1994 Fiscal Year Final Research Report Summary
Molecular analysis of dihydropteridine reductase deficiency
| Project/Area Number |
05670651
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Pediatrics
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| Research Institution | Tohoku University |
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Principal Investigator |
MATSUBARA Yoichi Tohoku University School of Medicine, Department of Biochemical Genetics Associate Professor, 医学部, 助教授 (00209602)
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| Co-Investigator(Kenkyū-buntansha) |
MIKAMI Hitoshi Tohoku University School of Medicine, Department of Pediatrics Assistant Profess, 医学部, 助手
SUZUKI Yoichi Tohoku University School of Medicine, Department of Biolchemical Genetics Assist, 医学部, 助手 (80216457)
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| Project Period (FY) |
1993 – 1994
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| Keywords | dihydropteridine reductase / genetic mutation / phenylketonuria / biopterin / PCR |
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| Research Abstract |
Dihydropteridine reductase (DHPR) plays an essential role in the metabolism of tetrahydrobiopterin, which is required as a cofactor for hydroxylation of phenylalanine, tyrosine and tryptophane. Deficiency of DHPR is inherited as an autosomal recessive disorder characterized by hyperphenylalaninemia and various neurological symptoms such as severe mental retardation and convulsion. We have studied three Japanese patients with DHPR deficiency. Northern blot analysis showed that DHPR mRNA was normally expressed in a cell culture obtained from case 1, whereas marked reduction of DHPR mRNA was observed in cultures from case 2 and 3. Sequencing analysis of DHPR mRNA from case 1 by reverse transcription/polymerase chain reaction (RT-PCR) identified a t-to-c change at nucleotide position 130, which resulted in a substitution of tryptophane with arginine at amino acid position 36 (W36R). Expression analysis of DHRP cDNA harboring W36R did not show DHPR activity, indicating that it is a disease-causing mutation. Case 1, a product of a consanguineous marriage, was homozygous for the mutation. RT-PCR analysis of case 2 revealed an abnormally spliced DHPR mRNA with a 152 bp-insertion between exon 3 and 4. The insertion encoded isoleucine and leucine followed by a stop codon. The analysis of genomic DNA revealed an a-to-g substitution in intron 3 which created a novel splicing donor site. The mutation probably activated a potential splicing acceptor site located 152 bp upstream, producing a "new" exon in intron 3. The mutation was present in homozygous form in case 2 as well as in his affected brother, while the mother was a heterozygote. Case 3 did not carry nucleotide substitution in the coding region of DHPR cDNA.The case probably has a mutation outside the coding region, which affects the expression or the stability of mRNA.
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