ABSTRACT
Previously, we reported that glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a binding partner of prolyl oligopeptidase (POP) in neuroblastoma NB-1 cells and that the POP inhibitor, SUAM-14746, inhibits cytosine arabinoside (Ara-C)-induced nuclear translocation of GAPDH and protects against Ara-C cytotoxicity. To carry out a more in-depth analysis of the interaction between POP and GAPDH, we generated POP-KO NB-1 cells and compared the nuclear translocation of GAPDH after Ara-C with or without SUAM-14746 treatment to wild-type NB-1 cells by western blotting and fluorescence immunostaining. Ara-C did not induce the nuclear translocation of GAPDH and SUAM-14746 did not protect against Ara-C cytotoxicity in POP-KO cells. These results indicate that the anticancer effects of Ara-C not only include the commonly known antimetabolic effects, but also the induction of cell death by nuclear transfer of GAPDH through interaction with POP.
Subject(s)
Cell Nucleus/drug effects , Cytarabine/pharmacology , Glyceraldehyde-3-Phosphate Dehydrogenases/metabolism , Prolyl Oligopeptidases/metabolism , Cell Death/drug effects , Cell Nucleus/metabolism , Cell Survival/drug effects , Cytarabine/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Humans , Proline/analogs & derivatives , Proline/pharmacology , Prolyl Oligopeptidases/antagonists & inhibitors , Prolyl Oligopeptidases/deficiency , Thiazolidines/pharmacology , Tumor Cells, CulturedABSTRACT
The nucleoside triphosphate hydrolases that are produced by Neospora caninum (NcNTPase) and Toxoplasma gondii (TgNTPase-I) have a different physiological function from the ubiquitous ecto-ATPases. The recombinant enzymes were crystallized at 293â K using polyethylene glycol 3350 as a precipitant and X-ray diffraction data sets were collected for NcNTPase (to 2.8â Å resolution) and TgNTPase-I (to 3.1â Å resolution) at 100â K using synchrotron radiation. The crystals of NcNTPase and TgNTPase-I belonged to the orthorhombic space group I222 (unit-cell parameters a = 93.6, b = 140.8, c = 301.1â Å) and the monoclinic space group P2(1) (unit-cell parameters a = 87.1, b = 123.5, c = 120.2â Å, ß = 96.6°), respectively, with two NcNTPase (V(M) = 3.7â Å(3)â Da(-1)) and four TgNTPase-I (V(M) = 2.7â Å(3)â Da(-1)) molecules per asymmetric unit. SAD phasing trials using a data set (λ = 0.97904â Å) collected from a crystal of selenomethionylated NcNTPase gave an initial electron-density map of sufficient quality to build a molecular model of NcNTPase.
