Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1988 Oct;85(19):7336–7340. doi: 10.1073/pnas.85.19.7336

Heterogeneous expression of protein and mRNA in pyruvate dehydrogenase deficiency.

I D Wexler 1, D S Kerr 1, L Ho 1, M M Lusk 1, R A Pepin 1, A A Javed 1, J E Mole 1, B W Jesse 1, T J Thekkumkara 1, G Pons 1, et al.
PMCID: PMC282181  PMID: 3140238

Abstract

Deficiency of pyruvate dehydrogenase [pyruvate:lipoamide 2-oxidoreductase (decarboxylating and acceptor-acetylating), EC 1.2.4.1], the first component of the pyruvate dehydrogenase complex, is associated with lactic acidosis and central nervous system dysfunction. Using both specific antibodies to pyruvate dehydrogenase and cDNAs coding for its two alpha and beta subunits, we characterized pyruvate dehydrogenase deficiency in 11 patients. Three different patterns were found on immunologic and RNA blot analyses. (i) Seven patients had immunologically detectable crossreactive material for the alpha and beta proteins of pyruvate dehydrogenase. (ii) Two patients had no detectable crossreactive protein for either the alpha or beta subunit but had normal amounts of mRNA for both alpha and beta subunits. (iii) The remaining two patients also had no detectable crossreactive protein but had diminished amounts of mRNA for the alpha subunit of pyruvate dehydrogenase only. These results indicate that loss of pyruvate dehydrogenase activity may be associated with either absent or catalytically inactive proteins, and in those cases in which this enzyme is absent, mRNA for one of the subunits may also be missing. When mRNA for one of the subunits is lacking, both protein subunits are absent, suggesting that a mutation affecting the expression of one of the subunit proteins causes the remaining uncomplexed subunit to be unstable. The results show that several different mutations account for the molecular heterogeneity of pyruvate dehydrogenase deficiency.

Full text

PDF
7336

Images in this article

7339Image
on p.7339
7339Image
on p.7339

Selected References

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

  1. Atkin B. M., Utter M. F., Weinberg M. B. Pyruvate carboxylase and phosphoenolpyruvate carboxykinase activity in leukocytes and fibroblasts from a patient with pyruvate carboxylase deficiency. Pediatr Res. 1979 Jan;13(1):38–43. doi: 10.1203/00006450-197901000-00009. [DOI] [PubMed] [Google Scholar]
  2. Barrera C. R., Namihira G., Hamilton L., Munk P., Eley M. H., Linn T. C., Reed L. J. -Keto acid dehydrogenase complexes. XVI. Studies on the subunit structure of the pyruvate dehydrogenase complexes from bovine kidney and heart. Arch Biochem Biophys. 1972 Feb;148(2):343–358. doi: 10.1016/0003-9861(72)90152-x. [DOI] [PubMed] [Google Scholar]
  3. Bhown A. S., Bennett J. C., Morgan P. H., Mole J. E. Use of fluorescamine as an effective blocking reagent to reduce the background in protein sequence analyses by the Beckman automated sequencer. Anal Biochem. 1981 Mar 15;112(1):158–162. doi: 10.1016/0003-2697(81)90274-8. [DOI] [PubMed] [Google Scholar]
  4. Blass J. P., Avigan J., Uhlendorf B. W. A defect in pyruvate decarboxylase in a child with an intermittent movement disorder. J Clin Invest. 1970 Mar;49(3):423–432. doi: 10.1172/JCI106251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. 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]
  7. Dahl H. H., Hunt S. M., Hutchison W. M., Brown G. K. The human pyruvate dehydrogenase complex. Isolation of cDNA clones for the E1 alpha subunit, sequence analysis, and characterization of the mRNA. J Biol Chem. 1987 May 25;262(15):7398–7403. [PubMed] [Google Scholar]
  8. De Meirleir L., MacKay N., Lam Hon Wah A. M., Robinson B. H. Isolation of a full-length complementary DNA coding for human E1 alpha subunit of the pyruvate dehydrogenase complex. J Biol Chem. 1988 Feb 5;263(4):1991–1995. [PubMed] [Google Scholar]
  9. Falk R. E., Cederbaum S. D., Blass J. P., Gibson G. E., Kark R. A., Carrel R. E. Ketonic diet in the management of pyruvate dehydrogenase deficiency. Pediatrics. 1976 Nov;58(5):713–721. [PubMed] [Google Scholar]
  10. Feinberg A. P., Vogelstein B. "A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity". Addendum. Anal Biochem. 1984 Feb;137(1):266–267. doi: 10.1016/0003-2697(84)90381-6. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Ho L., Hu C. W., Packman S., Patel M. S. Deficiency of the pyruvate dehydrogenase component in pyruvate dehydrogenase complex-deficient human fibroblasts. Immunological identification. J Clin Invest. 1986 Sep;78(3):844–847. doi: 10.1172/JCI112651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Ho L., Javed A. A., Pepin R. A., Thekkumkara T. J., Raefsky C., Mole J. E., Caliendo A. M., Kwon M. S., Kerr D. S., Patel M. S. Identification of a cDNA clone for the beta-subunit of the pyruvate dehydrogenase component of human pyruvate dehydrogenase complex. Biochem Biophys Res Commun. 1988 Feb 15;150(3):904–908. doi: 10.1016/0006-291x(88)90714-0. [DOI] [PubMed] [Google Scholar]
  14. Hod Y., Morris S. M., Hanson R. W. Induction by cAMP of the mRNA encoding the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) from the chicken. Identification and characterization of a cDNA clone for the enzyme. J Biol Chem. 1984 Dec 25;259(24):15603–15608. [PubMed] [Google Scholar]
  15. Jilka J. M., Rahmatullah M., Kazemi M., Roche T. E. Properties of a newly characterized protein of the bovine kidney pyruvate dehydrogenase complex. J Biol Chem. 1986 Feb 5;261(4):1858–1867. [PubMed] [Google Scholar]
  16. Johnston K., Newth C. J., Sheu K. F., Patel M. S., Heldt G. P., Schmidt K. A., Packman S. Central hypoventilation syndrome in pyruvate dehydrogenase complex deficiency. Pediatrics. 1984 Dec;74(6):1034–1040. [PubMed] [Google Scholar]
  17. Kerr D. S., Berry S. A., Lusk M. M., Ho L., Patel M. S. A deficiency of both subunits of pyruvate dehydrogenase which is not expressed in fibroblasts. Pediatr Res. 1988 Jul;24(1):95–100. doi: 10.1203/00006450-198807000-00022. [DOI] [PubMed] [Google Scholar]
  18. Kerr D. S., Ho L., Berlin C. M., Lanoue K. F., Towfighi J., Hoppel C. L., Lusk M. M., Gondek C. M., Patel M. S. Systemic deficiency of the first component of the pyruvate dehydrogenase complex. Pediatr Res. 1987 Sep;22(3):312–318. doi: 10.1203/00006450-198709000-00015. [DOI] [PubMed] [Google Scholar]
  19. Koike K., Ohta S., Urata Y., Kagawa Y., Koike M. Cloning and sequencing of cDNAs encoding alpha and beta subunits of human pyruvate dehydrogenase. Proc Natl Acad Sci U S A. 1988 Jan;85(1):41–45. doi: 10.1073/pnas.85.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kretzschmar H. A., DeArmond S. J., Koch T. K., Patel M. S., Newth C. J., Schmidt K. A., Packman S. Pyruvate dehydrogenase complex deficiency as a cause of subacute necrotizing encephalopathy (Leigh disease). Pediatrics. 1987 Mar;79(3):370–373. [PubMed] [Google Scholar]
  21. McKay N., Petrova-Benedict R., Thoene J., Bergen B., Wilson W., Robinson B. Lacticacidaemia due to pyruvate dehydrogenase deficiency, with evidence of protein polymorphism in the alpha-subunit of the enzyme. Eur J Pediatr. 1986 Feb;144(5):445–450. doi: 10.1007/BF00441736. [DOI] [PubMed] [Google Scholar]
  22. Morgan D. G., Routtenberg A. Evidence that a 41,000 dalton brain phosphoprotein is pyruvate dehydrogenase. Biochem Biophys Res Commun. 1980 Jul 31;95(2):569–576. doi: 10.1016/0006-291x(80)90822-0. [DOI] [PubMed] [Google Scholar]
  23. Otulakowski G., Robinson B. H. Isolation and sequence determination of cDNA clones for porcine and human lipoamide dehydrogenase. Homology to other disulfide oxidoreductases. J Biol Chem. 1987 Dec 25;262(36):17313–17318. [PubMed] [Google Scholar]
  24. Pons G., Raefsky-Estrin C., Carothers D. J., Pepin R. A., Javed A. A., Jesse B. W., Ganapathi M. K., Samols D., Patel M. S. Cloning and cDNA sequence of the dihydrolipoamide dehydrogenase component human alpha-ketoacid dehydrogenase complexes. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1422–1426. doi: 10.1073/pnas.85.5.1422. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Reed L. J. Regulation of mammalian pyruvate dehydrogenase complex by a phosphorylation-dephosphorylation cycle. Curr Top Cell Regul. 1981;18:95–106. doi: 10.1016/b978-0-12-152818-8.50012-8. [DOI] [PubMed] [Google Scholar]
  26. Robinson B. H., MacMillan H., Petrova-Benedict R., Sherwood W. G. Variable clinical presentation in patients with defective E1 component of pyruvate dehydrogenase complex. J Pediatr. 1987 Oct;111(4):525–533. doi: 10.1016/s0022-3476(87)80112-9. [DOI] [PubMed] [Google Scholar]
  27. Robinson B. H., Taylor J., Sherwood W. G. The genetic heterogeneity of lactic acidosis: occurrence of recognizable inborn errors of metabolism in pediatric population with lactic acidosis. Pediatr Res. 1980 Aug;14(8):956–962. doi: 10.1203/00006450-198008000-00013. [DOI] [PubMed] [Google Scholar]
  28. Sheu K. F., Hu C. W., Utter M. F. Pyruvate dehydrogenase complex activity in normal and deficient fibroblasts. J Clin Invest. 1981 May;67(5):1463–1471. doi: 10.1172/JCI110176. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stansbie D., Wallace S. J., Marsac C. Disorders of the pyruvate dehydrogenase complex. J Inherit Metab Dis. 1986;9(2):105–119. doi: 10.1007/BF01799447. [DOI] [PubMed] [Google Scholar]
  30. Thekkumkara T. J., Jesse B. W., Ho L., Raefsky C., Pepin R. A., Javed A. A., Pons G., Patel M. S. Isolation of a cDNA clone for the dihydrolipoamide acetyltransferase component of the human liver pyruvate dehydrogenase complex. Biochem Biophys Res Commun. 1987 Jun 15;145(2):903–907. doi: 10.1016/0006-291x(87)91050-3. [DOI] [PubMed] [Google Scholar]
  31. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Wicking C. A., Scholem R. D., Hunt S. M., Brown G. K. Immunochemical analysis of normal and mutant forms of human pyruvate dehydrogenase. Biochem J. 1986 Oct 1;239(1):89–96. doi: 10.1042/bj2390089. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES