title: The ISG15-specific protease USP18 regulates stability of PTEN.
abstract: The ubiquitin-like modifier interferon-stimulated gene 15 (ISG15) is implicated in both oncogenic and tumor suppressive programs. Yet, few ISGylation substrates are known and functionally validated in cancer biology. We previously found specific oncoproteins were substrates of ISGylation and were stabilized by the ISG15-specific deubiquitinase (DUB) ubiquitin specific peptidase 18 (USP18). Using reverse-phase protein arrays (RPPAs), this study reports that engineered loss of the DUB USP18 destabilized the tumor suppressor protein phosphatase and tensin homologue (PTEN) in both murine and human lung cancer cell lines. In contrast, engineered gain of USP18 expression in these same lung cancer cell lines stabilized PTEN protein. Using the protein synthesis inhibitor cycloheximide (CHX), USP18 knockdown was shown to destabilize PTEN whereas USP18 overexpression stabilized PTEN protein. Interestingly, repression of USP18 decreased cytoplasmic PTEN relative to nuclear PTEN protein levels. We sought to identify mechanisms engaged in this PTEN protein destabilization using immunoprecipitation assays and found ISG15 directly conjugated with PTEN. To confirm translational relevance of this work, USP18 and PTEN immunohistochemical expression were compared in comprehensive lung cancer arrays. There was a significant (P < 0.0001) positive correlation and association between PTEN and USP18 protein expression profiles in human lung cancers. Taken together, this study identified PTEN as a previously unrecognized substrate of the ISGylation post-translational modification pathway. The deconjugase USP18 serves as a novel regulator of PTEN stability. This indicates inhibition of ISGylation is therapeutically relevant in cancers.
authors.affiliation: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, NH, USA.
authors.forename: Lisa Maria
authors.initials: LM
authors.lastname: Mustachio
authors.affiliation: Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Masanori
authors.initials: M
authors.lastname: Kawakami
authors.affiliation: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, NH, USA.
authors.forename: Yun
authors.initials: Y
authors.lastname: Lu
authors.affiliation: Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Jaime
authors.initials: J
authors.lastname: Rodriguez-Canales
authors.affiliation: Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Barbara
authors.initials: B
authors.lastname: Mino
authors.affiliation: Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Carmen
authors.initials: C
authors.lastname: Behrens
authors.affiliation: Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Wistuba
authors.affiliation: Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Neus
authors.initials: N
authors.lastname: Bota-Rabassedas
authors.affiliation: Department of Biostatistics, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Yu
authors.affiliation: Department of Biostatistics, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: J Jack
authors.initials: JJ
authors.lastname: Lee
authors.affiliation: Melanoma Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.|Genomic Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Roszik
authors.affiliation: Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Lin
authors.initials: L
authors.lastname: Zheng
authors.affiliation: Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Xi
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, NH, USA.
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Freemantle
authors.affiliation: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, NH, USA.|Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.|Cancer Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
authors.forename: Ethan
authors.initials: E
authors.lastname: Dmitrovsky
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D016207:Cytokines / Q000235:genetics / Q000378:metabolism*; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D055786:Gene Knockout Techniques; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism; D051379:Mice; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D055550:Protein Stability; D043222:Ubiquitin Thiolesterase; D054875:Ubiquitination; D014452:Ubiquitins / Q000235:genetics / Q000378:metabolism*
chemical_list: D016207:Cytokines; D014452:Ubiquitins; C074034:ISG15 protein, human; D051059:PTEN Phosphohydrolase; D010450:Endopeptidases; C485560:USP18 protein, human; D043222:Ubiquitin Thiolesterase
keywords: ISG15; PTEN; USP18; and lung cancer; protein stability