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Research Articles: Therapeutics, Targets, and Development

UBE1L represses PML/RAR by targeting the PML domain for ISG15ylation

Departments of ¹ Pharmacology and Toxicology and ² Medicine and ³ Norris Cotton Cancer Center, Dartmouth Medical School, Hanover, New Hampshire and Dartmouth-Hitchcock Medical Center, Lebanon, New Hampshire

Requests for reprints: Ethan Dmitrovsky, Department of Pharmacology and Toxicology, Dartmouth Medical School, 7650 Remsen Building, Hanover, New Hampshire 03755. Phone: 603-650-1667; Fax: 603-650-1129. E-mail: Ethan.Dmitrovsky{at}Dartmouth.edu

Abstract

Acute promyelocytic leukemia (APL) is characterized by expression of promyelocytic leukemia (PML)/retinoic acid (RA) receptor (RAR) protein and all-trans-RA-mediated clinical remissions. RA treatment can confer PML/RAR degradation, overcoming dominant-negative effects of this oncogenic protein. The present study uncovered independent retinoid degradation mechanisms, targeting different domains of PML/RAR. RA treatment is known to repress PML/RAR and augment ubiquitin-activating enzyme-E1-like (UBE1L) protein expression in NB4-S1 APL cells. We previously reported RA-induced UBE1L and the IFN-stimulated gene, 15-kDa protein ISG15ylation in APL cells. Whether the ubiquitin-like protein ISG15 directly conjugates with PML/RAR was not explored previously and is examined in this study. Transient transfection experiments with different PML/RAR domains revealed that RA treatment preferentially down-regulated the RAR domain, whereas UBE1L targeted the PML domain for repression. As expected, ubiquitin-specific protease 18 (UBP43/USP18), the ISG15 deconjugase, opposed UBE1L but not RA-dependent PML/RAR degradation. In contrast, the proteasomal inhibitor, N-acetyl-leucinyl-leucinyl-norleucinal, inhibited both UBE1L- and RA-mediated PML/RAR degradation. Notably, UBE1L induced ISG15ylation of the PML domain of PML/RAR, causing its repression. These findings confirmed that RA triggers PML/RAR degradation through different domains and distinct mechanisms. Taken together, these findings advance prior work by establishing two pathways converge on the same oncogenic protein to cause its degradation and thereby promote antineoplastic effects. The molecular pharmacologic implications of these findings are discussed. [Mol Cancer Ther 2008;7(4):905–14]

Grant support: NIH, National Cancer Institute grants R01-CA087546, R01-CA062275, and R01-CA111422 (E. Dmitrovsky); Samuel Waxman Foundation Cancer Research Award (E. Dmitrovsky); American Cancer Society Institutional Grant (S.J. Freemantle); and NIH grant T32-CA00959 (S. Blumen).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

⁴ Supplementary material for this article is available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

Received 7/31/07; revised 1/18/08; accepted 2/21/08.