15-Deoxy-Delta12,14-Prostaglandin J2 Inhibits Tanscriptional Activity of Estrogen Receptor-Alpha via Covalent Modification of DNA-Binding Domain
The cyclopentenone 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)) inhibits proliferation of cancer cells, including breast cancers, by peroxisome proliferator-activated receptor-gamma (PPARgamma)-dependent and PPARgamma-independent mechanisms. However, little is known about its effect on the transcriptional activity of estrogen receptor-alpha (ERalpha) that plays vital roles in the growth of breast cancers. Here, we show that 15d-PGJ(2) inhibits both 17beta-estradiol (E(2))-dependent and E(2)-independent ERalpha transcriptional activity by PPARgamma-independent mechanism. In addition, 15d-PGJ(2) directly modifies ERalpha protein via its reactive cyclopentenone moiety, evidenced by incorporation of biotinylated 15d-PGJ(2) into ERalpha, both in vitro and in vivo. Nanoflow reverse-phase liquid chromatography tandem mass spectrometry analysis identifies two cysteines (Cys(227) and Cys(240)) within the COOH-terminal zinc finger of ERalpha DNA-binding domain (DBD) as targets for covalent modification by 15d-PGJ(2). Gel mobility shift and chromatin immunoprecipitation assays show that 15d-PGJ(2) inhibits DNA binding of ERalpha and subsequent repression of ERalpha target gene expression, such as pS2 and c-Myc. Therefore, our results suggest that 15d-PGJ(2) can block ERalpha function by covalent modification of cysteine residues within the vulnerable COOH-terminal zinc finger of ERalpha DBD, resulting in fundamental inhibition of both hormone-dependent and hormone-independent ERalpha transcriptional activity.
Amino acid sequence, breast neoplasms, cell line, tumor, cysteine, DNA, neoplasm, estradiol, estrogen antagonists, estrogen receptor alpha, protein structure, tertiary, transcriptional activation, transfection, zinc fingers
Kim, Han-Jong; Kim, Joon-Young; Meng, Zhaojing; Wang, Li Hua; Liu, Fa; Conrads, Thomas P.; Burke, Terrence R.; Veenstra, Timothy D.; and Farrar, William L., "15-Deoxy-Delta12,14-Prostaglandin J2 Inhibits Tanscriptional Activity of Estrogen Receptor-Alpha via Covalent Modification of DNA-Binding Domain" (2007). Pharmaceutical Sciences Faculty Publications. 385.