1999 Fiscal Year Final Research Report Summary
Biological and pharmaceutical studies on the mechanism of intracellular sorting of cholesterol and vitamin E
| Project/Area Number |
09470500
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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 | GRADUATE SCHOOL OF PHARMACEUTICAL SCIENCES, THE UNIVERSITY OF TOKYO |
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Principal Investigator |
KRAI Hiroyuki GRADUATE SCHOOL OF PARM. SCI., THE UNIV. TOKYO, ASSOCIATE PROFESSOR, 大学院・薬学系研究科, 助教授 (40167987)
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| Co-Investigator(Kenkyū-buntansha) |
AOKI Junken GRADUATE SCHOOL OF PARM. SCI., THE UNIV. TOKYO, ASSISTANT PROFESSOR, 大学院・薬学系研究科, 助手 (20250219)
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| Project Period (FY) |
1997 – 1999
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| Keywords | CHOLESTEROL / HDL / SR-BI / PDZ DOMAIN / VITAMIN E / TRANSPORT PROTEIN / LIVER / SERCRETION |
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| Research Abstract |
In this study we identified an SR-BI-associated protein from rat liver membrane extracts using affinity chromatography technique. This protein of 523 amino acids contains 4 PDZ domains and associates with the C-terminal of SR-BI using its N-terminal first PDZ domain. Therefore we denoted this protein as CLAMP (carboxy-terminal linking and modulating protein). CLAMP was located mostly in the sinusoidal membranes, while SR-BI was detected in both sinusoidal and canalicular membranes. After the solubilization of the liver membranes with Triton X-100, SR-BI was immunoprecipitated with anti-CLAMP monoclonal antibody, suggesting the association of these proteins in vivo By co-expressing SR-BI with CLAMP in CHO cells, we observed 1) the increase in the expression level of SR-BI, 2) the reduction in the deacylation rate of the cholesteryl esters taken up from HDL, and 3) the change in the intracellular distribution of fluorescent lipid DiI taken up from HDL. a-Tocopherol taken up by the liver
… More
with lipoprotein is thought to be re-secreted into the plasma in very low density lipoprotein (VLDL). a-Tocopherol transfer protein (aTTP) is a cytosolic liver protein and plays all important role in the efficient recycling of plasma vitamin E. Using using the hepatoma cell line, we found that the secretion of a-tocopherol was more efficient in cells expressing aTTP than in matched cells lacking aTTP. Brefeldin A, which effectively inhibits VLDL secretion by disrupting the Golgi apparatus, bad no effect on a-tocopherol secretion, indicating that aTTP-mediated a-tocopherol secretion is not coupled to VLDL secretion. Among other agents tested, only 25-hydroxycholesterol, a modulator of cholesterol metabolism, inhibited a-tocopherol secretion. This inhibition is most likely mediated by oxysterol binding protein. These results suggest that aTTP present in the liver cytosol functions to stimulate secretion of cellar a-tocopherol into the extracellular medium and that the reaction utilizes a novel non-Golgi mediated pathway which may be linked to cellular cholesterol metabolism and/or transport. Less
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