Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1986 Jul;6(7):2324–2333. doi: 10.1128/mcb.6.7.2324

Short repeated elements in the upstream regulatory region of the SUC2 gene of Saccharomyces cerevisiae.

L Sarokin, M Carlson
PMCID: PMC367785  PMID: 3097512

Abstract

Expression of secreted invertase from the SUC2 gene is regulated by carbon catabolite repression. Previously, an upstream regulatory region that is required for derepression of secreted invertase was identified and shown to confer glucose-repressible expression to the heterologous promoter of a LEU2-lacZ fusion. In this paper we show that tandem copies of a 32-base pair (bp) sequence from the upstream regulatory region activate expression of the same LEU2-lacZ fusion. The level of expression increased with the number of copies of the element, but was independent of their orientation; the expression from constructions containing four copies of the sequence was only twofold lower than that when the entire SUC2 upstream regulatory region was present. This activation was not significantly glucose repressible. The 32-bp sequence includes a 7-bp motif with the consensus sequence (A/C)(A/G)GAAAT that is repeated at five sites within the upstream regulatory region. Genetic evidence supporting the functional significance of this repeated motif was obtained by pseudoreversion of a SUC2 deletion mutant lacking part of the upstream region, including two copies of the 7-bp element. In three of five pseudorevertants, the mutations that restored high-level SUC2 expression altered one of the remaining copies of the 7-bp element.

Full text

PDF
2324

Selected References

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

  1. Andreadis A., Hsu Y. P., Hermodson M., Kohlhaw G., Schimmel P. Yeast LEU2. Repression of mRNA levels by leucine and primary structure of the gene product. J Biol Chem. 1984 Jul 10;259(13):8059–8062. [PubMed] [Google Scholar]
  2. Botstein D., Falco S. C., Stewart S. E., Brennan M., Scherer S., Stinchcomb D. T., Struhl K., Davis R. W. Sterile host yeasts (SHY): a eukaryotic system of biological containment for recombinant DNA experiments. Gene. 1979 Dec;8(1):17–24. doi: 10.1016/0378-1119(79)90004-0. [DOI] [PubMed] [Google Scholar]
  3. Carlson M., Botstein D. Organization of the SUC gene family in Saccharomyces. Mol Cell Biol. 1983 Mar;3(3):351–359. doi: 10.1128/mcb.3.3.351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carlson M., Botstein D. Two differentially regulated mRNAs with different 5' ends encode secreted with intracellular forms of yeast invertase. Cell. 1982 Jan;28(1):145–154. doi: 10.1016/0092-8674(82)90384-1. [DOI] [PubMed] [Google Scholar]
  5. Carlson M., Celenza J. L., Eng F. J. Evolution of the dispersed SUC gene family of Saccharomyces by rearrangements of chromosome telomeres. Mol Cell Biol. 1985 Nov;5(11):2894–2902. doi: 10.1128/mcb.5.11.2894. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Carlson M., Osmond B. C., Botstein D. Mutants of yeast defective in sucrose utilization. Genetics. 1981 May;98(1):25–40. doi: 10.1093/genetics/98.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Carlson M., Osmond B. C., Neigeborn L., Botstein D. A suppressor of SNF1 mutations causes constitutive high-level invertase synthesis in yeast. Genetics. 1984 May;107(1):19–32. doi: 10.1093/genetics/107.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Carlson M., Taussig R., Kustu S., Botstein D. The secreted form of invertase in Saccharomyces cerevisiae is synthesized from mRNA encoding a signal sequence. Mol Cell Biol. 1983 Mar;3(3):439–447. doi: 10.1128/mcb.3.3.439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Celenza J. L., Carlson M. Cloning and genetic mapping of SNF1, a gene required for expression of glucose-repressible genes in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Jan;4(1):49–53. doi: 10.1128/mcb.4.1.49. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Donahue T. F., Daves R. S., Lucchini G., Fink G. R. A short nucleotide sequence required for regulation of HIS4 by the general control system of yeast. Cell. 1983 Jan;32(1):89–98. doi: 10.1016/0092-8674(83)90499-3. [DOI] [PubMed] [Google Scholar]
  11. Everett R. D., Baty D., Chambon P. The repeated GC-rich motifs upstream from the TATA box are important elements of the SV40 early promoter. Nucleic Acids Res. 1983 Apr 25;11(8):2447–2464. doi: 10.1093/nar/11.8.2447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fromm M., Berg P. Deletion mapping of DNA regions required for SV40 early region promoter function in vivo. J Mol Appl Genet. 1982;1(5):457–481. [PubMed] [Google Scholar]
  13. Giniger E., Varnum S. M., Ptashne M. Specific DNA binding of GAL4, a positive regulatory protein of yeast. Cell. 1985 Apr;40(4):767–774. doi: 10.1016/0092-8674(85)90336-8. [DOI] [PubMed] [Google Scholar]
  14. Goff C. G., Moir D. T., Kohno T., Gravius T. C., Smith R. A., Yamasaki E., Taunton-Rigby A. Expression of calf prochymosin in Saccharomyces cerevisiae. Gene. 1984 Jan;27(1):35–46. doi: 10.1016/0378-1119(84)90236-1. [DOI] [PubMed] [Google Scholar]
  15. Goldstein A., Lampen J. O. Beta-D-fructofuranoside fructohydrolase from yeast. Methods Enzymol. 1975;42:504–511. doi: 10.1016/0076-6879(75)42159-0. [DOI] [PubMed] [Google Scholar]
  16. Guarente L., Lalonde B., Gifford P., Alani E. Distinctly regulated tandem upstream activation sites mediate catabolite repression of the CYC1 gene of S. cerevisiae. Cell. 1984 Feb;36(2):503–511. doi: 10.1016/0092-8674(84)90243-5. [DOI] [PubMed] [Google Scholar]
  17. Guarente L. Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. Methods Enzymol. 1983;101:181–191. doi: 10.1016/0076-6879(83)01013-7. [DOI] [PubMed] [Google Scholar]
  18. Hearing P., Shenk T. The adenovirus type 5 E1A transcriptional control region contains a duplicated enhancer element. Cell. 1983 Jul;33(3):695–703. doi: 10.1016/0092-8674(83)90012-0. [DOI] [PubMed] [Google Scholar]
  19. Hinnebusch A. G., Lucchini G., Fink G. R. A synthetic HIS4 regulatory element confers general amino acid control on the cytochrome c gene (CYC1) of yeast. Proc Natl Acad Sci U S A. 1985 Jan;82(2):498–502. doi: 10.1073/pnas.82.2.498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hinnen A., Hicks J. B., Fink G. R. Transformation of yeast. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1929–1933. doi: 10.1073/pnas.75.4.1929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Khoury G., Gruss P. Enhancer elements. Cell. 1983 Jun;33(2):313–314. doi: 10.1016/0092-8674(83)90410-5. [DOI] [PubMed] [Google Scholar]
  22. Martinez-Arias A., Yost H. J., Casadaban M. J. Role of an upstream regulatory element in leucine repression of the Saccharomyces cerevisiae leu2 gene. Nature. 1984 Feb 23;307(5953):740–742. doi: 10.1038/307740b0. [DOI] [PubMed] [Google Scholar]
  23. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  24. McKnight S. L. Functional relationships between transcriptional control signals of the thymidine kinase gene of herpes simplex virus. Cell. 1982 Dec;31(2 Pt 1):355–365. doi: 10.1016/0092-8674(82)90129-5. [DOI] [PubMed] [Google Scholar]
  25. Miller A. M., MacKay V. L., Nasmyth K. A. Identification and comparison of two sequence elements that confer cell-type specific transcription in yeast. Nature. 1985 Apr 18;314(6012):598–603. doi: 10.1038/314598a0. [DOI] [PubMed] [Google Scholar]
  26. Neigeborn L., Carlson M. Genes affecting the regulation of SUC2 gene expression by glucose repression in Saccharomyces cerevisiae. Genetics. 1984 Dec;108(4):845–858. doi: 10.1093/genetics/108.4.845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Neigeborn L., Rubin K., Carlson M. Suppressors of SNF2 mutations restore invertase derepression and cause temperature-sensitive lethality in yeast. Genetics. 1986 Apr;112(4):741–753. doi: 10.1093/genetics/112.4.741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  29. Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Genetic applications of yeast transformation with linear and gapped plasmids. Methods Enzymol. 1983;101:228–245. doi: 10.1016/0076-6879(83)01017-4. [DOI] [PubMed] [Google Scholar]
  30. Perlman D., Halvorson H. O., Cannon L. E. Presecretory and cytoplasmic invertase polypeptides encoded by distinct mRNAs derived from the same structural gene differ by a signal sequence. Proc Natl Acad Sci U S A. 1982 Feb;79(3):781–785. doi: 10.1073/pnas.79.3.781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Russell D. W., Smith M., Cox D., Williamson V. M., Young E. T. DNA sequences of two yeast promoter-up mutants. Nature. 1983 Aug 18;304(5927):652–654. doi: 10.1038/304652a0. [DOI] [PubMed] [Google Scholar]
  32. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sarokin L., Carlson M. Comparison of two yeast invertase genes: conservation of the upstream regulatory region. Nucleic Acids Res. 1985 Sep 11;13(17):6089–6103. doi: 10.1093/nar/13.17.6089. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sarokin L., Carlson M. Upstream region of the SUC2 gene confers regulated expression to a heterologous gene in Saccharomyces cerevisiae. Mol Cell Biol. 1985 Oct;5(10):2521–2526. doi: 10.1128/mcb.5.10.2521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sarokin L., Carlson M. Upstream region required for regulated expression of the glucose-repressible SUC2 gene of Saccharomyces cerevisiae. Mol Cell Biol. 1984 Dec;4(12):2750–2757. doi: 10.1128/mcb.4.12.2750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  37. Struhl K. Naturally occurring poly(dA-dT) sequences are upstream promoter elements for constitutive transcription in yeast. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8419–8423. doi: 10.1073/pnas.82.24.8419. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES