A Discrete Ubiquitin-Mediated Network Regulates the Strength of NOD2 Signaling

Abstract

Dysregulation of NOD2 signaling is implicated in the pathology of various inflammatory diseases, including Crohn's disease, asthma, and sarcoidosis, making signaling proteins downstream of NOD2 potential therapeutic targets. Inhibitor-of-apoptosis (IAP) proteins, particularly cIAP1, are essential mediators of NOD2 signaling, and in this work, we describe a molecular mechanism for cIAP1's regulation in the NOD2 signaling pathway. While cIAP1 promotes RIP2's tyrosine phosphorylation and subsequent NOD2 signaling, this positive regulation is countered by another E3 ubiquitin ligase, ITCH, through direct ubiquitination of cIAP1. This ITCH-mediated ubiquitination leads to cIAP1's lysosomal degradation. Pharmacologic inhibition of cIAP1 expression in ITCH^−/− macrophages attenuates heightened ITCH^−/− macrophage muramyl dipeptide-induced responses. Transcriptome analysis, combined with pharmacologic inhibition of cIAP1, further defines specific pathways within the NOD2 signaling pathway that are targeted by cIAP1. This information provides genetic signatures that may be useful in repurposing cIAP1-targeted therapies to correct NOD2-hyperactive states and identifies a ubiquitin-regulated signaling network centered on ITCH and cIAP1 that controls the strength of NOD2 signaling.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.01049-12.

ACKNOWLEDGMENTS

We thank Sylvia Kertesy for technical assistance with mouse husbandry and members of the Abbott, Pizarro (CWRU), Ley (La Jolla Institute for Allergy & Immunology, La Jolla, CA), and Cominelli (CWRU) labs for helpful discussions in the course of their program project grant (P01DK091222) meetings. We thank Peter Scacheri (CWRU Department of Genetics) for helpful advice on the interpretation and presentation of the RNA-Seq analysis data.

This work was supported by NIH research grants R01GM86550-01 and P01DK091222, a Burroughs Wellcome Career Award for Biomedical Scientists (10061206.01 to D.W.A.), and an American Cancer Society postdoctoral fellowship (120209-PF-11-058-01-MPC to J.T.T.-A.). Bioinformatic support was made possible by a core utilization grant from the Clinical and Translational Science Collaborative of Cleveland to D.W.A. and J.T.T.-A., UL1TR000439 from the National Center for Advancing Translational Sciences component of the National Institutes of Health, and the NIH Roadmap for Medical Research.

Justine T. Tigno-Aranjuez designed, performed, and interpreted the experiments. Derek Abbott designed, performed, and helped to interpret the experiments. Xiaodong Bai performed the bioinformatic analysis of the transcriptome data. Justine T. Tigno-Aranjuez and Derek Abbott wrote the manuscript.

We have no conflict of interest.