| Co-Investigator(Kenkyū-buntansha) |
KANEMATSU Takashi Faculty of Dental Science, Aso. Professor, 大学院・歯学研究院, 助教授 (10264053)
HIRATA Masato Faculty of Dental Science, Professor, 大学院・歯学研究院, 教授 (60136471)
OHISHI Masamichi Faculty of Dental Science, Professor, 大学院・歯学研究院, 教授 (70037505)
MATSUDA Miho Faculty of Dental Science, Ass. Professor, 大学院・歯学研究院, 助手 (40291520)
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| Research Abstract |
PRIP-1 was originally identified as an inositol 1,4,5-trisphosphate [Ins(1,4,5)P_3] binding protein similar to phospholipase C (PLC)-δ1, but lacking any phospholipase activity. In this study we analyzed the effects of the pleckstrin homology domain (PH domain) of PRIP-1 on the Ins(1,4,5)P_3-mediated Ca^<2+> signaling using permeabilized and intact HeLa cells, and found that PRIP-1PH domain had an inhibitory effect on Ca^<2+> signaling. PRIP-1PH domain inhibited the Ins(1,4,5)P_3-mediated Ca^<2+> release from permeabilized cells in a dose-dependent manner. When fura-2 loaded HeLa cells transfected with PRIP-1PH domain were stimulated with ATP, it was found that the agonist-induced increase in free Ca^<2+> concentration, observed in control cells, was inhibited in transfected cells. This inhibition was not accompanied by the reduced production of Ins(1,4,5)P_3. These results suggest that PRIP-1 has a potential to inhibit Ins(1,4,5)P_3-mediated Ca^<2+> signaling.
It has been reported that
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inositol hexakisphosphate (InsP_6, phytic acid), a natural product, has an anticancer role. In the present study, we investigated the mechanism by which InsP_6 acts as an anticancer agent. Treatment of HeLa cells with InsP_6 at 1 mM induced apoptosis, as assessed by counting the cell number, and by Hoechst and TUNEL staining. This is probably mediated by intracellular InsP_6 itself and/or the dephosphorylated forms of metabolized InsP_6, because incubation of HeLa cells with [^3H]InsP_6 produces dephosphorylated forms such as InsP_4 and InsP_5. Induction of apoptosis by InsP_6 was examined in two ways: inhibition of cell survival signaling and direct induction of apoptosis. As well as inhibiting the cell survival Akt-NF_κB pathway, InsP_6 itself caused mitochondrial permeabilization, followed by cytochrome c release, which later caused activation of the apoptotic machinery, caspase 9, 3 and poly (ADP-ribose) polymerase. These results revealed that extracellularly-applied InsP_6 directly activates the apoptotic machinery as well as inhibits the cell survival signaling, probably by the intracellular delivery followed by a dephosphorylation. Less
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