title: Characterization of the protein ubiquitination response induced by Doxorubicin.
abstract: Doxorubicin is commonly considered to exert its anti-tumor activity by triggering apoptosis of cancer cells through DNA damage. Recent reports have shown that Doxorubicin elicits a marked heat shock response, and that either inhibition or silencing of heat shock proteins enhance the Doxorubicin apoptotic effect in neuroblastoma cells. In order to investigate whether Doxorubicin may also act through protein modification, we performed a proteomic analysis of ubiquitinated proteins. Here we show that nanomolar Doxorubicin treatment of neuroblastoma cells caused: (a) dose-dependent over-ubiquitination of a specific set of proteins in the absence of measurable inhibition of proteasome, (b) protein ubiquination patterns similar to those with Bortezomib, a proteasome inhibitor, (c) depletion and loss of activity of ubiquitinated enzymes such as lactate dehydrogenase and α-enolase, and (d) a decrease in HSP27 solubility, probably a consequence of its binding to denatured proteins. These data strongly reinforce the hypothesis that Doxorubicin may also exert its effect by damaging proteins.
authors.affiliation: Department of Genetics, Biology and Biochemistry, University of Turin, Turin, Italy. giorgia.mandili@unito.it
authors.forename: Giorgia
authors.initials: G
authors.lastname: Mandili
authors.forename: Amina
authors.initials: A
authors.lastname: Khadjavi
authors.forename: Valentina
authors.initials: V
authors.lastname: Gallo
authors.forename: Valerio G
authors.initials: VG
authors.lastname: Minero
authors.forename: Luca
authors.initials: L
authors.lastname: Bessone
authors.forename: Franco
authors.initials: F
authors.lastname: Carta
authors.forename: Giuliana
authors.initials: G
authors.lastname: Giribaldi
authors.forename: Francesco
authors.initials: F
authors.lastname: Turrini
mesh_terms: D015153:Blotting, Western; D001897:Boronic Acids / Q000494:pharmacology; D000069286:Bortezomib; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000494:pharmacology*; D055551:HSP27 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D011719:Pyrazines / Q000494:pharmacology; D054875:Ubiquitination / Q000187:drug effects*
chemical_list: D001897:Boronic Acids; D055551:HSP27 Heat-Shock Proteins; D011719:Pyrazines; D000069286:Bortezomib; D004317:Doxorubicin; D007770:L-Lactate Dehydrogenase; D010751:Phosphopyruvate Hydratase