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. 1993 Feb;52(2):414–424.

Occurrence of a 2-bp (AT) deletion allele and a nonsense (G-to-T) mutant allele at the E2 (DBT) locus of six patients with maple syrup urine disease: multiple-exon skipping as a secondary effect of the mutations.

C W Fisher 1, C R Fisher 1, J L Chuang 1, K S Lau 1, D T Chuang 1, R P Cox 1
PMCID: PMC1682180  PMID: 8430702

Abstract

We have identified two novel mutant alleles in the transacylase (E2) gene of the human branched-chain alpha-keto acid dehydrogenase (BCKAD) complex in 6 of 38 patients with maple syrup urine disease (MSUD). One mutation, a 2-bp (AT) deletion in exon 2 of the E2 gene, causes a frameshift downstream of residue (-26) in the mitochondrial targeting presequence. The second mutation, a G-to-T transversion in exon 6 of the E2 gene, produces a premature stop codon at Glu-163 (E163*). Transfection of constructs harboring the E163* mutation into an E2-deficient MSUD cell line produced a truncated E2 subunit. However, this mutant E2 chain is unable to assemble into a 24-mer cubic structure and is degraded in the cell. The 2-bp (AT) deletion and the E163* mutant alleles occur in either the homozygous or compound-heterozygous state in the 6 of 38 unrelated MSUD patients studied. Moreover, an array of precise single- and multiple-exon deletions were observed in many amplified E2 mutant cDNAs. The latter results appear to represent secondary effects on RNA processing that are associated with the MSUD mutations at the E2 locus.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Chen E. Y., Seeburg P. H. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. doi: 10.1089/dna.1985.4.165. [DOI] [PubMed] [Google Scholar]
  2. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  3. Chuang D. T., Hu C. W., Ku L. S., Markovitz P. J., Cox R. P. Subunit structure of the dihydrolipoyl transacylase component of branched-chain alpha-keto acid dehydrogenase complex from bovine liver. Characterization of the inner transacylase core. J Biol Chem. 1985 Nov 5;260(25):13779–13786. [PubMed] [Google Scholar]
  4. Chuang D. T., Niu W. L., Cox R. P. Activities of branched-chain 2-oxo acid dehydrogenase and its components in skin fibroblasts from normal and classical-maple-syrup-urine-disease subjects. Biochem J. 1981 Oct 15;200(1):59–67. doi: 10.1042/bj2000059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Eisenstein R. S., Hoganson G., Miller R. H., Harper A. E. Altered phosphorylation state of branched-chain 2-oxo acid dehydrogenase in a branched-chain acyltransferase deficient human fibroblast cell line. J Inherit Metab Dis. 1991;14(1):37–44. doi: 10.1007/BF01804386. [DOI] [PubMed] [Google Scholar]
  6. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  7. Fisher C. R., Chuang J. L., Cox R. P., Fisher C. W., Star R. A., Chuang D. T. Maple syrup urine disease in Mennonites. Evidence that the Y393N mutation in E1 alpha impedes assembly of the E1 component of branched-chain alpha-keto acid dehydrogenase complex. J Clin Invest. 1991 Sep;88(3):1034–1037. doi: 10.1172/JCI115363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fisher C. W., Chuang J. L., Griffin T. A., Lau K. S., Cox R. P., Chuang D. T. Molecular phenotypes in cultured maple syrup urine disease cells. Complete E1 alpha cDNA sequence and mRNA and subunit contents of the human branched chain alpha-keto acid dehydrogenase complex. J Biol Chem. 1989 Feb 25;264(6):3448–3453. [PubMed] [Google Scholar]
  9. Fisher C. W., Lau K. S., Fisher C. R., Wynn R. M., Cox R. P., Chuang D. T. A 17-bp insertion and a Phe215----Cys missense mutation in the dihydrolipoyl transacylase (E2) mRNA from a thiamine-responsive maple syrup urine disease patient WG-34. Biochem Biophys Res Commun. 1991 Jan 31;174(2):804–809. doi: 10.1016/0006-291x(91)91489-y. [DOI] [PubMed] [Google Scholar]
  10. Griffin T. A., Lau K. S., Chuang D. T. Characterization and conservation of the inner E2 core domain structure of branched-chain alpha-keto acid dehydrogenase complex from bovine liver. Construction of a cDNA encoding the entire transacylase (E2b) precursor. J Biol Chem. 1988 Oct 5;263(28):14008–14014. [PubMed] [Google Scholar]
  11. Harrison B., Zimmerman S. B. T4 polynucleotide kinase: macromolecular crowding increases the efficiency of reaction at DNA termini. Anal Biochem. 1986 Nov 1;158(2):307–315. doi: 10.1016/0003-2697(86)90555-5. [DOI] [PubMed] [Google Scholar]
  12. Herring W. J., Litwer S., Weber J. L., Danner D. J. Molecular genetic basis of maple syrup urine disease in a family with two defective alleles for branched chain acyltransferase and localization of the gene to human chromosome 1. Am J Hum Genet. 1991 Feb;48(2):342–350. [PMC free article] [PubMed] [Google Scholar]
  13. Hu C. W., Griffin T. A., Lau K. S., Cox R. P., Chuang D. T. Subunit structure of the dihydrolipoyl transacylase component of branched-chain alpha-keto acid dehydrogenase complex from bovine liver. Mapping of the lipoyl-bearing domain by limited proteolysis. J Biol Chem. 1986 Jan 5;261(1):343–349. [PubMed] [Google Scholar]
  14. Lau K. S., Griffin T. A., Hu C. W., Chuang D. T. Conservation of primary structure in the lipoyl-bearing and dihydrolipoyl dehydrogenase binding domains of mammalian branched-chain alpha-keto acid dehydrogenase complex: molecular cloning of human and bovine transacylase (E2) cDNAs. Biochemistry. 1988 Mar 22;27(6):1972–1981. doi: 10.1021/bi00406a025. [DOI] [PubMed] [Google Scholar]
  15. Lau K. S., Herring W. J., Chuang J. L., McKean M., Danner D. J., Cox R. P., Chuang D. T. Structure of the gene encoding dihydrolipoyl transacylase (E2) component of human branched chain alpha-keto acid dehydrogenase complex and characterization of an E2 pseudogene. J Biol Chem. 1992 Nov 25;267(33):24090–24096. [PubMed] [Google Scholar]
  16. MENKES J. H., HURST P. L., CRAIG J. M. A new syndrome: progressive familial infantile cerebral dysfunction associated with an unusual urinary substance. Pediatrics. 1954 Nov;14(5):462–467. [PubMed] [Google Scholar]
  17. Margolskee R. F., Kavathas P., Berg P. Epstein-Barr virus shuttle vector for stable episomal replication of cDNA expression libraries in human cells. Mol Cell Biol. 1988 Jul;8(7):2837–2847. doi: 10.1128/mcb.8.7.2837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Matsuo M., Nishio H., Kitoh Y., Francke U., Nakamura H. Partial deletion of a dystrophin gene leads to exon skipping and to loss of an intra-exon hairpin structure from the predicted mRNA precursor. Biochem Biophys Res Commun. 1992 Jan 31;182(2):495–500. doi: 10.1016/0006-291x(92)91759-j. [DOI] [PubMed] [Google Scholar]
  19. Miles J. S., Guest J. R. Subgenes expressing single lipoyl domains of the pyruvate dehydrogenase complex of Escherichia coli. Biochem J. 1987 Aug 1;245(3):869–874. doi: 10.1042/bj2450869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mitsubuchi H., Nobukuni Y., Akaboshi I., Indo Y., Endo F., Matsuda I. Maple syrup urine disease caused by a partial deletion in the inner E2 core domain of the branched chain alpha-keto acid dehydrogenase complex due to aberrant splicing. A single base deletion at a 5'-splice donor site of an intron of the E2 gene disrupts the consensus sequence in this region. J Clin Invest. 1991 Apr;87(4):1207–1211. doi: 10.1172/JCI115120. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Padgett R. A., Grabowski P. J., Konarska M. M., Seiler S., Sharp P. A. Splicing of messenger RNA precursors. Annu Rev Biochem. 1986;55:1119–1150. doi: 10.1146/annurev.bi.55.070186.005351. [DOI] [PubMed] [Google Scholar]
  22. Reed R., Maniatis T. A role for exon sequences and splice-site proximity in splice-site selection. Cell. 1986 Aug 29;46(5):681–690. doi: 10.1016/0092-8674(86)90343-0. [DOI] [PubMed] [Google Scholar]
  23. Roche T. E., Reed L. J. Function of the nonidentical subunits of mammalian pyruvate dehydrogenase. Biochem Biophys Res Commun. 1972 Aug 21;48(4):840–846. doi: 10.1016/0006-291x(72)90684-5. [DOI] [PubMed] [Google Scholar]
  24. Somasekhar M. B., Mertz J. E. Exon mutations that affect the choice of splice sites used in processing the SV40 late transcripts. Nucleic Acids Res. 1985 Aug 12;13(15):5591–5609. doi: 10.1093/nar/13.15.5591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wynn R. M., Chuang J. L., Davie J. R., Fisher C. W., Hale M. A., Cox R. P., Chuang D. T. Cloning and expression in Escherichia coli of mature E1 beta subunit of bovine mitochondrial branched-chain alpha-keto acid dehydrogenase complex. Mapping of the E1 beta-binding region on E2. J Biol Chem. 1992 Jan 25;267(3):1881–1887. [PubMed] [Google Scholar]
  26. Yeaman S. J. The 2-oxo acid dehydrogenase complexes: recent advances. Biochem J. 1989 Feb 1;257(3):625–632. doi: 10.1042/bj2570625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Zneimer S. M., Lau K. S., Eddy R. L., Shows T. B., Chuang J. L., Chuang D. T., Cox R. P. Regional assignment of two genes of the human branched-chain alpha-keto acid dehydrogenase complex: the E1 beta gene (BCKDHB) to chromosome 6p21-22 and the E2 gene (DBT) to chromosome 1p31. Genomics. 1991 Jul;10(3):740–747. doi: 10.1016/0888-7543(91)90458-q. [DOI] [PubMed] [Google Scholar]

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