Abstract
In the yeast Saccharomyces cerevisiae, the phospholipid biosynthetic genes are highly regulated at the transcriptional level in response to the phospholipid precursors inositol and choline. In the absence of inositol and choline (derepressing), the products of the INO2 and INO4 genes form a heteromeric complex which binds to a 10-bp element, upstream activation sequence INO (UASINO), in the promoters of the phospholipid biosynthetic genes to activate their transcription. In the presence of inositol and choline (repressing), the product of the OPI1 gene represses transcription dictated by the UASINO element. Curiously, we identified a UASINO-like element in the promoters of both the INO2 and INO4 genes. The presence of the UASINO element in these two promoters suggested that the mechanism for the inositol-choline response would involved regulating expression of the two activator genes. Using a cat reporter gene, we find that INO2-cat expression was regulated 12-fold in response to inositol and choline but that INO4-cat was constitutively expressed. We further observed that INO2-cat was not expressed in either an ino2 or an ino4 mutant strain and was constitutively overexpressed in an opi1 mutant strain. Expression of the INO4-cat gene was affected only by mutation in the INO4 gene itself. Therefore, INO2-cat transcription is regulated by the products of both the INO2 and INO4 genes whereas INO4 must interact with another protein to activate its own transcription. Our data show that derepression of phospholipid biosynthetic gene expression involves two mechanisms: increasing the levels of the INO2 and INO4 gene products and inactivating the OPI1-mediated repression mechanism. We propose a model suggesting that this dual mechanism of regulation accounts for the observed cooperative stimulation of IN01 and CH01 gene expression (phospholipids biosynthetic genes).
Full Text
The Full Text of this article is available as a PDF (285.3 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Amati B., Land H. Myc-Max-Mad: a transcription factor network controlling cell cycle progression, differentiation and death. Curr Opin Genet Dev. 1994 Feb;4(1):102–108. doi: 10.1016/0959-437x(94)90098-1. [DOI] [PubMed] [Google Scholar]
- Ambroziak J., Henry S. A. INO2 and INO4 gene products, positive regulators of phospholipid biosynthesis in Saccharomyces cerevisiae, form a complex that binds to the INO1 promoter. J Biol Chem. 1994 May 27;269(21):15344–15349. [PubMed] [Google Scholar]
- Bailis A. M., Lopes J. M., Kohlwein S. D., Henry S. A. Cis and trans regulatory elements required for regulation of the CHO1 gene of Saccharomyces cerevisiae. Nucleic Acids Res. 1992 Mar 25;20(6):1411–1418. doi: 10.1093/nar/20.6.1411. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bailis A. M., Poole M. A., Carman G. M., Henry S. A. The membrane-associated enzyme phosphatidylserine synthase is regulated at the level of mRNA abundance. Mol Cell Biol. 1987 Jan;7(1):167–176. doi: 10.1128/mcb.7.1.167. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cai M., Davis R. W. Yeast centromere binding protein CBF1, of the helix-loop-helix protein family, is required for chromosome stability and methionine prototrophy. Cell. 1990 May 4;61(3):437–446. doi: 10.1016/0092-8674(90)90525-j. [DOI] [PubMed] [Google Scholar]
- Caponigro G., Muhlrad D., Parker R. A small segment of the MAT alpha 1 transcript promotes mRNA decay in Saccharomyces cerevisiae: a stimulatory role for rare codons. Mol Cell Biol. 1993 Sep;13(9):5141–5148. doi: 10.1128/mcb.13.9.5141. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Carman G. M., Henry S. A. Phospholipid biosynthesis in yeast. Annu Rev Biochem. 1989;58:635–669. doi: 10.1146/annurev.bi.58.070189.003223. [DOI] [PubMed] [Google Scholar]
- Culbertson M. R., Donahue T. F., Henry S. A. Control of inositol biosynthesis in Saccharomyces cerevisiae: properties of a repressible enzyme system in extracts of wild-type (Ino+) cells. J Bacteriol. 1976 Apr;126(1):232–242. doi: 10.1128/jb.126.1.232-242.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Donahue T. F., Henry S. A. myo-Inositol-1-phosphate synthase. Characteristics of the enzyme and identification of its structural gene in yeast. J Biol Chem. 1981 Jul 10;256(13):7077–7085. [PubMed] [Google Scholar]
- Elion E. A., Warner J. R. The major promoter element of rRNA transcription in yeast lies 2 kb upstream. Cell. 1984 Dec;39(3 Pt 2):663–673. doi: 10.1016/0092-8674(84)90473-2. [DOI] [PubMed] [Google Scholar]
- Gill G., Ptashne M. Negative effect of the transcriptional activator GAL4. Nature. 1988 Aug 25;334(6184):721–724. doi: 10.1038/334721a0. [DOI] [PubMed] [Google Scholar]
- Giniger E., Ptashne M. Cooperative DNA binding of the yeast transcriptional activator GAL4. Proc Natl Acad Sci U S A. 1988 Jan;85(2):382–386. doi: 10.1073/pnas.85.2.382. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Griggs D. W., Johnston M. Promoter elements determining weak expression of the GAL4 regulatory gene of Saccharomyces cerevisiae. Mol Cell Biol. 1993 Aug;13(8):4999–5009. doi: 10.1128/mcb.13.8.4999. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Griggs D. W., Johnston M. Regulated expression of the GAL4 activator gene in yeast provides a sensitive genetic switch for glucose repression. Proc Natl Acad Sci U S A. 1991 Oct 1;88(19):8597–8601. doi: 10.1073/pnas.88.19.8597. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guarente L. Regulatory proteins in yeast. Annu Rev Genet. 1987;21:425–452. doi: 10.1146/annurev.ge.21.120187.002233. [DOI] [PubMed] [Google Scholar]
- Hinnebusch A. G. Evidence for translational regulation of the activator of general amino acid control in yeast. Proc Natl Acad Sci U S A. 1984 Oct;81(20):6442–6446. doi: 10.1073/pnas.81.20.6442. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hirsch J. P., Henry S. A. Expression of the Saccharomyces cerevisiae inositol-1-phosphate synthase (INO1) gene is regulated by factors that affect phospholipid synthesis. Mol Cell Biol. 1986 Oct;6(10):3320–3328. doi: 10.1128/mcb.6.10.3320. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Homann M. J., Henry S. A., Carman G. M. Regulation of CDP-diacylglycerol synthase activity in Saccharomyces cerevisiae. J Bacteriol. 1985 Sep;163(3):1265–1266. doi: 10.1128/jb.163.3.1265-1266.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hoshizaki D. K., Hill J. E., Henry S. A. The Saccharomyces cerevisiae INO4 gene encodes a small, highly basic protein required for derepression of phospholipid biosynthetic enzymes. J Biol Chem. 1990 Mar 15;265(8):4736–4745. [PubMed] [Google Scholar]
- Hudak K. A., Lopes J. M., Henry S. A. A pleiotropic phospholipid biosynthetic regulatory mutation in Saccharomyces cerevisiae is allelic to sin3 (sdi1, ume4, rpd1). Genetics. 1994 Feb;136(2):475–483. doi: 10.1093/genetics/136.2.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Igarashi M., Segawa T., Nogi Y., Suzuki Y., Fukasawa T. Autogenous regulation of the Saccharomyces cerevisiae regulatory gene GAL80. Mol Gen Genet. 1987 May;207(2-3):273–279. doi: 10.1007/BF00331589. [DOI] [PubMed] [Google Scholar]
- Johnson P. F., McKnight S. L. Eukaryotic transcriptional regulatory proteins. Annu Rev Biochem. 1989;58:799–839. doi: 10.1146/annurev.bi.58.070189.004055. [DOI] [PubMed] [Google Scholar]
- Johnston M. A model fungal gene regulatory mechanism: the GAL genes of Saccharomyces cerevisiae. Microbiol Rev. 1987 Dec;51(4):458–476. doi: 10.1128/mr.51.4.458-476.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kelley M. J., Bailis A. M., Henry S. A., Carman G. M. Regulation of phospholipid biosynthesis in Saccharomyces cerevisiae by inositol. Inositol is an inhibitor of phosphatidylserine synthase activity. J Biol Chem. 1988 Dec 5;263(34):18078–18085. [PubMed] [Google Scholar]
- Landschulz W. H., Johnson P. F., McKnight S. L. The DNA binding domain of the rat liver nuclear protein C/EBP is bipartite. Science. 1989 Mar 31;243(4899):1681–1688. doi: 10.1126/science.2494700. [DOI] [PubMed] [Google Scholar]
- Leuther K. K., Johnston S. A. Nondissociation of GAL4 and GAL80 in vivo after galactose induction. Science. 1992 May 29;256(5061):1333–1335. doi: 10.1126/science.1598579. [DOI] [PubMed] [Google Scholar]
- Li Z., Brendel M. Co-regulation with genes of phospholipid biosynthesis of the CTR/HNM1-encoded choline/nitrogen mustard permease in Saccharomyces cerevisiae. Mol Gen Genet. 1993 Dec;241(5-6):680–684. doi: 10.1007/BF00279911. [DOI] [PubMed] [Google Scholar]
- Liao X., Butow R. A. RTG1 and RTG2: two yeast genes required for a novel path of communication from mitochondria to the nucleus. Cell. 1993 Jan 15;72(1):61–71. doi: 10.1016/0092-8674(93)90050-z. [DOI] [PubMed] [Google Scholar]
- Lopes J. M., Henry S. A. Interaction of trans and cis regulatory elements in the INO1 promoter of Saccharomyces cerevisiae. Nucleic Acids Res. 1991 Jul 25;19(14):3987–3994. doi: 10.1093/nar/19.14.3987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lopes J. M., Hirsch J. P., Chorgo P. A., Schulze K. L., Henry S. A. Analysis of sequences in the INO1 promoter that are involved in its regulation by phospholipid precursors. Nucleic Acids Res. 1991 Apr 11;19(7):1687–1693. doi: 10.1093/nar/19.7.1687. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Maniatis T., Goodbourn S., Fischer J. A. Regulation of inducible and tissue-specific gene expression. Science. 1987 Jun 5;236(4806):1237–1245. doi: 10.1126/science.3296191. [DOI] [PubMed] [Google Scholar]
- Mitchell P. J., Tjian R. Transcriptional regulation in mammalian cells by sequence-specific DNA binding proteins. Science. 1989 Jul 28;245(4916):371–378. doi: 10.1126/science.2667136. [DOI] [PubMed] [Google Scholar]
- Morimoto R. I. Transcription factors: positive and negative regulators of cell growth and disease. Curr Opin Cell Biol. 1992 Jun;4(3):480–487. doi: 10.1016/0955-0674(92)90015-5. [DOI] [PubMed] [Google Scholar]
- Murre C., McCaw P. S., Baltimore D. A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell. 1989 Mar 10;56(5):777–783. doi: 10.1016/0092-8674(89)90682-x. [DOI] [PubMed] [Google Scholar]
- Nikoloff D. M., Henry S. A. Functional characterization of the INO2 gene of Saccharomyces cerevisiae. A positive regulator of phospholipid biosynthesis. J Biol Chem. 1994 Mar 11;269(10):7402–7411. [PubMed] [Google Scholar]
- Nikoloff D. M., Henry S. A. Genetic analysis of yeast phospholipid biosynthesis. Annu Rev Genet. 1991;25:559–583. doi: 10.1146/annurev.ge.25.120191.003015. [DOI] [PubMed] [Google Scholar]
- Nikoloff D. M., McGraw P., Henry S. A. The INO2 gene of Saccharomyces cerevisiae encodes a helix-loop-helix protein that is required for activation of phospholipid synthesis. Nucleic Acids Res. 1992 Jun 25;20(12):3253–3253. doi: 10.1093/nar/20.12.3253. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Yeast transformation: a model system for the study of recombination. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6354–6358. doi: 10.1073/pnas.78.10.6354. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pabo C. O., Sauer R. T. Transcription factors: structural families and principles of DNA recognition. Annu Rev Biochem. 1992;61:1053–1095. doi: 10.1146/annurev.bi.61.070192.005201. [DOI] [PubMed] [Google Scholar]
- Pierrat B., Lacroute F., Losson R. The 5' untranslated region of the PPR1 regulatory gene dictates rapid mRNA decay in yeast. Gene. 1993 Sep 6;131(1):43–51. doi: 10.1016/0378-1119(93)90667-r. [DOI] [PubMed] [Google Scholar]
- Schmid A., Fascher K. D., Hörz W. Nucleosome disruption at the yeast PHO5 promoter upon PHO5 induction occurs in the absence of DNA replication. Cell. 1992 Nov 27;71(5):853–864. doi: 10.1016/0092-8674(92)90560-y. [DOI] [PubMed] [Google Scholar]
- Schüller H. J., Schorr R., Hoffmann B., Schweizer E. Regulatory gene INO4 of yeast phospholipid biosynthesis is positively autoregulated and functions as a transactivator of fatty acid synthase genes FAS1 and FAS2 from Saccharomyces cerevisiae. Nucleic Acids Res. 1992 Nov 25;20(22):5955–5961. doi: 10.1093/nar/20.22.5955. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seed B., Sheen J. Y. A simple phase-extraction assay for chloramphenicol acyltransferase activity. Gene. 1988 Jul 30;67(2):271–277. doi: 10.1016/0378-1119(88)90403-9. [DOI] [PubMed] [Google Scholar]
- Shimada H., Fukasawa T. Controlled transcription of the yeast regulatory gene GAL80. Gene. 1985;39(1):1–9. doi: 10.1016/0378-1119(85)90100-3. [DOI] [PubMed] [Google Scholar]
- Solomon D. L., Amati B., Land H. Distinct DNA binding preferences for the c-Myc/Max and Max/Max dimers. Nucleic Acids Res. 1993 Nov 25;21(23):5372–5376. doi: 10.1093/nar/21.23.5372. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Struhl G., Struhl K., Macdonald P. M. The gradient morphogen bicoid is a concentration-dependent transcriptional activator. Cell. 1989 Jun 30;57(7):1259–1273. doi: 10.1016/0092-8674(89)90062-7. [DOI] [PubMed] [Google Scholar]
- Struhl K. Molecular mechanisms of transcriptional regulation in yeast. Annu Rev Biochem. 1989;58:1051–1077. doi: 10.1146/annurev.bi.58.070189.005155. [DOI] [PubMed] [Google Scholar]
- White M. J., Hirsch J. P., Henry S. A. The OPI1 gene of Saccharomyces cerevisiae, a negative regulator of phospholipid biosynthesis, encodes a protein containing polyglutamine tracts and a leucine zipper. J Biol Chem. 1991 Jan 15;266(2):863–872. [PubMed] [Google Scholar]
- White M. J., Lopes J. M., Henry S. A. Inositol metabolism in yeasts. Adv Microb Physiol. 1991;32:1–51. doi: 10.1016/s0065-2911(08)60004-1. [DOI] [PubMed] [Google Scholar]