Skip to main content
Log in

Alteration in the Expression of Genes Encoding Primary Metabolism Enzymes and Plastid Transporters during the Culture Growth of Chlamydomonas reinhardtii

  • GENOMICS. TRANSCRIPTOMICS
  • Published:
Molecular Biology Aims and scope Submit manuscript

Abstract

In a mixotrophic Chlamydomonas reinhardtii culture, the expression levels of genes encoding primary metabolic enzymes and chloroplast plastid transporters were analyzed. For the majority of the genes studied, their expression levels decreased as they approached the final stages of culture growth. During the period of exponential growth, the expression profiles changed more intensively than during the stationary phase. In the middle of exponential growth, significant changes of mRNA profiles reflected reorganization of metabolism, with an emphasis on the induction of lipid synthesis, accompanied by alterations in carbon fluxes through biochemical pathways and a shift in the energy balance between the plastid and cytosol.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.
Fig. 5.
Fig. 6.
Fig. 7.

Similar content being viewed by others

REFERENCES

  1. Perez-Garcia O., Bashan Y. 2015. Microalgal heterotrophic and mixotrophic culturing for bio-refining: From metabolic routes to techno-economics. In Algal Biorefineries, vol. 2: Products and Refinery Design. Prokop A., Bajpai R.K., Zappi M.E., Eds. Springer, pp. 61‒131.

  2. Fouchard S., Hemschemeier A., Caruana A., Pruvost J., Legrand J., Happe T., Peltier G., Cournac L. 2005. Autotrophic and mixotrophic hydrogen photoproduction in sulfur-deprived chlamydomonas cells. Appl. Environ. Microbiol. 71, 6199–6205.

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Sager R., Granick S. 1953. Nutritional studies with Chlamydomanas reinhardtii.Ann. N. Y. Acad. Sci. 56, 831‒838.

    CAS  PubMed  Google Scholar 

  4. Rochaix J.D. 2002. Chlamydomonas, a model system for studying the assembly and dynamics of photosynthetic complexes. FEBS Lett. 529, 34‒38.

    CAS  PubMed  Google Scholar 

  5. Grossman A., Lohr M., Im C. 2004. Chlamydomonas reinhardtii in the landscape of pigments. Annu. Rev. Genet. 38, 119‒173.

    CAS  PubMed  Google Scholar 

  6. Grossman A. 2000. Acclimation of Chlamydomonas reinhardtii to its nutrient environment. Protist. 151, 201‒224.

    CAS  PubMed  Google Scholar 

  7. Bolling C., Fiehn O. 2005. Metabolite profiling of Chlamydomonas reinhardtii under nutrient deprivation. Plant Physiol. 139, 1995‒2005.

    PubMed  PubMed Central  Google Scholar 

  8. Glaesener A., Merchant S., Blaby-Haas C. 2013. Iron economy in Chlamydomonas reinhardtii.Front. Plant Sci. 4, article 337.

    PubMed  PubMed Central  Google Scholar 

  9. Jamers A., Blust R., De Coen W., Griffin J., Jones O. 2013. An omics based assessment of cadmium toxicity in the green alga Chlamydomonas reinhardtii.Aquatic Toxicol.126, 355‒364.

    CAS  Google Scholar 

  10. Schmollinger S., Schulz-Raffelt M., Strenkert D., Veyel D., Vallon O., Schroda M. 2013. Dissecting the heat stress response in Chlamydomonas by pharmaceutical and RNAi approaches reveals conserved and novel aspects. Mol. Plant. 6, 1795‒1813.

    CAS  PubMed  Google Scholar 

  11. Valledor L., Furuhashi T., Hanak A., Weckwerth W. 2013. Systemic cold stress adaptation of Chlamydomonas reinhardtii.Mol. Cell. Proteomics. 12, 2032‒2047.

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Moellering E., Benning C. 2009. RNA interference silencing of a major lipid droplet protein affects lipid droplet size in Chlamydomonas reinhardtii.Eukaryot. Cell. 9, 97‒106.

    PubMed  PubMed Central  Google Scholar 

  13. Lee D.Y., Park J.J., Barupa D.K., Fiehn O. 2012. System response of metabolic networks in Chlamydomonas reinhardtii to total available ammonium. Mol. Cell. Proteomics.11, 973‒988.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Mettler T., Mühlhaus T., Hemme D., Schöttler MA., Rupprecht J., Idoine A., Veyel D., Pal S.K., Yaneva-Roder L., Winck F.V., Sommer F., Vosloh D., Seiwert B., Erban A., Burgos A., et al. 2014. Systems analysis of the response of photosynthesis, metabolism, and growth to an increase in irradiance in the photosynthetic model organism Chlamydomonas reinhardtii.Plant Cell. 26, 2310‒2350.

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Lv H., Qu G., Qi X., Lu L., Tian C., Ma Y. 2013. Transcriptome analysis of Chlamydomonas reinhardtii during the process of lipid accumulation. Genomics. 101, 229‒237.

    CAS  PubMed  Google Scholar 

  16. Lee D.Y., Fiehn O. 2013. Metabolomic response of Chlamydomonas reinhardtii to the inhibition of target of rapamycin (TOR) by rapamycin. J. Microbiol. Biotechnol. 23, 923‒931.

    CAS  PubMed  Google Scholar 

  17. Velmurugan N., Sung M., Yim S.S., Park M.S., Yang J.W., Jeong K.J. 2013. Evaluation of intracellular lipid bodies in Chlamydomonas reinhardtii strains by flow cytometry. Bioresour. Technol.138, 30‒37.

    CAS  PubMed  Google Scholar 

  18. Singh H., Shukla M.R., Chary K.V., Rao B.J. 2014. Acetate and bicarbonate assimilation and metabolite formation in Chlamydomonas reinhardtii: A 13C-NMR study. PLoS One. 9, e106457.

    PubMed  PubMed Central  Google Scholar 

  19. Humby P., Snyder E., Durnford D. 2013. Conditional senescence in Chlamydomonas reinhardtii (Chlorophyceae). J. Phycol. 49, 389‒400.

    CAS  PubMed  Google Scholar 

  20. Lee D.Y., Fiehn O. 2008. High quality metabolomic data for Chlamydomonas reinhardtii.Plant Methods.4, 7.

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Puzanskiy R., Tarakhovskaya E., Shavarda A., Shishova M. 2018. Metabolomic and physiological changes of Chlamydomonas reinhardtii (Chlorophyceae, Chlorophyta) during batch culture development. J. Appl. Phycol.30, 803‒818.

    CAS  Google Scholar 

  22. Terashima M., Specht M., Naumann B., Hippler M. 2010. Characterizing the anaerobic response of Chlamy-domonas reinhardtii by quantitative proteomics. Mol. Cell. Proteomics. 9, 1514‒1532.

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Atteia A., van Lis R., Tielens A.G., Martin W.F. 2013. Anaerobic energy metabolism in unicellular photosynthetic eukaryotes. Biochim. Biophys. Acta. 1827, 210‒223.

    CAS  PubMed  Google Scholar 

  24. Yang W., Catalanotti C., D’Adamo S., Wittkopp T., Ingram-Smith C., Mackinder L. Miller T.E., Heuberger A.L., Peers G., Smith K.S., Jonikas M.C., Grossman A.R., Posewitz M.C. 2014. Alternative acetate production pathways in Chlamydomonas reinhardtii during dark anoxia and the dominant role of chloroplasts in fermentative acetate production. Plant Cell Online. 26, 4499‒4518.

    CAS  Google Scholar 

  25. Goodenough U., Blaby I., Casero D., Gallaher S., Goodson C., Johnson S., Lee J., Merchant, S.S., Pellegrini M., Roth R., Rusch J., Singh M., Umen J.G., Weiss T.L. Wulan T. 2014. The path to triacylglyceride obesity in the sta6 strain of Chlamydomonas reinhardtii.Eukaryot. Cell.13, 591‒613.

    PubMed  PubMed Central  Google Scholar 

  26. Deng X., Cai J., Fei X. 2013. Effect of the expression and knockdown of citrate synthase gene on carbon flux during triacylglycerol biosynthesis by green algae Chlamydomonas reinhardtii.BMC Biochem. 14, 38.

    PubMed  PubMed Central  Google Scholar 

  27. Johnson X., Alric J. 2013. Central carbon metabolism and electron transport in Chlamydomonas reinhardtii: metabolic constraints for carbon partitioning between oil and starch. Eukaryot. Cell. 12, 776‒793.

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Cronan J., Waldrop G. 2002. Multi-subunit acetyl-CoA carboxylases. Progr. Lipid Res. 41, 407‒435.

    CAS  Google Scholar 

  29. Heldt H., Rapley L. 1970. Specific transport of inorganic phosphate, 3-phosphoglycerate and dihydroxyacetonephosphate, and of dicarboxylates across the inner membrane of spinach chloroplasts. FEBS Lett. 10, 143‒148.

    CAS  PubMed  Google Scholar 

  30. Atteia A., Adrait A., Brugière S., Tardif M., van Lis R., Deusch O., Dagan T., Kuhn L., Gontero B., Martin W., Garin J., Joyard J., Rolland N. 2009. A proteomic survey of Chlamydomonas reinhardtii mitochondria sheds new light on the metabolic plasticity of the organelle and on the nature of the α-proteobacterial mitochondrial ancestor. Mol. Biol. Evol.26, 1533–1548.

    CAS  PubMed  Google Scholar 

  31. Weber A., Menzlaff E., Arbinger B., Gutensohn M., Eckerskorn C., Fluegge U. 1995. The 2-oxoglutarate/malate translocator of chloroplast envelope membranes: Molecular cloning of a transporter containing a 12-helix motif and expression of the functional protein in yeast cells. Biochemistry. 34, 2621‒2627.

    CAS  PubMed  Google Scholar 

  32. Terashima M., Specht M., Hipple M. 2011. The chloroplast proteome: A survey from the Chlamydomonas reinhardtii perspective with a focus on distinctive features. Curr. Genet. 57, 151‒168.

    CAS  PubMed  Google Scholar 

  33. Gorman D., Levine R. 1965. Cytochrome f and plastocyanin: Their sequence in the photosynthetic electron transport chain of Chlamydomonas reinhardi.Proc. Natl. Acad. Sci. U. S. A.54, 1665‒1669.

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Chomczynski P. 1987. Single-step method of RNA isolation by acid guanidinium thiocyanate–phenol–chloroform extraction. Anal. Biochem.162, 156‒159.

    CAS  PubMed  Google Scholar 

  35. Pootakham W., Gonzalez-Ballester D., Grossman A. 2010. Identification and regulation of plasma membrane sulfate transporters in Chlamydomonas.Plant Physiol. 153, 1653‒1668.

    CAS  PubMed  PubMed Central  Google Scholar 

  36. Maikova A., Zalutskaya Z., Lapina T., Ermilova E. 2016. The HSP70 chaperone machines of Chlamydomonas are induced by cold stress. J. Plant Physiol. 204, 85‒91.

    CAS  PubMed  Google Scholar 

  37. Schloss J. 1990. A Chlamydomonas gene encodes a G protein β-subunit-like polypeptide. Mol. Gen. Genet. 221, 443‒452.

    CAS  PubMed  Google Scholar 

  38. Liu C., Wu G., Huang X., Liu S., Cong B. 2012. Validation of housekeeping genes for gene expression studies in an ice alga Chlamydomonas during freezing acclimation. Extremophiles. 16, 419‒425.

    CAS  PubMed  Google Scholar 

  39. Zhang H., Wang W., Li Y., Yang W., Shen G. 2011. Mixotrophic cultivation of Botryococcus braunii.Biomass. Bioenergy. 35, 1710‒1715.

    CAS  Google Scholar 

  40. R Core Team. 2016. R: A Language and Environment For Statistical Computing. Vienna, Austria: R Foundation for Statistical Computing. https://www.R-project.org/.

  41. Murtagh F., Legendre P. 2014. Ward’s hierarchical agglomerative clustering method: Which algorithms implement Ward’s criterion? J. Classification. 31, 274‒295.

    Google Scholar 

  42. Stacklies W., Redestig H., Scholz M., Walther D., Selbig J. 2007. pcaMethods: A bioconductor package providing PCA methods for incomplete data. Bioinformatics. 23, 1164‒1167.

    CAS  PubMed  Google Scholar 

  43. Thevenot E.A., Roux A., Xu Y., Ezan E., Junot C. 2015. Analysis of the human adult urinary metabolome variations with age, body mass index and gender by implementing a comprehensive workflow for univariate and OPLS statistical analyses. J. Proteome Res. 14, 3322‒3335.

    CAS  PubMed  Google Scholar 

  44. Tai Y.C., Speed T.P. 2006. A multivariate empirical Bayes statistic for replicated microarray time course data. Ann. Stat.34, 2387‒2412.

    Google Scholar 

  45. Xia J., Sinelnikov I., Han B., Wishart D.S. 2015. MetaboAnalyst 3.0: Making metabolomics more meaningful. Nucleic Acids Res.43 (W1), W251‒W257.

    Google Scholar 

  46. Xia J., Wishart D.S. 2016. Using MetaboAnalyst 3.0 for comprehensive metabolomics data analysis. Curr. Protocols Bioinform. 55, 14.10.1‒14.10.91.

  47. Bogaert K.A., Manoharan-Basil S.S., Perez E., Levine R.D., Remacle F., Remacle C. 2018. Surprisal analysis of genome-wide transcript profiling identifies differentially expressed genes and pathways associated with four growth conditions in the microalga Chlamydomonas.PLoS One. 13, e0195142.

    PubMed  PubMed Central  Google Scholar 

  48. Bogaert K.A., Perez E., Rumin J., Giltay A., Carone M., Coosemans N., Radoux M., Eppe G., Levine R.D., Remacle F., Remacle C. 2019. Metabolic, physiological, and transcriptomics analysis of batch cultures of the green microalga Chlamydomonas grown on different acetate concentrations. Cells.8, 1367.

    CAS  PubMed Central  Google Scholar 

  49. Work V., Radakovits R., Jinkerson R., Meuser J., Elliott L., Vinyard D., Laurens L., Dismukes C., Posewitz M. 2010. Increased lipid accumulation in the Chlamydomonas reinhardtii sta7-10 starchless isoamylase mutant and increased carbohydrate synthesis in complemented strains. Eukaryot. Cell. 9, 1251‒1261.

    CAS  PubMed  PubMed Central  Google Scholar 

  50. Ramanan R., Kim B., Cho D., Ko S., Oh H., Kim H. 2013. Lipid droplet synthesis is limited by acetate availability in starchless mutant of Chlamydomonas reinhardtii.FEBS Lett.587, 370‒377.

    CAS  PubMed  Google Scholar 

  51. Lalibertè G., de la Noüie J. 1993. Auto-, hetero-, and mixotrophic growth of Chlamydomonas humicola (Cmloroimiyckak) on acetate. J. Phycol.29, 612‒620.

    Google Scholar 

  52. Therien J.B., Zadvornyy O.A., Posewitz M.C., Bryant D.A., Peters J.W. 2014. Growth of Chlamydomonas reinhardtii in acetate-free medium when co-cultured with alginate-encapsulated, acetate-producing strains of Synechococcus sp. PCC 7002. Biotechnol. Biofuels. 7, 154.

    PubMed  PubMed Central  Google Scholar 

  53. Eppley R., Gee R., Saltman P. 1963. Photometabolism of acetate by Chlamydomonas mundana.Physiol. Plant. 16, 777‒792.

    CAS  Google Scholar 

  54. Sugimoto T., Tanaka K., Monma M., Kawamura Y., Saio K. 1989. Phosphoenolpyruvate carboxylase level in soybean seed highly correlates to its contents of protein and lipid. Agricult. Biol. Chem. 53, 885‒887.

    CAS  Google Scholar 

  55. Chen J.Q., Lang C.X., Hu Z.H., Liu Z.H., Huang R.Z. 1999. Antisense PEP gene regulates to ratio of protein and lipid content in Brassica napus seeds. J. Agricult. Biotechnol.7, 316‒320.

    Google Scholar 

  56. Leyva L., Bashan Y., Mendoza A., de-Bashan L. 2014. Accumulation fatty acids of in Chlorella vulgaris under heterotrophic conditions in relation to activity of acetyl-CoA carboxylase, temperature, and co-immobilization with Azospirillum brasilense.Naturwissenschaften. 101, 819‒830.

    CAS  PubMed  Google Scholar 

  57. Radakovits R., Jinkerson R., Darzins A., Posewitz M. 2010. Genetic engineering of algae for enhanced biofuel production. Eukaryot. Cell. 9, 486‒501.

    CAS  PubMed  PubMed Central  Google Scholar 

  58. Mus F., Dubini A., Seibert M., Posewitz M., Grossman A. 2007. Anaerobic acclimation in Chlamydomonas reinhardtii: Anoxic gene expression, hydrogenase induction, and metabolic pathways. J. Biol. Chem. 282, 25475‒25486.

    CAS  PubMed  Google Scholar 

  59. Plancke C., Vigeolas H., Höhner R., Roberty S., Emonds-Alt B., Larosa V., Willamme R., Duby F., Onga D.D., Thonart P., Hiligsmann S., Franck F., Eppe G., Cardol P., Hippler M., Remacle C. 2014. Lack of isocitrate lyase in Chlamydomonas leads to changes in carbon metabolism and in the response to oxidative stress under mixotrophic growth. Plant J.77, 404‒417.

    CAS  PubMed  Google Scholar 

  60. Park J., Wang H., Gargouri M., Deshpande R., Skepper J., Holguin F., Juergens M.T., Shachar-Hill Y., Hicks L.M., Gang D.R. 2015. The response of Chlamy-domonas reinhardtii to nitrogen deprivation: A systems biology analysis. Plant J.81, 611‒624.

    CAS  PubMed  Google Scholar 

  61. Wienkoop S., Weiß J., May P., Kempa S., Irgang S., Recuenco-Munoz L., Pietzke M., Schwemmer T., Rupprecht J., Egelhofer V., Weckwerth W. 2010. Targeted proteomics for Chlamydomonas reinhardtii combined with rapid subcellular protein fractionation, metabolomics and metabolic flux analyses. Mol. Biosyst. 6, 1018‒1031.

    CAS  PubMed  Google Scholar 

  62. Davey M., Horst I., Duong G., Tomsett E., Litvinenko A., Howe C., Smith A. 2014. Triacylglyceride production and autophagous responses in Chlamydomonas reinhardtii depend on resource allocation and carbon source. Eukaryot. Cell.13, 392‒400.

    PubMed  PubMed Central  Google Scholar 

Download references

ACKNOWLEDGMENTS

Using the equipment of the Resource Center “Molecular and Cell Technologies” of St. Petersburg State University.

Funding

This work was supported by the Russian Foundation for Basic Research (project nos. 15-04-03090 and 16-34-01122) and St. Petersburg State University (Research project no. 1.40.495.2017).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to M. F. Shishova.

Ethics declarations

The authors declare that they have no conflict of interest. This article does not contain any studies involving animals or human participants performed by any of the authors.

Additional information

Translated by D. Timchenko

Abbreviations: PC, principal component; PCA, principal component analysis; MEBA, multivariate empirical Bayes analysis; (O)PLS‒DA, (orthogonal) projections on latent structures – discriminant analysis); DBI, Davies–Bouldin index; VIP, Variable Importance in Projection; PPP, pentose phosphate pathway; PEP, phosphoenolpyruvate; TAG, triacylglycerol; ACC, acetyl-CoA carboxylase; SD, standard deviation; AU p, approximately unbiased p-value.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Puzanskiy, R.K., Romanyuk, D.A., Kirpichnikova, A.A. et al. Alteration in the Expression of Genes Encoding Primary Metabolism Enzymes and Plastid Transporters during the Culture Growth of Chlamydomonas reinhardtii . Mol Biol 54, 503–519 (2020). https://doi.org/10.1134/S0026893320040147

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1134/S0026893320040147

Keywords:

Navigation