-Linked Polyubiquitin Chains

Abstract

K63 polyubiquitin chains spatially and temporally link innate immune signaling effectors such that cytokine release can be coordinated. Crohn's disease is a prototypical inflammatory disorder in which this process may be faulty as the major Crohn's disease-associated protein, NOD2 (nucleotide oligomerization domain 2), regulates the formation of K63-linked polyubiquitin chains on the I kappa kinase (IKK) scaffolding protein, NEMO (NF-B essential modifier). In this work, we study these K63-linked ubiquitin networks to begin to understand the biochemical basis for the signaling cross talk between extracellular pathogen Toll-like receptors (TLRs) and intracellular pathogen NOD receptors. This work shows that TLR signaling requires the same ubiquitination event on NEMO to properly signal through NF-B. This ubiquitination is partially accomplished through the E3 ubiquitin ligase TRAF6. TRAF6 is activated by NOD2, and this activation is lost with a major Crohn's disease-associated NOD2 allele, L1007insC. We further show that TRAF6 and NOD2/RIP2 share the same biochemical machinery (transforming growth factor ␤-activated kinase 1 [TAK1]/TAB/Ubc13) to activate NF-B, allowing TLR signaling and NOD2 signaling to synergistically augment cytokine release. These findings suggest a biochemical mechanism for the faulty cytokine balance seen in Crohn's disease.

RIP kinases: key decision makers in cell death and innate immunity

“…This results in ubiquitination of RIP2 followed by recruitment of the TAK1 and IKK complexes and downstream activation of NFkB and MAP kinase pathways by an analogous mechanism to that described above for RIP1 signalling in the TNFR-1 pathway. 148,[163][164][165] This results in the expression of a range of inflammatory proteins, anti-bacterial proteins, activation of autophagy and antigen presentation. [166][167][168] Notably, RIP2 is required to mediate all of the in vivo host responses to MDP.…”

The ubiquitin system: a critical regulator of innate immunity and pathogen–host interactions

“…In addition, NOD2 activation leads to NEMO ubiquitination, which was shown to amplify NF-κB signaling and is partially accomplished by NOD2-activated TRAF6. 18 Finally, a recent study suggests that the ubiquitin molecule might compete with RIP2 to associate with CARD of NOD1/2, thereby negatively affecting NOD1/2 signaling. 19 Intriguingly, XIAP (X-linked Inhibitor of Apoptosis) was also recently observed to be present in the NOD2 signaling complex.…”

LRRC19 expressed in the kidney induces TRAF2/6-mediated signals to prevent infection by uropathogenic bacteria

“…16). Although proteins are frequently targeted to the proteosome for degradation by conjugation with K48-linked ubiquitin chains, K63-linked ubiquitin is a common activating signal 29,30 . Thus, to determine the type of ubiquitin on TRAF6 that was affected by LRRC19, we transfected HEK293T cells to express TRAF6 with LRRC19 in the presence of vector encoding total ubiquitin, K48-linked ubiquitin or K63-linked ubiquitin.…”