Homologous expression of lipid droplet protein-enhanced neutral lipid accumulation in the marine diatom Phaeodactylum tricornutum

Journal of Applied Phycology volume 30, pages2793–2802(2018)Cite this article

Abstract

Lipid droplets are ubiquitous cellular compartments that store neutral lipids and specific proteins localize on their surface. These proteins work as a scaffold in maintaining the lipid droplet structure or as regulators of lipogenesis or lipolysis. Previously, the most abundant lipid droplet protein, namely stramenopile-type lipid droplet protein (StLDP), was identified in the marine diatom Phaeodactylum tricornutum; however, its function remains unclear because StLDP does not reveal homology with known lipid droplet proteins and lacks a predictable domain. In this study, P. tricornutum was transformed to express a homologous StLDP gene under an fcpA promoter in order to determine its function. StLDP expression was strongly enhanced in the mutant (H8), especially in nitrogen-sufficient conditions; however, it was attenuated in nitrogen-deficient conditions. Despite the strong expression, no significant difference was observed in the lipid composition between the wild type (WT) and H8 under nitrogen-sufficient conditions. After cultivation in nitrogen-free medium for 6 days, neutral lipid content significantly increased in H8 than in WT. After 2 days of cultivation in nitrogen-free medium, 97.0% of single cells in WT formed one or two lipid droplets, whereas in H8, this proportion decreased to 78.8%, and the proportion of cells forming three or four lipid droplets increased. Thus, the function of StLDP was speculated to sequester triacylglycerol on the initial lipid droplet formation.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

References

  1. Abida H, Dolch LJ, Meï C, Villanova V, Conte M, Block MA, Finazzi G, Bastien O, Tirichine L, Bowler C, Rébeillé F (2015) Membrane glycerolipid remodeling triggered by nitrogen and phosphorus starvation in Phaeodactylum tricornutum. Plant Physiol 167:118–136

    CAS  Article  Google Scholar 

  2. Abramoff MD, Magalhaes PJ, Ram SJ (2004) Image processing with image. J Biophot Int 11:36–42

    Google Scholar 

  3. Alipanah L, Rohloff J, Winge P, Bones AM, Brembu T (2015) Whole-cell response to nitrogen deprivation in the diatom Phaeodactylum tricornutum. J Exp Bot 66:6281–6296

    CAS  Article  Google Scholar 

  4. Chapman KD, Dyer JM, Mullen RT (2012) Biogenesis and functions of lipid droplets in plants. J Lipid Res 53:215–216

    CAS  Article  Google Scholar 

  5. Chisti Y (2007) Biodiesel from microalgae. Biotechnol Adv 25:294–306

    CAS  Article  Google Scholar 

  6. Davidi L, Katz A, Pick U (2012) Characterization of major lipid droplet proteins from Dunaliella. Planta 236:19–33

    CAS  Article  Google Scholar 

  7. Fajardo AR, Cerdan LE, Medina AR, Fermamdez FGA, Moreno PAG, Grima EM (2007) Lipid extraction from the microalga Phaeodactylum tricornutum. Eur J Lipid Sci Technol 109:120–126

    CAS  Article  Google Scholar 

  8. Folch J, Lee M, Stanley GHS (1957) A simple method for the isolation and purification of total lipids from animal tissues. J Biol Chem 226:497–509

    CAS  PubMed  Google Scholar 

  9. Guiheneuf F, Leu S, Zarka A, Khozin-Goldberg I, Khalilov I, Boussiba S (2011) Cloning and molecular characterization of a novel acyl-CoA: diacylglycerol acyltransferase 1-like gene (PtDGAT1) from the diatom Phaeodactylum tricornutum. FEBS J 278:3651–3666

    CAS  Article  Google Scholar 

  10. Guillard RRL, Ryther JH (1962) Studies of marine planktonic diatoms. I. Cyclotella nana Hustedt and Detonula confervaceae (Cleve) Gran. Can J Microbiol 8:229–239

    CAS  Article  Google Scholar 

  11. Huang G, Chen F, Wei D, Zhang X, Chen G (2010) Biodiesel production by microalgal biotechnology. Appl Energy 87:38–46

    CAS  Article  Google Scholar 

  12. Jacquier N, Mishra S, Choudhary V, Schneiter R (2013) Expression of oleosin and perilipins in yeast promotes formation of lipid droplets from the endoplasmic reticulum. J Cell Sci 126:5198–5209

    CAS  Article  Google Scholar 

  13. Javee A, Sulochana SB, Pallissery SJ, Arumugam M (2016) Major lipid body protein: a conserved structural component of lipid body accumulated during abiotic stress in S. quadricauda CASA-CC302. Front Energ Res 4:37

    Article  Google Scholar 

  14. Kimmel AR, Brasaemle DL, McAndrews-Hill M, Sztalryd C, Londos C (2010) Adoption of PERILIPIN as a unifying nomenclature for the mammalian PAT-family of intracellular lipid storage droplet proteins. J Lipid Res 51:468–471

    CAS  Article  Google Scholar 

  15. Lin I, Jiang P, Chen C, Tzen JTC (2012) A unique caleosin serving as the major integral protein in oil bodies isolated from Chlorella sp. cells cultured with limited nitrogen. Plant Physiol Biochem 61:80–87

    CAS  Article  Google Scholar 

  16. Liu WX, Liu HL, Qu LQ (2013) Embryo-specific expression of soybean oleosin altered oil body morphogenesis and increased lipid content in transgenic rice seeds. Theor Appl Genet 126:2289–2297

    CAS  Article  Google Scholar 

  17. Martin S, Parton RG (2006) Lipid droplets: a unified view of a dynamic organelle. Nat Rev Mol Cell Bio 7:373–378

    CAS  Article  Google Scholar 

  18. Mata TM, Martins AA, Caetano NS (2010) Microalgae for biodiesel production and other applications: a review. Renew Sust Energ Rev 14:217–232

    CAS  Article  Google Scholar 

  19. Miyoshi H, Souza SC, Endo M, Sawada T, Perfield JW, Shimizu C, Stancheva Z, Nagai S, Strissel KJ, Yoshioka N, Obin MS (2010) Perilipin overexpression in mice protects against diet-induced obesity. J Lipid Res 51:975–982

    CAS  Article  Google Scholar 

  20. Moellering ER, Benning C (2010) RNA interference silencing of a major lipid droplet protein affects lipid droplet size in Chlamydomonas reinhardtii. Eukaryot Cell 9:97–106

    CAS  Article  Google Scholar 

  21. Murphy DJ, Vance J (1999) Mechanisms of lipid-body formation. Trends Biochem Sci 24:109–115

    CAS  Article  Google Scholar 

  22. Murphy DJ (2012) The dynamic roles of intracellular lipid droplets: from archaea to mammals. Protoplasma 249:541–585

    CAS  Article  Google Scholar 

  23. Nguyen HM, Baudet M, Cuine S, Adriano JM, Barthe D, Billon E, Bruley C, Beisson F, Peltier G, Ferro M, Li-Beisson Y (2011) Proteomic profiling of oil bodies isolated from the unicellular green microalga Chlamydomonas reinhardtii: with focus on proteins involved in lipid metabolism. Proteomics 11:4266–4273

    CAS  Article  Google Scholar 

  24. Nojima D, Yoshino T, Maeda Y, Tanaka M, Nemoto M, Tanaka T (2013) Proteomics analysis of oil body associated proteins in the oleaginous diatom. J Proteome Res 12:5293–5301

    CAS  Article  Google Scholar 

  25. Ohsaki Y, Suzuki M, Fujimoto T (2014) Open questions in lipid droplet biology. Chem Biol 21:86–96

    CAS  Article  Google Scholar 

  26. Pasaribu B, Lin IP, Tzen JT, Jauh GY, Fan TY, Ju YM, Cheng JO, Chen CS, Jiang PL (2014) SLDP: a novel protein related to caleosin is associated with the endosymbiotic Symbiodinium lipid droplets from Euphyllia glabrescens. Mar Biotechnol 16:560–571

    CAS  Article  Google Scholar 

  27. Peled E, Leu S, Zarka A, Weiss M, Pick U, Khozin-Goldberg I, Boussiba S (2011) Isolation of a novel oil globule protein from the green alga Haematococcus pluvialis (Chlorophyceae). Lipids 46:851–861

    CAS  Article  Google Scholar 

  28. Pol A, Gross SP, Parton RG (2014) Biogenesis of the multifunctional lipid droplet: lipids, proteins, and sites. J Cell Biol 204:635–646

    CAS  Article  Google Scholar 

  29. Pollak NM, Schweiger M, Jaeger D, Kolb D, Kumari M, Schreiber R, Kolleritsch S, Markolin P, Grabner GF, Heier C, Zierler KA (2013) Cardiac-specific overexpression of perilipin 5 provokes severe cardiac steatosis via the formation of a lipolytic barrier. J Lipid Res 54:1092–1102

    CAS  Article  Google Scholar 

  30. Sawada T, Miyoshi H, Shimada K, Suzuki A, Okamatsu-Ogura Y, Perfield JW II, Kondo T, Nagai S, Shimizu C, Yoshioka N, Greenberg AS (2010) Perilipin overexpression in white adipose tissue induces a brown fat-like phenotype. PLoS One 5:e14006

    Article  Google Scholar 

  31. Schmidt MA, Herman EM (2008) Suppression of soybean oleosin produces micro-oil bodies that aggregate into oil body/ER complexes. Mol Plant 1:910–924

    CAS  Article  Google Scholar 

  32. Shemesh Z, Leu S, Khozin-Goldberg I, Didi-Cohen S, Zarka A, Boussiba S (2016) Inducible expression of Haematococcus oil globule protein in the diatom Phaeodactylum tricornutum: association with lipid droplets and enhancement of TAG accumulation under nitrogen starvation. Algal Res 18:321–331

    Article  Google Scholar 

  33. Shi Q, Araie H, Bakku RK, Fukao Y, Rakwal R, Suzuki I, Shiraiwa Y (2015) Proteomic analysis of lipid body from the alkenone-producing marine haptophyte alga Tisochrysis lutea. Proteomics 15:4145–4158

    CAS  Article  Google Scholar 

  34. Siegler H, Valerius O, Ischebeck T, Popko J, Tourasse NJ, Vallon O, Khozin-Goldberg I, Braus GH, Feussner I (2017) Analysis of the lipid body proteome of the oleaginous alga Lobosphaera incisa. BMC Plant Biol 17:98

    Article  Google Scholar 

  35. Siloto RMP, Findlay K, Villalobos AL, Yeung EC, Nykiforuk CL, Moloney MM (2006) The accumulation of oleosins determines the size of seed oilbodies in Arabidopsis. Plant Cell 18:1961–1974

    CAS  Article  Google Scholar 

  36. Sun Z, Miller RA, Patel RT, Chen J, Dhir R, Wang H, Zhang D, Graham MJ, Unterman TG, Shulman GI, Sztalryd C (2012) Hepatic Hdac3 promotes gluconeogenesis by repressing lipid synthesis and sequestration. Nat Med 18:934–942

    CAS  Article  Google Scholar 

  37. Sztalryd C, Kimmel AR (2014) Perilipins: lipid droplet coat proteins adapted for tissue-specific energy storage and utilization, and lipid cytoprotection. Biochimie 96:96–101

    CAS  Article  Google Scholar 

  38. Trentacoste EM, Shrestha RP, Smith SR, Gle C, Hartmann AC, Hildebrand M, Gerwick WH (2013) Metabolic engineering of lipid catabolism increases microalgal lipid accumulation without compromising growth. Proc Natl Acad Sci U S A 110:19748–19753

    CAS  Article  Google Scholar 

  39. Vieler A, Brubaker SB, Vick B, Benning C (2012) A lipid droplet protein on Nannochloropsis with functions partially analogous to plant oleosins. Plant Physiol 158:1562–1569

    CAS  Article  Google Scholar 

  40. Wang H, Sreenivasan U, Gong DW, O’Connell KA, Dabkowski ER, Hecker PA, Ionica N, Konig M, Mahurkar A, Sun Y, Stanley WC (2013) Cardiomyocyte-specific perilipin 5 overexpression leads to myocardial steatosis and modest cardiac dysfunction. J Lipid Res 54:953–965

    CAS  Article  Google Scholar 

  41. Yang L, Ding Y, Chen Y, Zhang S, Huo C, Wang Y, Yu J, Zhang P, Na H, Zhang H, Ma Y (2012) The proteomics of lipid droplets: structure, dynamics, and functions of organelle conserved from bacteria to humans. J Lipid Res 53:1245–1253

    CAS  Article  Google Scholar 

  42. Yoneda K, Yoshida M, Suzuki I, Watanabe MM (2016) Identification of a major lipid droplet protein in a marine diatom Phaeodactylum tricornutum. Plant Cell Physiol 57:397–406

    CAS  Article  Google Scholar 

  43. Zaslavskaia LA, Lippmeier JC, Kroth PG, Grossman AR, Apt KE (2000) Transformation of the diatom Phaeodactylum tricornutum (Bacillariophyceae) with a variety of selectable marker and reporter genes. J Phycol 36:379–386

    CAS  Article  Google Scholar 

Download references

Acknowledgments

This study was supported by the Grant-in-Aid for Japan Society for the Promotion of Science (JSPS) Research Fellow [JSPS KAKENHI Grant Number JP17J03747 to K.Y.].

Author information

Affiliations

  1. Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8572, Japan

    Kohei Yoneda, Masaki Yoshida, Iwane Suzuki & Makoto M. Watanabe

Authors
  1. Kohei Yoneda
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Masaki Yoshida
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Iwane Suzuki
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Makoto M. Watanabe
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Masaki Yoshida.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Electronic supplementary material

ESM 1

(DOCX 11 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Yoneda, K., Yoshida, M., Suzuki, I. et al. Homologous expression of lipid droplet protein-enhanced neutral lipid accumulation in the marine diatom Phaeodactylum tricornutum. J Appl Phycol 30, 2793–2802 (2018). https://doi.org/10.1007/s10811-018-1402-9

Download citation

Keywords

  • Diatom
  • Nitrogen starvation
  • Overexpression
  • StLDP
  • Triacylglycerol