OTULIN prevents liver inflammation and hepatocellular carcinoma by inhibiting FADD- and RIPK1 kinase-mediated hepatocyte apoptosis

Abstract

Inflammatory signaling pathways are tightly regulated to avoid chronic inflammation and the development of inflammatory pathologies. OTULIN is a deubiquitinating enzyme that specifically cleaves linear ubiquitin chains generated by the linear ubiquitin chain assembly complex (LUBAC), and OTULIN deficiency causes OTULIN-related autoinflammatory syndrome (ORAS) in humans. OTULIN was shown to negatively control NF-κB signaling in response to various stimuli, but also to protect cells from tumor necrosis factor (TNF)-induced apoptosis. To investigate the importance of OTULIN in liver homeostasis and pathology, we developed a novel mouse line specifically lacking OTULIN in liver parenchymal cells. These mice spontaneously develop a severe liver disease, characterized by liver inflammation, hepatocyte apoptosis and compensatory hepatocyte proliferation, leading to steatohepatitis, fibrosis and hepatocellular carcinoma (HCC). Genetic ablation of Fas-associated death domain (FADD) completely rescues the severe liver pathology, and knock-in expression of kinase inactive receptor-interacting protein kinase 1 (RIPK1) significantly protects from developing liver disease, demonstrating that death receptor-mediated apoptosis of OTULIN-deficient hepatocytes triggers disease pathogenesis in this model. Finally, we demonstrate that type I interferons contribute to disease pathogenesis in hepatocyte-specific OTULIN deficient mice. Together, our study reveals the critical importance of OTULIN in protecting hepatocytes from death, and thereby avoid development of chronic liver inflammation and HCC in mice.

Ubiquitin-dependent and -independent functions of OTULIN in cell fate control and beyond

“…Surprisingly, OTULIN deficiency in non-hematopoietic cells resulted in severe liver pathology in mice [25]. Hepatocyte-specific OTULIN knock-out mice develop chronic inflammatory liver disease with premalignant alterations, ultimately leading to hepatocellular carcinoma [25,68]. Loss of OTULIN expression in mouse hepatocytes increases cellular Met1 Ub levels, decreases LUBAC levels and does not induce NF-κB hyper-signaling, but sensitizes to apoptosis [25,68], in line with previous reports [7,23].…”

“…Hepatocyte-specific OTULIN knock-out mice develop chronic inflammatory liver disease with premalignant alterations, ultimately leading to hepatocellular carcinoma [25,68]. Loss of OTULIN expression in mouse hepatocytes increases cellular Met1 Ub levels, decreases LUBAC levels and does not induce NF-κB hyper-signaling, but sensitizes to apoptosis [25,68], in line with previous reports [7,23]. Importantly, co-deletion of TNFR1 in OTULIN-deficient mice does not ameliorate liver pathology, suggesting TNFαindependent mechanisms [25,68].…”

“…Loss of OTULIN expression in mouse hepatocytes increases cellular Met1 Ub levels, decreases LUBAC levels and does not induce NF-κB hyper-signaling, but sensitizes to apoptosis [25,68], in line with previous reports [7,23]. Importantly, co-deletion of TNFR1 in OTULIN-deficient mice does not ameliorate liver pathology, suggesting TNFαindependent mechanisms [25,68]. This is in contrast with dysregulated TNFR1 signaling commonly observed in liver disease and cancer and the fact that liver-specific deletion of CYLD induces TNFR1-mediated liver inflammation and development of hepatocellular carcinoma [69,70].…”

The Balance of TNF Mediated Pathways Regulates Inflammatory Cell Death Signaling in Healthy and Diseased Tissues

“…However, the lack of protection with RIP3 loss alone and the extensive cleaved caspase-3 labeling suggests that apoptosis is the primary driver of the pro-inflammatory phenotype in these mice. In line with these observations in systemic, inducible Otulin or Birc2/3 knock-out mice, hepatocyte specific deletion of Otulin results in hepatocyte apoptosis with resultant compensatory hyperplasia and inflammation that can progress to hepatocellular carcinoma ( Damgaard et al, 2020 ; Verboom et al, 2020 ). Increased cell death and steatosis is evident in these mice by postnatal day 9.…”

“…Interestingly, steatosis and increased liver enzymes were also identified in an OTULIN deficient patient ( Damgaard et al, 2020 ). This hepatic injury can be alleviated by the loss of RIP1 kinase activity due to expression of the RIP1 D138N kinase dead protein, and more completely rescued by hepatocyte-specific Fadd deletion, indicating that the injury is driven by apoptosis signaling ( Verboom et al, 2020 ), although loss of TNFR1 is not sufficient to protect against liver pathology in these mice ( Damgaard et al, 2020 ). mTOR signaling is also increased in livers with hepatocyte-specific OTULIN deficiency.…”