title: Inhibition of the transcription factor Yin Yang 1 activity by S-nitrosation.
abstract: Treatment of several prostate cancer (CaP) cell lines (PC-3, CL-1, and DU-145) with the nitric oxide (NO) donor DETA/NONOate upregulated Fas expression and sensitized the CaP cells to the Fas ligand CH-11 agonist monoclonal antibody-induced apoptosis. Previous findings demonstrated that the transcription repressor Yin Yang 1 (YY1), which is inhibited by NO, negatively regulates Fas transcription [H.J. Garban, B. Bonavida, Nitric oxide inhibits the transcription repressor Yin-Yang 1 binding activity at the silencer region of the Fas promoter: a pivotal role for nitric oxide in the upregulation of Fas gene expression in human tumor cells, J. Immunol. 167 (2001) 75-81]. YY1 is a zinc finger protein and thus, we hypothesized that NO inhibits YY1 activity via S-nitrosation of critical cysteines residues coordinated by Zn2+. Treatment of PC-3 cells with DETA/NONOate inhibited the constitutive DNA-binding activity of YY1 as assessed by EMSA. Further, treatment with DETA/NONOate resulted in S-nitrosation of YY1 as detected by two different methods. The DAN-based method examined NO-treated tumor-derived cell lysates that were immunoprecipitated with an anti-YY1 specific antibody and the NO released was determined quantitatively by fluorometry. The second method consisted of immunoprecipitation of the tumor cell lysates by an anti-SNO cysteine antibody and the immunoprecipitate was immunoblotted with anti-YY1 antibody. Both methods revealed significant S-nitrosation of YY1 by DETA/NONOate treatment over control untreated cells. The S-nitrosation of YY1 was further corroborated by immunohistochemistry using dual color immunofluorescence. The direct role of YY1 in the negative regulation of Fas expression was demonstrated by transfection of cells with siRNA YY1. The transfectants exhibited upregulation of Fas expression in the absence of treatment with DETA/NONOate and were sensitized to CH-11-induced apoptosis. Altogether, these findings reveal that NO inhibits YY1 DNA-binding activity through S-nitrosation and consequently results in upregulation of Fas expression and tumor cell sensitization to Fas-induced apoptosis.
authors.affiliation: Department of Microbiology, Immunology and Molecular Genetics, David Geffen School of Medicine and Jonsson Comprehensive Cancer Center at the University of California, Los Angeles, USA.
authors.forename: Fumiya
authors.initials: F
authors.lastname: Hongo
authors.forename: Hermes
authors.initials: H
authors.lastname: Garban
authors.forename: Sara
authors.initials: S
authors.lastname: Huerta-Yepez
authors.forename: Mario
authors.initials: M
authors.lastname: Vega
authors.forename: Ali R
authors.initials: AR
authors.lastname: Jazirehi
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Mizutani
authors.forename: Tsuneharu
authors.initials: T
authors.lastname: Miki
authors.forename: Benjamin
authors.initials: B
authors.lastname: Bonavida
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects; D050981:Erythroid-Specific DNA-Binding Factors; D006801:Humans; D008297:Male; D009569:Nitric Oxide / Q000378:metabolism*; D015538:Nitrosation / Q000187:drug effects; D009603:Nitroso Compounds / Q000008:administration & dosage*; D011471:Prostatic Neoplasms / Q000378:metabolism*; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D050986:YY1 Transcription Factor; D019014:fas Receptor
chemical_list: D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; C491901:FAS protein, human; D009603:Nitroso Compounds; D018124:Receptors, Tumor Necrosis Factor; D014157:Transcription Factors; D050986:YY1 Transcription Factor; C494698:YY1 protein, human; D019014:fas Receptor; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; D009569:Nitric Oxide; D004247:DNA