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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
. 1990 Apr;87(8):2916–2920. doi: 10.1073/pnas.87.8.2916

Cloning of three human multifunctional de novo purine biosynthetic genes by functional complementation of yeast mutations.

D Schild 1, A J Brake 1, M C Kiefer 1, D Young 1, P J Barr 1
PMCID: PMC53804  PMID: 2183217

Abstract

Functional complementation of mutations in the yeast Saccharomyces cerevisiae has been used to clone three multifunctional human genes involved in de novo purine biosynthesis. A HepG2 cDNA library constructed in a yeast expression vector was used to transform yeast strains with mutations in adenine biosynthetic genes. Clones were isolated that complement mutations in the yeast ADE2, ADE3, and ADE8 genes. The cDNA that complemented the ade8 (phosphoribosylglycinamide formyltransferase, GART) mutation, also complemented the ade5 (phosphoribosylglycinamide synthetase) and ade7 [phosphoribosylaminoimidazole synthetase (AIRS; also known as PAIS)] mutations, indicating that it is the human trifunctional GART gene. Supporting data include homology between the AIRS and GART domains of this gene and the published sequence of these domains from other organisms, and localization of the cloned gene to human chromosome 21, where the GART gene has been shown to map. The cDNA that complemented ade2 (phosphoribosylaminoimidazole carboxylase) also complemented ade1 (phosphoribosylaminoimidazole succinocarboxamide synthetase), supporting earlier data suggesting that in some organisms these functions are part of a bifunctional protein. The cDNA that complemented ade3 (formyltetrahydrofolate synthetase) is different from the recently isolated human cDNA encoding this enzyme and instead appears to encode a related mitochondrial enzyme.

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Selected References

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

  1. Aruffo A., Seed B. Molecular cloning of a CD28 cDNA by a high-efficiency COS cell expression system. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8573–8577. doi: 10.1073/pnas.84.23.8573. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Birnstiel M. L., Busslinger M., Strub K. Transcription termination and 3' processing: the end is in site! Cell. 1985 Jun;41(2):349–359. doi: 10.1016/s0092-8674(85)80007-6. [DOI] [PubMed] [Google Scholar]
  4. Bélanger C., MacKenzie R. E. Isolation and characterization of cDNA clones encoding the murine NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase. J Biol Chem. 1989 Mar 25;264(9):4837–4843. [PubMed] [Google Scholar]
  5. Carbon J., Ratzkin B., Clarke L., Richardson D. The expression of cloned eukaryotic DNA in prokaryotes. Brookhaven Symp Biol. 1977 May 12;(29):277–296. [PubMed] [Google Scholar]
  6. Colicelli J., Birchmeier C., Michaeli T., O'Neill K., Riggs M., Wigler M. Isolation and characterization of a mammalian gene encoding a high-affinity cAMP phosphodiesterase. Proc Natl Acad Sci U S A. 1989 May;86(10):3599–3603. doi: 10.1073/pnas.86.10.3599. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Collart F. R., Huberman E. Cloning and sequence analysis of the human and Chinese hamster inosine-5'-monophosphate dehydrogenase cDNAs. J Biol Chem. 1988 Oct 25;263(30):15769–15772. [PubMed] [Google Scholar]
  8. Daubner S. C., Young M., Sammons R. D., Courtney L. F., Benkovic S. J. Structural and mechanistic studies on the HeLa and chicken liver proteins that catalyze glycinamide ribonucleotide synthesis and formylation and aminoimidazole ribonucleotide synthesis. Biochemistry. 1986 May 20;25(10):2951–2957. doi: 10.1021/bi00358a033. [DOI] [PubMed] [Google Scholar]
  9. Ebbole D. J., Zalkin H. Cloning and characterization of a 12-gene cluster from Bacillus subtilis encoding nine enzymes for de novo purine nucleotide synthesis. J Biol Chem. 1987 Jun 15;262(17):8274–8287. [PubMed] [Google Scholar]
  10. Fitzhugh A. L., Chabner B. A., Sholar P. W., Baram J., Allegra C. J. Antifolate metabolism and sites of action: implications for the design of new antifolates. NCI Monogr. 1987;(5):153–157. [PubMed] [Google Scholar]
  11. Fuller R. S., Brake A., Thorner J. Yeast prohormone processing enzyme (KEX2 gene product) is a Ca2+-dependent serine protease. Proc Natl Acad Sci U S A. 1989 Mar;86(5):1434–1438. doi: 10.1073/pnas.86.5.1434. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hards R. G., Benkovic S. J., Van Keuren M. L., Graw S. L., Drabkin H. A., Patterson D. Assignment of a third purine biosynthetic gene (glycinamide ribonucleotide transformylase) to human chromosome 21. Am J Hum Genet. 1986 Aug;39(2):179–185. [PMC free article] [PubMed] [Google Scholar]
  13. Henikoff S., Eghtedarzadeh M. K. Conserved arrangement of nested genes at the Drosophila Gart locus. Genetics. 1987 Dec;117(4):711–725. doi: 10.1093/genetics/117.4.711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Henikoff S., Furlong C. E. Sequence of a Drosophila DNA segment that functions in Saccharomyces cerevisiae and its regulation by a yeast promoter. Nucleic Acids Res. 1983 Feb 11;11(3):789–800. doi: 10.1093/nar/11.3.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Henikoff S., Keene M. A., Fechtel K., Fristrom J. W. Gene within a gene: nested Drosophila genes encode unrelated proteins on opposite DNA strands. Cell. 1986 Jan 17;44(1):33–42. doi: 10.1016/0092-8674(86)90482-4. [DOI] [PubMed] [Google Scholar]
  16. Henikoff S. Multifunctional polypeptides for purine de novo synthesis. Bioessays. 1987 Jan;6(1):8–13. doi: 10.1002/bies.950060104. [DOI] [PubMed] [Google Scholar]
  17. Henikoff S., Sloan J. S., Kelly J. D. A Drosophila metabolic gene transcript is alternatively processed. Cell. 1983 Sep;34(2):405–414. doi: 10.1016/0092-8674(83)90374-4. [DOI] [PubMed] [Google Scholar]
  18. Henikoff S., Tatchell K., Hall B. D., Nasmyth K. A. Isolation of a gene from Drosophila by complementation in yeast. Nature. 1981 Jan 1;289(5793):33–37. doi: 10.1038/289033a0. [DOI] [PubMed] [Google Scholar]
  19. Henikoff S. The Saccharomyces cerevisiae ADE5,7 protein is homologous to overlapping Drosophila melanogaster Gart polypeptides. J Mol Biol. 1986 Aug 20;190(4):519–528. doi: 10.1016/0022-2836(86)90238-x. [DOI] [PubMed] [Google Scholar]
  20. Hum D. W., Bell A. W., Rozen R., MacKenzie R. E. Primary structure of a human trifunctional enzyme. Isolation of a cDNA encoding methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase-formyltetrahydrofolate synthetase. J Biol Chem. 1988 Nov 5;263(31):15946–15950. [PubMed] [Google Scholar]
  21. Janssen J. W., Collard J. G., Tulp A., Cox D., Millington-Ward A., Pearson P. Construction and analysis of an EMBL-3 phage library containing partially digested human chromosome 21-specific DNA inserts (15-20 kb). Cytometry. 1986 Sep;7(5):411–417. doi: 10.1002/cyto.990070504. [DOI] [PubMed] [Google Scholar]
  22. Kramer B., Kramer W., Williamson M. S., Fogel S. Heteroduplex DNA correction in Saccharomyces cerevisiae is mismatch specific and requires functional PMS genes. Mol Cell Biol. 1989 Oct;9(10):4432–4440. doi: 10.1128/mcb.9.10.4432. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kronstad J. W., Wang J., Covert S. F., Holden D. W., McKnight G. L., Leong S. A. Isolation of metabolic genes and demonstration of gene disruption in the phytopathogenic fungus Ustilago maydis. Gene. 1989 Jun 30;79(1):97–106. doi: 10.1016/0378-1119(89)90095-4. [DOI] [PubMed] [Google Scholar]
  24. Lee M. G., Nurse P. Complementation used to clone a human homologue of the fission yeast cell cycle control gene cdc2. Nature. 1987 May 7;327(6117):31–35. doi: 10.1038/327031a0. [DOI] [PubMed] [Google Scholar]
  25. Lugo T. G., Handelin B., Killary A. M., Housman D. E., Fournier R. E. Isolation of microcell hybrid clones containing retroviral vector insertions into specific human chromosomes. Mol Cell Biol. 1987 Aug;7(8):2814–2820. doi: 10.1128/mcb.7.8.2814. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. McKnight G. L., Kato H., Upshall A., Parker M. D., Saari G., O'Hara P. J. Identification and molecular analysis of a third Aspergillus nidulans alcohol dehydrogenase gene. EMBO J. 1985 Aug;4(8):2093–2099. doi: 10.1002/j.1460-2075.1985.tb03897.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. McKnight G. L., McConaughy B. L. Selection of functional cDNAs by complementation in yeast. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4412–4416. doi: 10.1073/pnas.80.14.4412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mejia N. R., MacKenzie R. E. NAD-dependent methylenetetrahydrofolate dehydrogenase is expressed by immortal cells. J Biol Chem. 1985 Nov 25;260(27):14616–14620. [PubMed] [Google Scholar]
  29. Mejia N. R., MacKenzie R. E. NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase in transformed cells is a mitochondrial enzyme. Biochem Biophys Res Commun. 1988 Aug 30;155(1):1–6. doi: 10.1016/s0006-291x(88)81040-4. [DOI] [PubMed] [Google Scholar]
  30. Okayama H., Berg P. A cDNA cloning vector that permits expression of cDNA inserts in mammalian cells. Mol Cell Biol. 1983 Feb;3(2):280–289. doi: 10.1128/mcb.3.2.280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Patey C. A., Shaw G. Purification and properties of an enzyme duet, phosphoribosylaminoimidazole carboxylase and phosphoribosylaminoimidazolesuccinocarboxamide synthetase, involved in the biosynthesis of purine nucleotides de novo. Biochem J. 1973 Nov;135(3):543–545. doi: 10.1042/bj1350543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Patterson D., Graw S., Jones C. Demonstration, by somatic cell genetics, of coordinate regulation of genes for two enzymes of purine synthesis assigned to human chromosome 21. Proc Natl Acad Sci U S A. 1981 Jan;78(1):405–409. doi: 10.1073/pnas.78.1.405. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Peri K. G., Belanger C., Mackenzie R. E. Nucleotide sequence of the human NAD-dependent methylene tetrahydrofolate dehydrogenase-cyclohydrolase. Nucleic Acids Res. 1989 Nov 11;17(21):8853–8853. doi: 10.1093/nar/17.21.8853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Shannon K. W., Rabinowitz J. C. Isolation and characterization of the Saccharomyces cerevisiae MIS1 gene encoding mitochondrial C1-tetrahydrofolate synthase. J Biol Chem. 1988 Jun 5;263(16):7717–7725. [PubMed] [Google Scholar]
  35. Sherman F., Stewart J. W., Tsunasawa S. Methionine or not methionine at the beginning of a protein. Bioessays. 1985 Jul;3(1):27–31. doi: 10.1002/bies.950030108. [DOI] [PubMed] [Google Scholar]
  36. Smith J. M., Daum H. A., 3rd Identification and nucleotide sequence of a gene encoding 5'-phosphoribosylglycinamide transformylase in Escherichia coli K12. J Biol Chem. 1987 Aug 5;262(22):10565–10569. [PubMed] [Google Scholar]
  37. Van Keuren M. L., Hart I. M., Kao F. T., Neve R. L., Bruns G. A., Kurnit D. M., Patterson D. A somatic cell hybrid with a single human chromosome 22 corrects the defect in the CHO mutant (Ade-I) lacking adenylosuccinase activity. Cytogenet Cell Genet. 1987;44(2-3):142–147. doi: 10.1159/000132358. [DOI] [PubMed] [Google Scholar]
  38. White J. H., DiMartino J. F., Anderson R. W., Lusnak K., Hilbert D., Fogel S. A DNA sequence conferring high postmeiotic segregation frequency to heterozygous deletions in Saccharomyces cerevisiae is related to sequences associated with eucaryotic recombination hotspots. Mol Cell Biol. 1988 Mar;8(3):1253–1258. doi: 10.1128/mcb.8.3.1253. [DOI] [PMC free article] [PubMed] [Google Scholar]

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