*3.1. OTULIN*

OTULIN (also called FAM105B and Gumby) is an OTU-family DUB with the catalytic Cys129 residue (Figure 3). OTULIN exclusively hydrolyzes the M1-linked peptide bond between ubiquitins, but none of the K-linked isopeptide linkages [85–87]. Therefore, OTULIN downregulates the LUBAC-mediated innate immune responses. Interestingly, OTULIN binds to the N-terminal PUB domain of HOIP through the PUB domain-interacting motif (PIM), and the phosphorylation of Tyr56 in the PIM of OTULIN abrogated the HOIP binding [25,88,89]. In humans, genetic mutations in *OTULIN* cause multiple symptoms, such as recurrent fevers, autoantibodies, diarrhea, panniculitis, and arthritis, which are collectively referred to as OTULIN-related autoinflammatory syndrome (ORAS). These mutations deregulate the LUBAC-mediated linear ubiquitination signal [90–92], and the *OTULIN*-deficiency causes cell-type-specific LUBAC degradation [85,93]. Interestingly, knock-in mice expressing the active site mutant of Otulin(C129A) showed enhanced cell death through apoptosis and necroptosis, and the increased production of type I IFN, due to the reduced LUBAC activity [94]. Therefore, under physiological conditions, OTULIN prevents the auto-linear ubiquitination of LUBAC, and functions to maintain the LUBAC activity. During cell death, OTULIN regulates the linear ubiquitination of RIP1, which is cleaved at Asp31 by caspase 3. Moreover, the phosphorylation of Tyr56 in OTULIN is increased, and this is counteracted by the dual-specificity phosphatase 14 (DUSP14) during necroptosis [95]. The OTULIN-mediated suppression of hepatocyte apoptosis plays a crucial role in liver pathogeneses, such as hepatitis, fibrosis, and hepatocellular carcinoma [96]. TRIM32, a RING-type E3, conjugates the K63-linked ubiquitin chain to OTULIN to interfere with its interaction with HOIP in human embryonic kidney (HEK) 293 cells [97], and the interaction between TRIM32 and SNX27, which regulates endosome-to-plasma membrane trafficking, has been confirmed [98]. Thus, OTULIN is a crucial pathophysiological regulator in the linear ubiquitin code.
