| Project/Area Number |
13671196
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Metabolomics
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| Research Institution | Kyoto Prefectural University of Medicine |
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Principal Investigator |
NOSAKA Kazuto Kyoto Prefectural University of Medicine, Grad. Sch. Med., Associate Professor, 医学研究科, 助教授 (10228314)
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| Co-Investigator(Kenkyū-buntansha) |
YAMANISHI Kiyofumi Mie University, Sch. Med, Associate Professor, 医学部, 助教授 (10182586)
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| Project Period (FY) |
2001 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2003: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2002: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2001: ¥700,000 (Direct Cost: ¥700,000)
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| Keywords | thiamin / thiamin-responsive megaloblastic anemia / thiamin transporter / thiamin pyrophosphokinase |
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| Research Abstract |
Thiamin-responsive megaloblastic anemia (TRMA ; OMIM 249270) is an autosomal recessive disorder defined by the occurrence of megaloblastic anemia, diabetes mellitus, sensorineural deafness, and responding to thiamin treatment. It has been speculated that TRMA is caused by the aberration of one of the two proteins both required for thiamin pyrophosphate synthesis, thiamin pyrophosphokinase (TPK) and thiamin transporter. We and others identified SLC192A as the disease gene that encodes the high affinity thiamin transporter (THTR1).
In this study, we isolated the complementary and genome DNAs of mouse thiamin transporter (mTHTR1). Using 5'-RACE two transcriptional start sites for SLC192A located at -175 and -183 relative to the translation start codon were identified in mouse liver cells. We planed to generate a mouse model of SLC192A deficiency to understand the pathogenesis of TRMA. On the other hand, we also isolated human TPK cDNA (hTPK1) and analyzed the enzymatic properties. Furthermore, we identified the minimal 5'-promoter region required for the expression of hTPK1 gene in HepG2 cells which was found to be encoded in a sequence between -540〜-490,and included a Sp1 binding site. Mutation in Sp1 site in the region led to a decrease in the promoter activity. Using Electrophoretic mobility shift analysis with the nuclear extracts from HepG2 cells and the synthetic 24bp oligonucleotide corresponding to -514〜-491 sequence of hTPK1 promoter region, specific DNA-protein complexes were identified. These results suggested the importance of Sp1 in regulating the hTPK1 expression.
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