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. 1993 May;13(5):2706–2717. doi: 10.1128/mcb.13.5.2706

Conservation between human and fungal squalene synthetases: similarities in structure, function, and regulation.

G W Robinson 1, Y H Tsay 1, B K Kienzle 1, C A Smith-Monroy 1, R W Bishop 1
PMCID: PMC359645  PMID: 8474436

Abstract

Squalene synthetase (farnesyl diphosphate:farnesyl diphosphate farnesyltransferase; EC 2.5.1.21) is thought to represent a major control point of isoprene and sterol biosynthesis in eukaryotes. We demonstrate structural and functional conservation between the enzymes from humans, a budding yeast (Saccharomyces cerevisiae), and a fission yeast (Schizosaccharomyces pombe). The amino acid sequences of the human and S. pombe proteins deduced from cloned cDNAs were compared to those of the known S. cerevisiae protein. All are predicted to encode C-terminal membrane-spanning proteins of approximately 50 kDa with similar hydropathy profiles. Extensive sequence conservation exists in regions of the enzyme proposed to interact with its prenyl substrates (i.e., two farnesyl diphosphate molecules). Many of the highly conserved regions are also present in phytoene and prephytoene diphosphate synthetases, enzymes which catalyze prenyl substrate condensation reactions analogous to that of squalene synthetase. Expression of cDNA clones encoding S. pombe or hybrid human-S. cerevisiae squalene synthetases reversed the ergosterol requirement of S. cerevisiae cells bearing ERG9 gene disruptions, showing that these enzymes can functionally replace the S. cerevisiae enzyme. Inhibition of sterol synthesis in S. cerevisiae and S. pombe cells or in cultured human fibroblasts by treatment with the 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor lovastatin resulted in elevated levels of squalene synthetase mRNA in all three cell types.

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Selected References

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  1. Agnew W. S. Squalene synthetase. Methods Enzymol. 1985;110:359–373. doi: 10.1016/s0076-6879(85)10094-7. [DOI] [PubMed] [Google Scholar]
  2. Andersson S., Davis D. L., Dahlbäck H., Jörnvall H., Russell D. W. Cloning, structure, and expression of the mitochondrial cytochrome P-450 sterol 26-hydroxylase, a bile acid biosynthetic enzyme. J Biol Chem. 1989 May 15;264(14):8222–8229. [PubMed] [Google Scholar]
  3. Armstrong G. A., Alberti M., Hearst J. E. Conserved enzymes mediate the early reactions of carotenoid biosynthesis in nonphotosynthetic and photosynthetic prokaryotes. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9975–9979. doi: 10.1073/pnas.87.24.9975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bartley G. E., Viitanen P. V., Bacot K. O., Scolnik P. A. A tomato gene expressed during fruit ripening encodes an enzyme of the carotenoid biosynthesis pathway. J Biol Chem. 1992 Mar 15;267(8):5036–5039. [PubMed] [Google Scholar]
  5. Basson M. E., Thorsness M., Finer-Moore J., Stroud R. M., Rine J. Structural and functional conservation between yeast and human 3-hydroxy-3-methylglutaryl coenzyme A reductases, the rate-limiting enzyme of sterol biosynthesis. Mol Cell Biol. 1988 Sep;8(9):3797–3808. doi: 10.1128/mcb.8.9.3797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Biller S. A., Forster C., Gordon E. M., Harrity T., Scott W. A., Ciosek C. P., Jr Isoprenoid (phosphinylmethyl)phosphonates as inhibitors of squalene synthetase. J Med Chem. 1988 Oct;31(10):1869–1871. doi: 10.1021/jm00118a001. [DOI] [PubMed] [Google Scholar]
  7. Brown M. S., Goldstein J. L. Multivalent feedback regulation of HMG CoA reductase, a control mechanism coordinating isoprenoid synthesis and cell growth. J Lipid Res. 1980 Jul;21(5):505–517. [PubMed] [Google Scholar]
  8. Chang T. Y., Limanek J. S. Regulation of cytosolic acetoacetyl coenzyme A thiolase, 3-hydroxy-3-methylglutaryl coenzyme A synthase, 3-hydroxy-3-methylglutaryl coenzyme A reductase, and mevalonate kinase by low density lipoprotein and by 25-hydroxycholesterol in Chinese hamster ovary cells. J Biol Chem. 1980 Aug 25;255(16):7787–7795. [PubMed] [Google Scholar]
  9. 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]
  10. Dogbo O., Laferriére A., D'Harlingue A., Camara B. Carotenoid biosynthesis: Isolation and characterization of a bifunctional enzyme catalyzing the synthesis of phytoene. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7054–7058. doi: 10.1073/pnas.85.19.7054. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Endo A., Kuroda M., Tanzawa K. Competitive inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase by ML-236A and ML-236B fungal metabolites, having hypocholesterolemic activity. FEBS Lett. 1976 Dec 31;72(2):323–326. doi: 10.1016/0014-5793(76)80996-9. [DOI] [PubMed] [Google Scholar]
  12. Fegueur M., Richard L., Charles A. D., Karst F. Isolation and primary structure of the ERG9 gene of Saccharomyces cerevisiae encoding squalene synthetase. Curr Genet. 1991 Nov;20(5):365–372. doi: 10.1007/BF00317063. [DOI] [PubMed] [Google Scholar]
  13. Fikes J. D., Becker D. M., Winston F., Guarente L. Striking conservation of TFIID in Schizosaccharomyces pombe and Saccharomyces cerevisiae. Nature. 1990 Jul 19;346(6281):291–294. doi: 10.1038/346291a0. [DOI] [PubMed] [Google Scholar]
  14. Goldstein J. L., Brown M. S. Regulation of the mevalonate pathway. Nature. 1990 Feb 1;343(6257):425–430. doi: 10.1038/343425a0. [DOI] [PubMed] [Google Scholar]
  15. Haffey M. L., Stevens J. T., Terry B. J., Dorsky D. I., Crumpacker C. S., Wietstock S. M., Ruyechan W. T., Field A. K. Expression of herpes simplex virus type 1 DNA polymerase in Saccharomyces cerevisiae and detection of virus-specific enzyme activity in cell-free lysates. J Virol. 1988 Dec;62(12):4493–4498. doi: 10.1128/jvi.62.12.4493-4498.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jackson M. R., Nilsson T., Peterson P. A. Identification of a consensus motif for retention of transmembrane proteins in the endoplasmic reticulum. EMBO J. 1990 Oct;9(10):3153–3162. doi: 10.1002/j.1460-2075.1990.tb07513.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jennings S. M., Tsay Y. H., Fisch T. M., Robinson G. W. Molecular cloning and characterization of the yeast gene for squalene synthetase. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6038–6042. doi: 10.1073/pnas.88.14.6038. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  19. Köhrer K., Domdey H. Preparation of high molecular weight RNA. Methods Enzymol. 1991;194:398–405. doi: 10.1016/0076-6879(91)94030-g. [DOI] [PubMed] [Google Scholar]
  20. M'Baya B., Fegueur M., Servouse M., Karst F. Regulation of squalene synthetase and squalene epoxidase activities in Saccharomyces cerevisiae. Lipids. 1989 Dec;24(12):1020–1023. doi: 10.1007/BF02544072. [DOI] [PubMed] [Google Scholar]
  21. McKenzie T. L., Jiang G., Straubhaar J. R., Conrad D. G., Shechter I. Molecular cloning, expression, and characterization of the cDNA for the rat hepatic squalene synthase. J Biol Chem. 1992 Oct 25;267(30):21368–21374. [PubMed] [Google Scholar]
  22. Misawa N., Nakagawa M., Kobayashi K., Yamano S., Izawa Y., Nakamura K., Harashima K. Elucidation of the Erwinia uredovora carotenoid biosynthetic pathway by functional analysis of gene products expressed in Escherichia coli. J Bacteriol. 1990 Dec;172(12):6704–6712. doi: 10.1128/jb.172.12.6704-6712.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Molowa D. T., Cimis G. M. Co-ordinate regulation of low-density-lipoprotein receptor and 3-hydroxy-3-methylglutaryl-CoA reductase and synthase gene expression in HepG2 cells. Biochem J. 1989 Jun 15;260(3):731–736. doi: 10.1042/bj2600731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Möller W., Amons R. Phosphate-binding sequences in nucleotide-binding proteins. FEBS Lett. 1985 Jul 1;186(1):1–7. doi: 10.1016/0014-5793(85)81326-0. [DOI] [PubMed] [Google Scholar]
  25. Naumovski L., Friedberg E. C. Saccharomyces cerevisiae RAD2 gene: isolation, subcloning, and partial characterization. Mol Cell Biol. 1984 Feb;4(2):290–295. doi: 10.1128/mcb.4.2.290. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Popják G., Agnew W. S. Squalene synthetase. Mol Cell Biochem. 1979 Oct 15;27(2):97–116. doi: 10.1007/BF00218354. [DOI] [PubMed] [Google Scholar]
  27. Qureshi A. A., Barnes F. J., Semmler E. J., Porter J. W. Biosynthesis of prelycopersene pyrophosphate and lycopersene by squalene synthetase. J Biol Chem. 1973 Apr 25;248(8):2755–2767. [PubMed] [Google Scholar]
  28. Rine J., Hansen W., Hardeman E., Davis R. W. Targeted selection of recombinant clones through gene dosage effects. Proc Natl Acad Sci U S A. 1983 Nov;80(22):6750–6754. doi: 10.1073/pnas.80.22.6750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Rogers S., Wells R., Rechsteiner M. Amino acid sequences common to rapidly degraded proteins: the PEST hypothesis. Science. 1986 Oct 17;234(4774):364–368. doi: 10.1126/science.2876518. [DOI] [PubMed] [Google Scholar]
  30. 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]
  31. Sasiak K., Rilling H. C. Purification to homogeneity and some properties of squalene synthetase. Arch Biochem Biophys. 1988 Feb 1;260(2):622–627. doi: 10.1016/0003-9861(88)90490-0. [DOI] [PubMed] [Google Scholar]
  32. Schafer B. L., Bishop R. W., Kratunis V. J., Kalinowski S. S., Mosley S. T., Gibson K. M., Tanaka R. D. Molecular cloning of human mevalonate kinase and identification of a missense mutation in the genetic disease mevalonic aciduria. J Biol Chem. 1992 Jul 5;267(19):13229–13238. [PubMed] [Google Scholar]
  33. Schiestl R. H., Gietz R. D. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier. Curr Genet. 1989 Dec;16(5-6):339–346. doi: 10.1007/BF00340712. [DOI] [PubMed] [Google Scholar]
  34. Schild D., Brake A. J., Kiefer M. C., Young D., Barr P. J. Cloning of three human multifunctional de novo purine biosynthetic genes by functional complementation of yeast mutations. Proc Natl Acad Sci U S A. 1990 Apr;87(8):2916–2920. doi: 10.1073/pnas.87.8.2916. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sentenac H., Bonneaud N., Minet M., Lacroute F., Salmon J. M., Gaymard F., Grignon C. Cloning and expression in yeast of a plant potassium ion transport system. Science. 1992 May 1;256(5057):663–665. doi: 10.1126/science.1585180. [DOI] [PubMed] [Google Scholar]
  36. Servouse M., Karst F. Regulation of early enzymes of ergosterol biosynthesis in Saccharomyces cerevisiae. Biochem J. 1986 Dec 1;240(2):541–547. doi: 10.1042/bj2400541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sheares B. T., White S. S., Molowa D. T., Chan K., Ding V. D., Kroon P. A., Bostedor R. G., Karkas J. D. Cloning, analysis, and bacterial expression of human farnesyl pyrophosphate synthetase and its regulation in Hep G2 cells. Biochemistry. 1989 Oct 3;28(20):8129–8135. doi: 10.1021/bi00446a025. [DOI] [PubMed] [Google Scholar]
  38. Shechter I., Klinger E., Rucker M. L., Engstrom R. G., Spirito J. A., Islam M. A., Boettcher B. R., Weinstein D. B. Solubilization, purification, and characterization of a truncated form of rat hepatic squalene synthetase. J Biol Chem. 1992 Apr 25;267(12):8628–8635. [PubMed] [Google Scholar]
  39. Suzue G., Tsukada K., Tanaka S. Occurrence of dehydrosqualene (C30 phytoene) in Staphylococcus aureus. Biochim Biophys Acta. 1968 Sep 2;164(1):88–93. doi: 10.1016/0005-2760(68)90074-x. [DOI] [PubMed] [Google Scholar]
  40. Takatsuji H., Nishino T., Izui K., Katsuki H. Formation of dehydrosqualene catalyzed by squalene synthetase in Saccharomyces cerevisiae. J Biochem. 1982 Mar;91(3):911–921. doi: 10.1093/oxfordjournals.jbchem.a133780. [DOI] [PubMed] [Google Scholar]
  41. Wiedemann L. M., Perry R. P. Characterization of the expressed gene and several processed pseudogenes for the mouse ribosomal protein L30 gene family. Mol Cell Biol. 1984 Nov;4(11):2518–2528. doi: 10.1128/mcb.4.11.2518. [DOI] [PMC free article] [PubMed] [Google Scholar]

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