title: Receptor-Interacting Protein Kinase-2 Inhibition by CYLD Impairs Antibacterial Immune Responses in Macrophages.
abstract: Upon infection with intracellular bacteria, nucleotide oligomerization domain protein 2 recognizes bacterial muramyl dipeptide and binds, subsequently, to receptor-interacting serine/threonine kinase 2 (RIPK2), which activates immune responses via the nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB) and extracellular signal-regulated kinase (ERK) pathways. Activation of RIPK2 depends on its K63 ubiquitination by E3 ligases, whereas the deubiquitinating enzyme A20 counter regulates RIPK2 activity by cleaving K63-polyubiquitin chains from RIPK2. Here, we newly identify the deubiquitinating enzyme CYLD as a new inhibitor of RIPK2. We show that CYLD binds to and removes K63-polyubiquitin chains from RIPK2 in Listeria monocytogenes (Lm) infected murine bone marrow-derived macrophages. CYLD-mediated K63 deubiquitination of RIPK2 resulted in an impaired activation of both NF-κB and ERK1/2 pathways, reduced production of proinflammatory cytokines interleukin-6 (IL-6), IL-12, anti-listerial reactive oxygen species (ROS) and nitric oxide (NO), and, finally, impaired pathogen control. In turn, RIPK2 inhibition by siRNA prevented activation of NF-κB and ERK1/2 and completely abolished the protective effect of CYLD deficiency with respect to the production of IL-6, NO, ROS, and pathogen control. Noteworthy, CYLD also inhibited autophagy of Listeria in a RIPK2-ERK1/2-dependent manner. The protective function of CYLD deficiency was dependent on interferon gamma (IFN-γ) prestimulation of infected macrophages. Interestingly, the reduced NF-κB activation in CYLD-expressing macrophages limited the protective effect of IFN-γ by reducing NF-κB-dependent signal transducers and activators of transcription-1 (STAT1) activation. Taken together, our study identifies CYLD as an important inhibitor of RIPK2-dependent antibacterial immune responses in macrophages.
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg , Magdeburg , Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Wex
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg , Magdeburg , Germany.
authors.forename: Ursula
authors.initials: U
authors.lastname: Schmid
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg , Magdeburg , Germany.
authors.forename: Sissy
authors.initials: S
authors.lastname: Just
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg , Magdeburg , Germany.
authors.forename: Xu
authors.initials: X
authors.lastname: Wang
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg , Magdeburg , Germany.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Wurm
authors.affiliation: Institute of Experimental Internal Medicine, Otto-von-Guericke University Magdeburg , Magdeburg , Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Naumann
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg, Magdeburg, Germany; Organ-Specific Immune Regulation, Helmholtz Centre for Infection Research, Braunschweig, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Schlüter
authors.affiliation: Institute of Medical Microbiology and Hospital Hygiene, Otto-von-Guericke University Magdeburg, Magdeburg, Germany; Organ-Specific Immune Regulation, Helmholtz Centre for Infection Research, Braunschweig, Germany.
authors.forename: Gopala
authors.initials: G
authors.lastname: Nishanth
keywords: CYLD; Listeria monocytogenes; antibacterial immune responses; macrophages; receptor-interacting protein kinase-2
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