90K Glycoprotein Promotes Degradation of Mutant β-Catenin Lacking the ISGylation or Phosphorylation Sites in the N-terminus

Abstract

β-Catenin is a major transducer of the Wnt signaling pathway, which is aberrantly expressed in colorectal and other cancers. Previously, we showed that β-catenin is downregulated by the 90K glycoprotein via ISGylation-dependent degradation. However, the further mechanisms of β-catenin degradation by 90K-mediated ISGylation pathway were not investigated. This study aimed to identify the β-catenin domain responsible for the action of 90K and to compare the mechanism of 90K on β-catenin degradation with phosphorylation-dependent ubiquitinational degradation of β-catenin. The deletion mutants of β-catenin lacking N- or C-terminal domain or mutating the N-terminal lysine or nonlysine residue were employed to delineate the characteristics of β-catenin degradation by 90K-mediated ISGylation pathway. 90K induced Herc5 and ISG15 expression and reduced β-catenin levels in HeLa and CSC221 cells. The N-terminus of β-catenin is required for 90K-induced β-catenin degradation, but the N-terminus of β-catenin is not essential for interaction with Herc5. However, substituting lysine residues in the N-terminus of β-catenin with arginine or deleting serine or threonine residue containing domains from the N-terminus does not affect 90K-induced β-catenin degradation, indicating that the N-terminal 86 amino acids of β-catenin are crucial for 90K-mediated ISGylation/degradation of β-catenin in which the responsible lysine or nonlysine residues were not identified. Our present results highlight the action of 90K on promoting degradation of mutant β-catenin lacking the phosphorylation sites in the N-terminus. It provides further insights into the discrete pathway downregulating the stabilized β-catenin via acquiring mutations at the serine/threonine residues in the N-terminus.

Covalent conjugation of ubiquitin-like ISG15 to apoptosis-inducing factor exacerbates toxic stimuli-induced apoptotic cell death

“…Although ubiquitination mainly targets the lysine residues, numerous studies have reported the non-lysine-linked ubiquitination, occurring through non-amine groups, such as sulfhydryl groups of cysteine or hydroxyl groups of threonine and serine residues [46][47][48][49][50]. Supporting to our data, there was a report suggesting that "non-canonical" conjugation of ISG15 to its target protein could also be possible [51]. In this case the target protein has no cysteine residue except for lysine residues, and all serine and threonine residues which widely reported to be alternative conjugation sites in non-canonical ubiquitination studies were substituted for the assay of ISG15-conjugation.…”

Role of galectin 3 binding protein in cancer progression: a potential novel therapeutic target

“…Finally, in a successive work, the same research group aimed to identify the β-catenin domain responsible of this mechanism. Results from this study demonstrated that LGALS3BP promotes degradation of mutant β-catenins lacking the ISGylation or phosphorylation sites [ 78 ]. Role of LGALS3BP in inducing β-catenin degradation was also confirmed in a study from Pikkarainen et al [ 79 ].…”

The Functional Roles of ISG15/ISGylation in Cancer

“…On the other hand, Yoo et al demonstrated that ISGylation promotes the E3 ubiquitin ligase activity of Hsp70-interacting protein (CHIP), which subsequently reduces the level of oncogenic c-Myc, one of its many ubiquitination targets, and inhibits A549 cells and tumor growth [ 76 ]. Furthermore, CYP1B1 can inhibit the expression and covalent modification of ISG15 in the Hela cell line, preventing β-catenin degradation by ISG15 modification and activating the Wnt/β-catenin signaling pathway, resulting in abnormal cell proliferation and differentiation [ 77 ]. In lung cancer model mice, Mustachi et al found that the elevated level of ISGylation could inhibit tumor growth by upregulating autophagy [ 78 ].…”