title: A genome-wide small interfering RNA (siRNA) screen reveals nuclear factor-κB (NF-κB)-independent regulators of NOD2-induced interleukin-8 (IL-8) secretion.
abstract: NOD2 encodes an intracellular multidomain pattern recognition receptor that is the strongest known genetic risk factor in the pathogenesis of Crohn disease (CD), a chronic relapsing inflammatory disorder of the intestinal tract. NOD2 functions as a sensor for bacterial cell wall components and activates proinflammatory and antimicrobial signaling pathways. Here, using a genome-wide small interfering RNA (siRNA) screen, we identify numerous genes that regulate secretion of the proinflammatory cytokine IL-8 in response to NOD2 activation. Moreover, many of the identified IL-8 regulators are linked by protein-protein interactions, revealing subnetworks of highly connected IL-8 regulators implicated in processes such as vesicle formation, mRNA stability, and protein ubiquitination and trafficking. A TNFα counterscreen to induce IL-8 secretion in an NOD2-independent manner reveals that the majority of the identified regulators affect IL-8 secretion irrespective of the initiating stimuli. Using immortalized macrophages, we validate the ubiquitin protease, USP8, and the endosomal sorting protein, VPS28, as negative regulators of NOD2-induced cytokine secretion. Interestingly, several genes that affect NOD2-induced IL-8 secretion are present in loci associated with CD risk by genome-wide association studies, supporting a role for the NOD2/IL-8 pathway, and not just NOD2, in the pathogenesis of CD. Overall, this screen provides a valuable resource in the advancement of our understanding of the genes that regulate the secretion of IL-8.
authors.affiliation: From the Department of Pathology and.
authors.forename: Neil
authors.initials: N
authors.lastname: Warner
authors.affiliation: From the Department of Pathology and.
authors.forename: Aaron
authors.initials: A
authors.lastname: Burberry
authors.affiliation: the Comprehensive Cancer Center, University of Michigan, Ann Arbor, Michigan 48109 and.
authors.forename: Maria
authors.initials: M
authors.lastname: Pliakas
authors.affiliation: the Department of Pathobiology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195.
authors.forename: Christine
authors.initials: C
authors.lastname: McDonald
authors.affiliation: From the Department of Pathology and the Comprehensive Cancer Center, University of Michigan, Ann Arbor, Michigan 48109 and gabriel.nunez@umich.edu.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Núñez
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000378:metabolism / Q000473:pathology; D002461:Cell Line, Transformed; D003424:Crohn Disease / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010450:Endopeptidases / Q000235:genetics* / Q000378:metabolism; D056827:Endosomal Sorting Complexes Required for Transport / Q000235:genetics* / Q000378:metabolism; D056426:Genetic Loci; D055106:Genome-Wide Association Study; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D016209:Interleukin-8 / Q000819:agonists / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D053473:Nod2 Signaling Adaptor Protein / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D025941:Protein Interaction Mapping; D021381:Protein Transport; D020871:RNA Stability; D034741:RNA, Small Interfering / Q000235:genetics* / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism; D043222:Ubiquitin Thiolesterase / Q000235:genetics* / Q000378:metabolism; D054875:Ubiquitination
chemical_list: D056827:Endosomal Sorting Complexes Required for Transport; D016209:Interleukin-8; D016328:NF-kappa B; C429018:NOD2 protein, human; D053473:Nod2 Signaling Adaptor Protein; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; C476409:VPS28 protein, human; D010450:Endopeptidases; C541534:USP8 protein, human; D043222:Ubiquitin Thiolesterase
keywords: Cytokine; IL-8; Inflammation; Inflammatory Bowel Disease (IBD); JAK2; NOD-like Receptor (NLR); NOD2; Secretion; USP8