2001 Fiscal Year Final Research Report Summary
Molecular mechanism for the assembly of red cell membrane skeletons based on pathobiology of congenital hemolytic anemia in cattle
| Project/Area Number |
12460137
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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 |
Applied veterinary science
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| Research Institution | The University of Tokyo |
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Principal Investigator |
INABA Mutsumi Graduate School of Agriculture and Life Sciences, The University of Tokyo, Asssociate Professor, 大学院・農学生命科学研究科, 助教授 (00183179)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAKUWA Yuichi Tokyo Women's Medical University, Professor, 医学部, 教授 (40113740)
ONO Ken-ichiro Graduate School of Agriculture and Life Sciences, The University of Tokyo, Professor, 大学院・農学生命科学研究科, 教授 (50111480)
YAMAMOTO Masayuki Tsukuba University TARA Center, Professor, 基礎医学系・先端学際領域研究センター, 教授 (50166823)
MATSUKI Naoaki Graduate School of Agriculture and Life Sciences, The University of Tokyo, Research Associate, 大学院・農学生命科学研究科, 助手 (40251417)
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| Project Period (FY) |
2000 – 2001
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| Keywords | red cell membranes / membrane skeleton / band 3 / ankyrin / congenital hemolytic anemia / proeasome / vesicle trafficking / cattle |
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| Research Abstract |
Stable transfectants of normal bovine band 3 (bebWT) or the mutant band 3 with R664X mutation (bebRX) were established in K562 cells and HEK293 cells using retoriviral vectors. Transfected cells expressing EGFP-bebWT and EGFP-bebRX, and N-terminal domain of ankyrin (AnkN90) in combination with band 3 proteins were also prepared.
The bebWT and EGFP-bebWT showed stable expression on the plasma membrane of the transfected cells, whereas the mutant proteins, bebRX and EGFP-bebRX were degraded by Ub-proteasome system soon after synthesis on the ER or after retrograde transported from the Golgi apparatus to the ER. Stability of the bebWT was extremely reduced when bebRX was co-transfected. AnkN90 showed membrane localization within the cells and was destabilized in the cells that had the mutant band 3.
These findings indicate that the mutant band 3 (bebRX) plays a dominant-negative role on the expression of normal band 3 and a partner in the membrane skeleton, ankyrin, and the interaction of band 3 with ankyrin occurs on the ER membrane soon after band 3 synthesis is started during erythroid development.
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[Publications] Tamahara, S., Inaba, M., Sato, K., Matsuki, N., Hikasa, Y., and Ono, K.: "Nonessential roles of cysteine residues in functional expression and redox regulatory pathways for canine glutamate/aspartate transporter based on mutagenic analysis"Biochem. J.. 367. 107-111 (2002)
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[Publications] An, X.-L., Takakuwa, Y., Manno, S., Han, E.G., Gascard, P., and Mohandas, N.: "Structural and functional characterization of protein 4.1R-phosphatidylserine interaction: potential tole in 4.1R sorting within cells"J. Biol. Chem.. 276. 35778-35785 (2001)
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[Publications] Sato, K., Inaba, M., Baba, K., Tamahara, S., Koshino, I., Hikasa, Y., Ono, K., and Kagota, K.: "Cloning and characterization of excitatory amino acid transporters GLT-1 and EAAC1 in canine brain"J. Vet. Med. Sci.. 63. 997-1002 (2001)
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[Publications] Sato, K., Inaba, M., Suwa, Y., Matsuu, A., Hikasa, Y., Ono, K., and Kagota, K.: "Inherited defects of Na-dependent glutamate transport mediated by glutamate/aspartate transporter in canine red cells due to a decreased level of transporter protein expression"J. Biol. Chem.. 275. 6620-6627 (2000)
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「研究成果報告書概要(欧文)」より
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