ISG15 and ISGylation in Human Diseases

Abstract

Type I Interferons (IFNs) induce the expression of >500 genes, which are collectively called ISGs (IFN-stimulated genes). One of the earliest ISGs induced by IFNs is ISG15 (Interferon-Stimulated Gene 15). Free ISG15 protein synthesized from the ISG15 gene is post-translationally conjugated to cellular proteins and is also secreted by cells into the extracellular milieu. ISG15 comprises two ubiquitin-like domains (UBL1 and UBL2), each of which bears a striking similarity to ubiquitin, accounting for its earlier name ubiquitin cross-reactive protein (UCRP). Like ubiquitin, ISG15 harbors a characteristic β-grasp fold in both UBL domains. UBL2 domain has a conserved C-terminal Gly-Gly motif through which cellular proteins are appended via an enzymatic cascade similar to ubiquitylation called ISGylation. ISG15 protein is minimally expressed under physiological conditions. However, its IFN-dependent expression is aberrantly elevated or compromised in various human diseases, including multiple types of cancer, neurodegenerative disorders (Ataxia Telangiectasia and Amyotrophic Lateral Sclerosis), inflammatory diseases (Mendelian Susceptibility to Mycobacterial Disease (MSMD), bacteriopathy and viropathy), and in the lumbar spinal cords of veterans exposed to Traumatic Brain Injury (TBI). ISG15 and ISGylation have both inhibitory and/or stimulatory roles in the etiology and pathogenesis of human diseases. Thus, ISG15 is considered a "double-edged sword" for human diseases in which its expression is elevated. Because of the roles of ISG15 and ISGylation in cancer cell proliferation, migration, and metastasis, conferring anti-cancer drug sensitivity to tumor cells, and its elevated expression in cancer, neurodegenerative disorders, and veterans exposed to TBI, both ISG15 and ISGylation are now considered diagnostic/prognostic biomarkers and therapeutic targets for these ailments. In the current review, we shall cover the exciting journey of ISG15, spanning three decades from the bench to the bedside.

Keywords: ISG15; ISG15-deficient inflammatory diseases; ISGylation; cancer; neurodegenerative diseases; ubiquitin.

Figure 1

Processing of ISG15 and mechanism of ISG15 conjugation. (A) The schematic depicts that a 17 kDa precursor (Pro-ISG15) contains two ubiquitin-like domains joined by a flexible polypeptide hinge [69]. An ISG15-specific isopeptidase Ubp1-related protein exposes the C-terminal Gly-Gly motif necessary for conjugation to cellular proteins [87]. (B) E1 activating enzyme (Uba7/UBE1L) forms a high-energy thioester intermediate with ISG15 in an ATP-dependent manner (Step 1) [67,91]. E1 catalyzes the transfer of ISG15 thioester intermediate to the ISG15-specific conjugating enzyme E2 (UBCH8) to form UBCH8~ISG15 thioester (Step 2) [68]. ISG15-specific E3 ligase (HERC5, TRIM25, or ARIH1) binds the UBCH8~ISG15 thioester (Step 3) and catalyzes the aminolytic cleavage of UBCH8~ISG15 followed by the formation of an isopeptide bond with ε-NH2 of the Lys residue of the target protein (Step 4) [66,92,95]. UBP43 (USP18) catalyzes the process of deISGylation (Step 5) [88].

Figure 2

Functions of free and conjugated ISG15. (A) Extracellular free ISG15 serves as an immunomodulatory cytokine secreted in response to type I interferon induction [63,64]. ISG15 stimulates the production of IFN-γ in bovine peripheral blood mononuclear cells [60,106], T lymphocytes, and CD3⁺ T cells [63,107]. ISG15 stimulates the proliferation of CD56⁺ natural killer cells in the presence of CD3⁺ T cells, which leads to an enhanced lymphokine-activated non-major histocompatibility complex-restricted target cell lysis [63]. The binding of ISG15 to the LFA1 receptor on NK cells stimulates the release of IFN-γ and IL-10 from IL-12 primed NK cells [80]. Secreted ISG15 can also influence neutrophil chemotaxis [108]. (B) Free ISG15 protein synthesized from the ISG15 gene is found in cells in its free and conjugated intracellular forms. Intracellular free ISG15 stabilizes (e.g., USP18) or destabilizes (e.g., Cyclin D1) some cellular proteins [72,109]. Intracellular ISG15 conjugates to a multitude of targets within the cell upon interferon stimulation. These protein targets are involved in every facet of cellular function, including DNA replication/repair, metabolism, signal transduction, and cytoskeletal organization, among several others [73,110,111,112,113]. The majority of the nuclear targets for ISG15 are involved in chromatin remodeling/RNA polymerase II transcription or RNA processing [114]. Protein ISGylation inhibits proteasome-mediated protein degradation and increases protein degradation by selective autophagy [70,76,115,116,117]. ISGylation confers topoisomerase-targeted drug sensitivity to tumor cells [74]. The consequences of ISGylation for ubiquitin-mediated protein degradation of substrates are variable and may be context-specific.