OTUB1 prevents lethal hepatocyte necroptosis through stabilization of c-IAP1 during murine liver inflammation

Abstract

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.

The TWEAK/Fn14/CD163 axis—implications for metabolic disease

“…In their absence TWEAK signaling and gene expression are greatly diminished [52]. OTUB1 works by inhibiting cytokine gene transcription within the immune system and regulates c-IAP1 via K48-linked polyubiquitination [53]. When OTUB1 is downregulated, it leads to much faster degradation of c-IAP1 and as a consequence, weaker TWEAK signaling [52,53].…”

“…OTUB1 works by inhibiting cytokine gene transcription within the immune system and regulates c-IAP1 via K48-linked polyubiquitination [53]. When OTUB1 is downregulated, it leads to much faster degradation of c-IAP1 and as a consequence, weaker TWEAK signaling [52,53]. Interestingly, only canonical NF-κB and MAPK signaling are affected by the downregulation of OTUB1, which temporarily (as the non-canonical pathway is still functional) lowers the volume of produced cytokines including TNFα [32,52].…”

Necroptosis contributes to chronic inflammation and fibrosis in aging liver

“…We found that hepatocytes isolated from old mice exhibit increased necroptosis relative to hepatocytes from young mice, suggesting that necroptotic hepatocytes might be a major source of DAMPs in aging liver. Hepatocyte necroptosis has previously been reported in bacterial hepatitis and TNF challenge (Koschel et al, 2021), ischemia and reperfusion injury (Zhong et al, 2020), chronic alcoholic liver disease (Lu et al, 2016), nonalcoholic fatty liver disease (Afonso et al, 2015;Wu & Nagy, 2020) and from hepatic O-GlcNAc transferase deficiency in mice (Zhang et al, 2019). Similar to hepatocytes, liver macrophages in old mice also expressed necroptosis markers.…”

OTUD1 deubiquitylase regulates NF-κB- and KEAP1-mediated inflammatory responses and reactive oxygen species-associated cell death pathways

“…The ubiquitylation and deubiquitylation of RIP1/RIP3 regulate the NF-κB activation, apoptotic, and necroptotic pathways 52 , and DUBs such as CYLD 53 , A20 54 , and OTUB1 55 are involved in the regulation of the K63 ubiquitylation of RIP1/RIP3. In this study, we showed that the genetic deficiency of OTUD1 accelerated the TNF-α-induced apoptotic and necroptotic cell death pathways, and the basal K63 ubiquitylation of RIP3 was enhanced in Otud1 -/--MEFs (Figs.…”