In bacterial and sterile inflammation of the liver, hepatocyte apoptosis is, in contrast to necroptosis, a common feature. The molecular mechanisms preventing hepatocyte necroptosis and the potential consequences of hepatocyte necroptosis are largely unknown. Apoptosis and necroptosis are critically regulated by the ubiquitination of signaling molecules but especially the regulatory function of deubiquitinating enzymes (DUBs) is imperfectly defined. Here, we addressed the role of the DUB OTU domain aldehyde binding-1 (OTUB1) in hepatocyte cell death upon both infection with the hepatocyte-infecting bacterium Listeria monocytogenes (Lm) and D-Galactosamine (DGal)/Tumor necrosis factor (TNF)-induced sterile inflammation. Combined in vivo and in vitro experiments comprising mice lacking OTUB1 specifically in liver parenchymal cells (OTUB1LPC-KO) and human OTUB1-deficient HepG2 cells revealed that OTUB1 prevented hepatocyte necroptosis but not apoptosis upon infection with Lm and DGal/TNF challenge. Lm-induced necroptosis in OTUB1LPC-KO mice resulted in increased alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) release and rapid lethality. Treatment with the receptor-interacting serine/threonine-protein kinase (RIPK) 1 inhibitor necrostatin-1s and deletion of the pseudokinase mixed lineage kinase domain-like protein (MLKL) prevented liver damage and death of infected OTUB1LPC-KO mice. Mechanistically, OTUB1 reduced K48-linked polyubiquitination of the cellular inhibitor of apoptosis 1 (c-IAP1), thereby diminishing its degradation. In the absence of OTUB1, c-IAP1 degradation resulted in reduced K63-linked polyubiquitination and increased phosphorylation of RIPK1, RIPK1/RIPK3 necrosome formation, MLKL-phosphorylation and hepatocyte death. Additionally, OTUB1-deficiency induced RIPK1-dependent extracellular-signal-regulated kinase (ERK) activation and TNF production in Lm-infected hepatocytes. Collectively, these findings identify OTUB1 as a novel regulator of hepatocyte-intrinsic necroptosis and a critical factor for survival of bacterial hepatitis and TNF challenge.
Cerebral malaria is a severe complication of human malaria and may lead to death of Plasmodium falciparum-infected individuals. Cerebral malaria is associated with sequestration of parasitized red blood cells within the cerebral microvasculature resulting in damage of the blood–brain barrier and brain pathology. Although CD8+ T cells have been implicated in the development of murine experimental cerebral malaria (ECM), several other studies have shown that CD8+ T cells confer protection against blood-stage infections. Since the role of host deubiquitinating enzymes (DUBs) in malaria is yet unknown, we investigated how the DUB cylindromatosis (CYLD), an important inhibitor of several cellular signaling pathways, influences the outcome of ECM. Upon infection with Plasmodium berghei ANKA (PbA) sporozoites or PbA-infected red blood cells, at least 90% of Cyld−/− mice survived the infection, whereas all congenic C57BL/6 mice displayed signatures of ECM, impaired parasite control, and disruption of the blood–brain barrier integrity. Cyld deficiency prevented brain pathology, including hemorrhagic lesions, enhanced activation of astrocytes and microglia, infiltration of CD8+ T cells, and apoptosis of endothelial cells. Furthermore, PbA-specific CD8+ T cell responses were augmented in the blood of Cyld−/− mice with increased production of interferon-γ and granzyme B and elevated activation of protein kinase C-θ and nuclear factor “kappa light-chain enhancer” of activated B cells. Importantly, accumulation of CD8+ T cells in the brain of Cyld−/− mice was significantly reduced compared to C57BL/6 mice. Bone marrow chimera experiments showed that the absence of ECM signatures in infected Cyld−/− mice could be attributed to hematopoietic and radioresistant parenchymal cells, most likely endothelial cells that did not undergo apoptosis. Together, we were able to show that host deubiqutinating enzymes play an important role in ECM and that CYLD promotes ECM supporting it as a potential therapeutic target for adjunct therapy to prevent cerebral complications of severe malaria.
Sublethal Lm infection deregulates hepatic drug metabolizing pathways. This finding indicates the need to monitor drug administration along Lm infections, especially in all patients needing constant medication.
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