title: Induction of apoptosis in human prostate cancer cells by insulin-like growth factor binding protein-3 does not require binding to retinoid X receptor-alpha.
abstract: IGF binding protein (IGFBP)-3 can induce apoptosis in human prostate cancer cells directly without sequestering IGF-I and -II. The molecular mechanisms responsible for the IGF-independent actions of IGFBP-3 remain unclear. IGFBP-3, a secreted protein, can be internalized and translocate to the nucleus. It binds to the nuclear retinoid X receptor (RXR)-alpha. Binding to RXR-alpha has been proposed to be required for IGFBP-3 to induce apoptosis. The present study tests this hypothesis in the PC-3 human prostate cancer cell line. PC-3 cells express RXR-alpha, and apoptosis is induced by incubation with RXR-specific ligand. A COOH-terminal region in IGFBP-3 (residues 215-232) contains a nuclear localization signal, and binding domains for RXR-alpha and heparin (HBD). Different combinations of the 11 amino acids in this region that differ from IGFBP-1, a related IGFBP, which does not localize to the nucleus or bind RXR-alpha, were mutated to the IGFBP-1 sequence. By confocal imaging, mutation of residues 228-KGRKR-232 in nonsecreted IGFBP-3 diminished its nuclear localization. IGFBP-3 binding to glutathione S-transferase-RXR-alpha only was lost when all 11 sites were mutated (HBD-11m-IGFBP-3). Expressed nuclear RXR-alpha did not transport cytoplasmic IGFBP-3 nuclear localization signal mutants that can bind RXR-alpha to the nucleus even after treatment with RXR ligand. Expressed HBD-11m-IGFBP-3 still induced apoptosis in PC-3 cells in an IGF-independent manner as determined by flow cytometric analysis of Annexin V staining. We conclude that in PC-3 cells, RXR-alpha is not required for the nuclear translocation of IGFBP-3 and that IGFBP-3 can induce apoptosis in human prostate cancer cells without binding RXR-alpha.
authors.affiliation: Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 9000 Rockville Pike, Bethesda, MD 20892, USA.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Zappala
authors.forename: Cem
authors.initials: C
authors.lastname: Elbi
authors.forename: Joanna
authors.initials: J
authors.lastname: Edwards
authors.forename: Julie
authors.initials: J
authors.lastname: Gorenstein
authors.forename: Matthew M
authors.initials: MM
authors.lastname: Rechler
authors.forename: Nisan
authors.initials: N
authors.lastname: Bhattacharyya
mesh_terms: D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D017209:Apoptosis; D045744:Cell Line, Tumor; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000737:chemistry / Q000502:physiology*; D008297:Male; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D011471:Prostatic Neoplasms / Q000473:pathology*; D047490:Retinoid X Receptor alpha / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D018972:Insulin-Like Growth Factor Binding Protein 3; D019913:Nuclear Localization Signals; D047490:Retinoid X Receptor alpha