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. 1999 Mar;64(3):698–705. doi: 10.1086/302295

Human molybdopterin synthase gene: identification of a bicistronic transcript with overlapping reading frames.

B Stallmeyer 1, G Drugeon 1, J Reiss 1, A L Haenni 1, R R Mendel 1
PMCID: PMC1377786  PMID: 10053003

Abstract

A universal molybdenum-containing cofactor (MoCo) is essential for the activity of all human molybdoenzymes, including sulphite oxidase. The free cofactor is highly unstable, and all organisms share a similar biosynthetic pathway. The involved enzymes exhibit homologies, even between bacteria and humans. We have exploited these homologies to isolate a cDNA for the heterodimeric molybdopterin (MPT)-synthase. This enzyme is necessary for the conversion of an unstable precursor into molybdopterin, the organic moiety of MoCo. The corresponding transcript shows a bicistronic structure, encoding the small and large subunits of the MPT-synthase in two different open reading frames (ORFs) that overlap by 77 nucleotides. In various human tissues, only one size of mRNA coinciding with the bicistronic transcript was detected. In vitro translation and mutagenesis experiments demonstrated that each ORF is translated independently, leading to the synthesis of a 10-kDa protein and a 21-kDa protein for the small and large subunits, respectively, and indicated that the 3'-proximal ORF of the bicistronic transcript is translated by leaky scanning.

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Selected References

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  1. Amy C. M., Williams-Ahlf B., Naggert J., Smith S. Intron-exon organization of the gene for the multifunctional animal fatty acid synthase. Proc Natl Acad Sci U S A. 1992 Feb 1;89(3):1105–1108. doi: 10.1073/pnas.89.3.1105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Andrews J., Smith M., Merakovsky J., Coulson M., Hannan F., Kelly L. E. The stoned locus of Drosophila melanogaster produces a dicistronic transcript and encodes two distinct polypeptides. Genetics. 1996 Aug;143(4):1699–1711. doi: 10.1093/genetics/143.4.1699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Davidson J. N., Chen K. C., Jamison R. S., Musmanno L. A., Kern C. B. The evolutionary history of the first three enzymes in pyrimidine biosynthesis. Bioessays. 1993 Mar;15(3):157–164. doi: 10.1002/bies.950150303. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Hille Russ. The Mononuclear Molybdenum Enzymes. Chem Rev. 1996 Nov 7;96(7):2757–2816. doi: 10.1021/cr950061t. [DOI] [PubMed] [Google Scholar]
  6. Hoff T., Schnorr K. M., Meyer C., Caboche M. Isolation of two Arabidopsis cDNAs involved in early steps of molybdenum cofactor biosynthesis by functional complementation of Escherichia coli mutants. J Biol Chem. 1995 Mar 17;270(11):6100–6107. doi: 10.1074/jbc.270.11.6100. [DOI] [PubMed] [Google Scholar]
  7. Iizuka N., Chen C., Yang Q., Johannes G., Sarnow P. Cap-independent translation and internal initiation of translation in eukaryotic cellular mRNA molecules. Curr Top Microbiol Immunol. 1995;203:155–177. doi: 10.1007/978-3-642-79663-0_8. [DOI] [PubMed] [Google Scholar]
  8. Jang S. K., Kräusslich H. G., Nicklin M. J., Duke G. M., Palmenberg A. C., Wimmer E. A segment of the 5' nontranslated region of encephalomyocarditis virus RNA directs internal entry of ribosomes during in vitro translation. J Virol. 1988 Aug;62(8):2636–2643. doi: 10.1128/jvi.62.8.2636-2643.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Jobling S. A., Cuthbert C. M., Rogers S. G., Fraley R. T., Gehrke L. In vitro transcription and translational efficiency of chimeric SP6 messenger RNAs devoid of 5' vector nucleotides. Nucleic Acids Res. 1988 May 25;16(10):4483–4498. doi: 10.1093/nar/16.10.4483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Johnson J. L., Wuebbens M. M., Mandell R., Shih V. E. Molybdenum cofactor biosynthesis in humans. Identification of two complementation groups of cofactor-deficient patients and preliminary characterization of a diffusible molybdopterin precursor. J Clin Invest. 1989 Mar;83(3):897–903. doi: 10.1172/JCI113974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kozak M. The scanning model for translation: an update. J Cell Biol. 1989 Feb;108(2):229–241. doi: 10.1083/jcb.108.2.229. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lee S. J. Expression of growth/differentiation factor 1 in the nervous system: conservation of a bicistronic structure. Proc Natl Acad Sci U S A. 1991 May 15;88(10):4250–4254. doi: 10.1073/pnas.88.10.4250. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lennon G., Auffray C., Polymeropoulos M., Soares M. B. The I.M.A.G.E. Consortium: an integrated molecular analysis of genomes and their expression. Genomics. 1996 Apr 1;33(1):151–152. doi: 10.1006/geno.1996.0177. [DOI] [PubMed] [Google Scholar]
  14. Matsufuji S., Matsufuji T., Miyazaki Y., Murakami Y., Atkins J. F., Gesteland R. F., Hayashi S. Autoregulatory frameshifting in decoding mammalian ornithine decarboxylase antizyme. Cell. 1995 Jan 13;80(1):51–60. doi: 10.1016/0092-8674(95)90450-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Mendel R. R. Molybdenum cofactor of higher plants: biosynthesis and molecular biology. Planta. 1997 Dec;203(4):399–405. doi: 10.1007/s004250050206. [DOI] [PubMed] [Google Scholar]
  16. Pelletier J., Sonenberg N. Internal initiation of translation of eukaryotic mRNA directed by a sequence derived from poliovirus RNA. Nature. 1988 Jul 28;334(6180):320–325. doi: 10.1038/334320a0. [DOI] [PubMed] [Google Scholar]
  17. Pitterle D. M., Johnson J. L., Rajagopalan K. V. In vitro synthesis of molybdopterin from precursor Z using purified converting factor. Role of protein-bound sulfur in formation of the dithiolene. J Biol Chem. 1993 Jun 25;268(18):13506–13509. [PubMed] [Google Scholar]
  18. Rajagopalan K. V., Johnson J. L. The pterin molybdenum cofactors. J Biol Chem. 1992 May 25;267(15):10199–10202. [PubMed] [Google Scholar]
  19. Reiss J., Christensen E., Kurlemann G., Zabot M. T., Dorche C. Genomic structure and mutational spectrum of the bicistronic MOCS1 gene defective in molybdenum cofactor deficiency type A. Hum Genet. 1998 Dec;103(6):639–644. doi: 10.1007/s004390050884. [DOI] [PubMed] [Google Scholar]
  20. Reiss J., Cohen N., Dorche C., Mandel H., Mendel R. R., Stallmeyer B., Zabot M. T., Dierks T. Mutations in a polycistronic nuclear gene associated with molybdenum cofactor deficiency. Nat Genet. 1998 Sep;20(1):51–53. doi: 10.1038/1706. [DOI] [PubMed] [Google Scholar]
  21. Reiss J., Dorche C., Stallmeyer B., Mendel R. R., Cohen N., Zabot M. T. Human molybdopterin synthase gene: genomic structure and mutations in molybdenum cofactor deficiency type B. Am J Hum Genet. 1999 Mar;64(3):706–711. doi: 10.1086/302296. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Ritchie H., Wang L. H. A mammalian bicistronic transcript encoding two dentin-specific proteins. Biochem Biophys Res Commun. 1997 Feb 13;231(2):425–428. doi: 10.1006/bbrc.1997.6126. [DOI] [PubMed] [Google Scholar]
  23. Schägger H., von Jagow G. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. Anal Biochem. 1987 Nov 1;166(2):368–379. doi: 10.1016/0003-2697(87)90587-2. [DOI] [PubMed] [Google Scholar]
  24. Stallmeyer B., Nerlich A., Schiemann J., Brinkmann H., Mendel R. R. Molybdenum co-factor biosynthesis: the Arabidopsis thaliana cDNA cnx1 encodes a multifunctional two-domain protein homologous to a mammalian neuroprotein, the insect protein Cinnamon and three Escherichia coli proteins. Plant J. 1995 Nov;8(5):751–762. doi: 10.1046/j.1365-313x.1995.08050751.x. [DOI] [PubMed] [Google Scholar]
  25. Stiefel E. I. Molybdenum bolsters the bioinorganic brigade. Science. 1996 Jun 14;272(5268):1599–1600. doi: 10.1126/science.272.5268.1599. [DOI] [PubMed] [Google Scholar]
  26. Unkles S. E., Smith J., Kanan G. J., Millar L. J., Heck I. S., Boxer D. H., Kinghorn J. R. The Aspergillus nidulans cnxABC locus is a single gene encoding two catalytic domains required for synthesis of precursor Z, an intermediate in molybdenum cofactor biosynthesis. J Biol Chem. 1997 Nov 7;272(45):28381–28390. doi: 10.1074/jbc.272.45.28381. [DOI] [PubMed] [Google Scholar]
  27. Wilson R., Ainscough R., Anderson K., Baynes C., Berks M., Bonfield J., Burton J., Connell M., Copsey T., Cooper J. 2.2 Mb of contiguous nucleotide sequence from chromosome III of C. elegans. Nature. 1994 Mar 3;368(6466):32–38. doi: 10.1038/368032a0. [DOI] [PubMed] [Google Scholar]

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