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Functional analysis of very long-chain fatty acid elongase gene, HpELO2, in the methylotrophic yeast Hansenula polymorpha


We describe the cloning and functional characterization of the fatty acid elongase gene HpELO2, a homologue of the HpELO1 gene required for the production of C24:0 in the yeast Hansenula polymorpha. The open reading frame (ORF) of HpELO2 consists of 1,035 bp, encoding 344 amino acids, sharing about 65% identity with that of Saccharomyces cerevisiae Elo2. Expression of HpELO2 rescued the lethality of the S. cerevisiae elo2Δ elo3Δ double disruptant. An accumulation of C18:0 and a significant increase and decrease in the levels of C24:0 and C26:0, respectively, were observed in the Hpelo2Δ disruptant. These results supported an idea that HpELO2 encodes a fatty acid elongase involved in the elongation of C18:0 to very long-chain fatty acids. The Hpelo1Δ Hpelo2Δ double disruption was nonviable, suggesting that HpELO1 and HpELO2 are the only two genes necessary for the biosynthesis in H. polymorpha. Interestingly, transcription of HpELO2 and HpELO1 were found to be transiently up-regulated by exogenous long-chain fatty acids; however, this up-regulation was not observed with HpELO1 and HpELO2 genes driven by the constitutively expressed promoter of the HpACT gene, suggesting that exogenous fatty acids specifically trigger the transcriptional induction of HpELO1 and HpELO2 through their promoter regions.

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  1. Amberg DC, Burke DJ, Stratern JN (eds) (2005) Methods in yeast genetics: PCR-mediated gene disruption. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, pp 155–160

  2. Anamnart S, Tomita T, Fukui F, Fujimori K, Harashima S, Yamada Y, Oshima Y (1997) The P-Ole1 gene of Pichia angusta encodes a Δ9-fatty acid desaturase and complements the ole1 mutation of Saccharomyces cerevisiae. Gene 184:299–306

  3. Anamnart S, Tolstorukuv I, Kaneko Y, Harashima S (1998) Fatty acid desaturation in methylotrophic yeast Hansenula polymorpha strain CBS1976 and unsaturated fatty acid auxotrophic mutants. J Ferment Bioeng 85:476–482

  4. Cassagne C, Lessire R, Bessoule JJ, Moreau P, Creach A, Schneider F, Sturbois B (1994) Biosynthesis of very long chain fatty acids in higher plants. Prog Lipid Res 33:55–69

  5. Chirala SS (1992) Coordinated regulation and inositol-mediated and fatty acid mediated repression of fatty acid synthase genes in Saccharomyces cerevisiae. Proc Natl Acad Sci USA 89:10232–10236

  6. Cinti CL, Cook L, Nagi MN, Suneja SK (1992) The fatty acid chain elongation system of mammalian endoplasmic reticulum. Prog Lipid Res 31:1–51

  7. Dunn TM, Haak D, Monagham E, Beeler TJ (1998) Synthesis of monohydroxylated inositolphosphorylceramide (IPC-C) in Saccharomyces cerevisiae requires Scs7p, a protein with both a cytochrome b5-like domain and a hydroxylase/desaturase domain. Yeast 14:311–321

  8. Eisenkolb M, Zenzmaier C, Leitner E, Schneiter R (2002) A specific structural requirement for ergosterol in long-chain fatty acid synthesis mutants important for maintaining raft in yeast. Mol Biol Cell 13:4414–4428

  9. Fox BG, Shanklin J, Ai J, Loehr TM, Sanders-Loehr J (1994) Resonance Raman evidence for an Fe–O–Fe center in stearoyl-ACP desaturase: primary sequence identity with other diiron-oxo proteins. Biochemistry 33:12776–12786

  10. Gaigg B, Toulmay A, Schneither R (2006) Very long-chain fatty acid-containing lipids rather than sphingolipids per se are required for raft association and stable surface transport of newly synthesized plasma membrane ATPase in yeast. J Biol Chem 281:34135–34145

  11. Gasch AP, Spellman PT, Kao CM, Carnel-Harel O, Eisen MB, Storz G, Botstein D, Brown PO (2000) Genomic expression programs in the response of yeast cells to environmental changes. Mol Biol Cell 11:4241–4257

  12. Haan GJ, van Dijk R, Kiel JAKW, Veenhuis M (2001) Characterization of the Hansenula polymorpha PUR7 gene and its use as selectable marker for targeted chromosomal integration. FEMS Yeast Res 2:17–24

  13. Kamiryo T, Parthasarathy S, Numa S (1976) Evidence that acetyl coenzyme A synthase activity is required for repression of yeast acetyl coenzyme A carboxylase by exogenous fatty acids. Proc Natl Acad Sci USA 73:386–390

  14. Kamiryo T, Nishikawa Y, Mishina M, Terao M, Numa S (1979) Involvement of long-chain acyl coenzyme A for lipid synthesis in repression of acetyl-coenzyme A carboxylase in Candida lipolytica. Proc Natl Acad Sci USA 76:4390–4394

  15. Kaneko Y, Rueksomtawin K, Maekawa T, Harashima S (2003) Structural and functional analysis of the FAS1 gene encoding fatty acid synthase β-subunit in methylotrophic yeast Hansenula polymorpha. In: Murata N, Yamada M, Nishida I, Okuyama H, Sekiya J, Wada H (eds) Advanced research on plant lipids. Kluwer, London, pp 61–64

  16. Kang HA, Hong W-K, Sohn J-H, Choi E-S, Rhee SK (2001) Molecular characterization of the actin-encoding gene and the use of its promoter for a dominant selection system in the methylotrophic yeast Hansenula polymorpha. Appl Microbiol Biotechnol 55:734–741

  17. Laoteng K, Ruenwai R, Tanticharoen M, Cheev-adhanarak S (2005) Genetic modification of essential fatty acids biosynthesis in Hansenula polymorpha. FEMS Microbiol Lett 245:169–178

  18. Leonard AE, Pereira SL, Howard S, Huang Y-S (2004) Elongation of long-chain fatty acids. Prog Lipid Res 43:36–54

  19. Lu S-F, Tolstorukov II, Anamnart S, Kaneko Y, Harashima S (2000) Cloning, sequencing, and functional analysis of H-OLE1 gene encoding Δ9-fatty acid desaturase in Hansenula polymorpha. Appl Microbiol Biotechnol 54:499–509

  20. Mcdonough VM, Stukey JE, Martin CE (1992) Specificity of unsaturated fatty acid-regulated expression of the Saccharomyces cerevisiae OLE1 gene. J Biol Chem 267:5931–5936

  21. Mukaiyama H, Oku M, Baba M, Samizo T, Hammond AT, Glick BS, Kato N, Sakai Y (2002) Paz2 and 13 other PAZ gene products regulate vacuolar engulfment of peroxisome during micropexophagy. Genes Cells 7:75–90

  22. Oh C-S, Toke DA, Mandala S, Martin CE (1997) ELO2 and ELO3, homologues of the Saccharomyces cerevisiae ELO1 gene, function in fatty acid elongation and are required for sphingolipid formation. J Biol Chem 272:17376–17384

  23. Ohlrogge JB, Jaworski JG (1997) Regulation of fatty acid synthesis. Annu Rev Plant Physiol Plant Mol Biol 48:109–136

  24. Oshiro T, Aiba H, Mizuno T (2003) A defect in a fatty acyl-CoA synthase gene, lcf1+, results in a decrease in viability after entry into the stationary phase in fission yeast. Mol Genet Genomics 269:437–442

  25. Paul S, Gable K, Beaudoin F, Cahoon E, Jaworski J, Napier JA, Dunn TM (2006) Members of the Arabidopsis FAE1-like 3-ketoacyl-CoA synthase gene family substitute for the Elop proteins of Saccharomyces cerevisiae. J Biol Chem 281:9018–9029

  26. Prasitchoke P, Kaneko Y, Bamba T, Fukusaki E, Kobayashi A, Harashima S (2007) Identification and characterization of a very-long-chain fatty acid elongase gene in the methylotrophic yeast, Hansenula polymorpha. Gene 391:16–25

  27. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory: Cold Spring Harbor, New York

  28. Schweizer M (2004) Lipids and membranes. In: Dickinson JR, Schweizer M (eds) The metabolism and molecular physiology of Saccharomyces cerevisiae. CRC LLC, New York, pp 140–223

  29. Shanklin J, Whittle E, Fox BG (1994) Eight histidine residues are catalytically essential in a membrane-associated iron enzyme, stearoyl-CoA desaturase, and are conserved in alkane hydroxylase and xylene monooxygenase. Biochemistry 33:12787–12794

  30. Stock SD, Hama H, Radding JA, Young DA, Takemoto JY (2000) Syringomycin E inhibition of Saccharomyces cerevisiae: requirement for biosynthesis of sphingolipids with very-long-chain fatty acids and mannose- and phosphoinositol-containing head groups. Antimicrob Agents Chemother 44:1174–1180

  31. Toke DA, Martin CE (1996) Isolation and characterization of a gene affecting fatty acid elongation in Saccharomyces cerevisiae. J Biol Chem 271:18413–18422

  32. Tong F, Black PN, Bivins L, Quackenbush S, Ctrnacta V, DiRusso CC (2006) Direct interaction of Saccharomyces cerevisiae Faa1p with the Omi/HtrA protease orthologue Ynm3p alters lipid homeostasis. Mol Genet Genomics 275:330–343

  33. Wijeyaratne SC, Ohta K, Chavanich S, Mahamontri V, Ninubol N, Hayashida S (1986) Lipid composition of a thermotolerant yeast, Hansenula polymorpha. Agric Biol Chem 50:827–832

  34. Zeidel ML (1996) Low permeabilities of apical membranes of barrier epithelia: what makes watertight membranes watertight? Am J Physiol 271:F243–F245

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We would like to thank Y. Sakai for providing plasmid pREMI-Z, M. Veenhuis for providing plasmid pHIPA4, and H. A. Kang for providing plasmid pHACT850-HyL.

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Correspondence to Satoshi Harashima.

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Prasitchoke, P., Kaneko, Y., Sugiyama, M. et al. Functional analysis of very long-chain fatty acid elongase gene, HpELO2, in the methylotrophic yeast Hansenula polymorpha . Appl Microbiol Biotechnol 76, 417–427 (2007).

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  • Methylotrophic yeast
  • Hansenula polymorpha
  • Elongase for very long-chain fatty acids
  • Transient transcriptional induction