| Project/Area Number |
12144205
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| Research Category |
Grant-in-Aid for Scientific Research on Priority Areas
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| Allocation Type | Single-year Grants |
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| Review Section |
Biological Sciences
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| Research Institution | Toyama Medical and Pharmaceutical University |
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Principal Investigator |
SAKAI Hideki (2004) Toyama Medical and Pharmaceutical University, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (60242509)
竹口 紀晃 (2000-2003) 富山医科薬科大学, 副学長 (00019126)
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| Co-Investigator(Kenkyū-buntansha) |
MORII Magotoshi Toyama Medical and Pharmaceutical University, Faculty of Pharmaceutical Sciences, Associate Professor, 薬学部, 講師 (60019130)
酒井 秀紀 富山医科薬科大学, 薬学部, 助教授 (60242509)
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| Project Period (FY) |
2000 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥33,800,000 (Direct Cost: ¥33,800,000)
Fiscal Year 2004: ¥8,400,000 (Direct Cost: ¥8,400,000)
Fiscal Year 2003: ¥8,400,000 (Direct Cost: ¥8,400,000)
Fiscal Year 2002: ¥8,500,000 (Direct Cost: ¥8,500,000)
Fiscal Year 2001: ¥8,500,000 (Direct Cost: ¥8,500,000)
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| Keywords | gastric proton pump / gastric acid secretion / phosphorvlation / proteasome / acid pump antagonist / flippase / thromboxane / Na^+.K^+-ATPase / リン脂質 / 分子動力学 / Cl^-チャネル / 大腸 / 安定発現細胞 / プロトンポンプ阻害剤 / 大腸がん / 遺伝子発現 / 大腸プロトンポンプ / イオンポンプ / ナトリウムポンプ |
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| Research Abstract |
We investigated vectorial-transporting mechanisms of gastric acid secretion and intestinal ion-secretion.
1. We established the stable cell lines expressing the gastric proton a-and/or B-subunits. The a-subunit was retained in intracellular compartment and was unstable in the absence of β-subunit, but it was stabilized and reached the cell surface in the presence of β-subunit. We found that the extracellular three disulfide bonds of β-subunit are essential for assembly with a-subunit and expression of the pump activity as well as for cell surface delivery of a-subunit. The ubiquitin/proteasome system was involved in degradation of unassembled α-subunits in ER.
2. We found that Glu-345 in the fourth transmembrane (M4) segment of the proton pump is involved in cation-induced conformational change in the pump. We also found that Leu-819 and Glu-822 in the M6 segment are involved in K+-dependent dephosphorylation, and that Asp-826, Ile-827 and Leu-833 in the M6 are involved in phosphorylation of the pump.
3. We clarified that the binding site of SCH 28080, an acid pump antagonist, is a cavity in the E2 form of the proton pump a-subunit, and Tyr-801 faces the cavity.
4. We found that the phospholipid flippase activity is part of the proton pump reaction.
5. We confirmed that ClC-2 Cl^-channel is not a Cl^-transporting protein for gastric acid secretion in parietal cells.
6. We established new gastric epithelial cell lines expressing proton a-subunit mRNA from mice transgenic for temperature-sensitive simian virus 40 large T antigen.
7. We found that thromboxane A2 is involved in the mechanism of secretory diarrhea in colon, and that thromboxane A2 is released only in pathophysiological condition.
8. We found that a decrease in Na^+,K^+-ATPase al-isoform expression and an increase in Na^+,K^+-ATPase a3-isoform expression are associated with human colorectal cancer.
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