title: Sialyl-glycoconjugates in cholesterol-rich microdomains of P388 cells are the triggers for apoptosis induced by Rana catesbeiana oocyte ribonuclease.
abstract: SBL/RC-RNase was originally isolated from frog (Rana catesbeiana) oocytes and purified as a novel sialic acid-binding lectin (SBL) that displayed strong anti-cancer activity. SBL was later shown to be identical to a ribonuclease (RC-RNase) from oocytes of the same species. The administration of SBL/RC-RNase induced apoptosis (with nuclear condensation and DNA fragmentation) in mouse leukemia P388 cells but did not kill umbilical vein endothelial or fibroblast cells derived from normal tissues. The cytotoxic activity of SBL/RC-RNase was inhibited by desialylation of P388 cells and/or the co-presence of free bovine submaxillary mucin. FACS analysis showed that SBL/RC-RNase was incorporated into cells after attachment to cholesterol-rich microdomains. Addition of the cholesterol remover methyl-β-cyclodextrin reduced SBL/RC-RNase-induced apoptosis. Apoptosis occurred through the caspase-3 pathway following activation of caspase-8 by SBL/RC-RNase. A heat shock cognate protein (Hsc70) and a heat shock protein (Hsp70) (each 70 kDa) on the cell membrane were shown to bind to SBL/RC-RNase by mass spectrometric and flow cytometric analyses. Quercetin, an inhibitor of Hsc70 and Hsp70, significantly reduced SBL/RC-RNase-induced apoptosis. Taken together, our findings suggest that sialyl-glycoconjugates present in cholesterol-rich microdomains form complexes with Hsc70 or Hsp70 that act as triggers for SBL/RC-RNase to induce apoptosis through a pathway involving the activation of caspase-3 and caspase-8.
authors.affiliation: Divisions of Microbiology and Functional Morphology, Department of Pharmacy, Faculty of Pharmaceutical Science, Nagasaki International University, 2825-7 Huis Ten Bosch, Sasebo, Nagasaki, 859-3298, Japan, yogawa@niu.ac.jp.
authors.forename: Y
authors.initials: Y
authors.lastname: Ogawa
authors.forename: S
authors.initials: S
authors.lastname: Sugawara
authors.forename: T
authors.initials: T
authors.lastname: Tatsuta
authors.forename: M
authors.initials: M
authors.lastname: Hosono
authors.forename: K
authors.initials: K
authors.lastname: Nitta
authors.forename: Y
authors.initials: Y
authors.lastname: Fujii
authors.forename: H
authors.initials: H
authors.lastname: Kobayashi
authors.forename: T
authors.initials: T
authors.lastname: Fujimura
authors.forename: H
authors.initials: H
authors.lastname: Taka
authors.forename: Y
authors.initials: Y
authors.lastname: Koide
authors.forename: I
authors.initials: I
authors.lastname: Hasan
authors.forename: R
authors.initials: R
authors.lastname: Matsumoto
authors.forename: H
authors.initials: H
authors.lastname: Yasumitsu
authors.forename: R A
authors.initials: RA
authors.lastname: Kanaly
authors.forename: S M A
authors.initials: SM
authors.lastname: Kawsar
authors.forename: Y
authors.initials: Y
authors.lastname: Ozeki
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002470:Cell Survival; D002784:Cholesterol / Q000737:chemistry*; D006001:Glycoconjugates / Q000378:metabolism*; D050883:HSC70 Heat-Shock Proteins / Q000378:metabolism; D050884:HSP72 Heat-Shock Proteins / Q000378:metabolism; D007941:Leukemia P388; D021962:Membrane Microdomains / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D019158:N-Acetylneuraminic Acid / Q000737:chemistry; D009865:Oocytes / Q000201:enzymology*; D011892:Rana catesbeiana; D012260:Ribonucleases / Q000378:metabolism*
chemical_list: D006001:Glycoconjugates; D050883:HSC70 Heat-Shock Proteins; D050884:HSP72 Heat-Shock Proteins; D002784:Cholesterol; D012260:Ribonucleases; D019158:N-Acetylneuraminic Acid