Functional proteomics analysis of non-vesicular release mechanisms in stress exposed neuron-role of stress-induced non-vesicular release Prion, NDI and FGF-1 in neuron
| Project/Area Number |
15390028
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biological pharmacy
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| Research Institution | Nagasaki University |
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Principal Investigator |
UEDA Hiroshi Nagasaki University, Graduate School of Biomedical Sciences, Professor, 大学院・医歯薬学総合研究科, 教授 (00145674)
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| Co-Investigator(Kenkyū-buntansha) |
SAKAGUCHI Suehiro Nagasaki University, Graduate School of Biomedical Sciences, Associate Professor, 大学院・医歯薬学総合研究科, 助教授 (60274635)
INOUE Makoto Nagasaki University, Graduate School of Biomedical Sciences, Assistant Professor, 大学院・医歯薬学総合研究科, 講師 (60380987)
FUJITA Ryousuke Nagasaki University, Graduate School of Biomedical Sciences, Research Associate, 大学院・医歯薬学総合研究科, 助手 (70380855)
水野 恭伸 長崎大学, 大学院・医歯薬学総合研究科, 助手 (40311865)
吉田 明 長崎大学, 大学院・医歯薬学総合研究科, 助教授 (70257187)
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| Project Period (FY) |
2003 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥15,400,000 (Direct Cost: ¥15,400,000)
Fiscal Year 2004: ¥5,900,000 (Direct Cost: ¥5,900,000)
Fiscal Year 2003: ¥9,500,000 (Direct Cost: ¥9,500,000)
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| Keywords | Stress / non-vesicular release / S100A13 / FGF-1 / serum starvation / Ca^<2+> / STI / NDI / 神経細胞死 / ネクローシス / 分泌 / 核移行 / 低酸素 / S100 |
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| Research Abstract |
The purpose of this research is to demonstrate the stress-induced mechanism of non-vesicular release FGF-1, NDI and STI, as prion (PrP^c) associated protein using the proteomics analyses. We attempted to characterize the non-vesicular mode of FGF-1 and NDI release in the analyses using immunocytochemistry and immunoblot of conditioned medium (CM) from astrocytes. FGF-1 was completely released from astrocytes upon serum-deprivation stress in a Brefeldin A-insensitive manner. In the immunoprecipitation study using anti-FGF-1 IgG, S100A13 was identified to be the major protein co-eluted with FGF-1. The interaction between GST-FGF-1 and Strep-tag II S100A13 was found to be Ca^<2+>-sensitive, and to require the C-terminal 11 amino acid peptide sequence of S100A13. The overexpression of D88-98 mutant of S100A13 selectively inhibited the serum-deprivation stress-induced release of FGF-1, but not the release of S100A13 mutant from C6 glioma cells. However, amlexanox, anti-allergic drug whose t
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arget is S100A13, completely inhibited the stress-induced release of FGF-1 as well as S100A13. The stress-induced release of both proteins was also abolished by BAPTA-AM, an intracellular Ca^<2+> chelating agent. The serum-deprivation caused Ca^<2+> spikes in v-conotoxin GVIA and thapsigargin-sensitive manner. These same results observed NDI protein experiments. These results suggest that S100A13 is a cargo molecule for the serum-deprivation stress-induced non-vesicular release of FGF-1 and NDI, and that its driving force of protein-protein interaction and release is possibly mediated by Ca^<2+>-induced Ca^<2+> release (CICR) coupled to N-type Ca^<2+> channel activity. On the other hand, STI have neuronal survival activity. STI inhibitor, as a PrP^c c terminal peptide (113-132), treated neuron was enhanced decrease in survival activity under the oxygen-glucose deprivation. Moreover, STI inhibitor induced neuronal cell death under the serum containing normal condition. These results suggested that PrP^c have increase in neuronal survival activity under the normal condition. Less
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Report
(3 results)
Research Products
(8 results)
