Molecular Biology

, Volume 52, Issue 4, pp 520–531 | Cite as

In silico Analyses of Transcriptomes of the Marine Green Microalga Dunaliella tertiolecta: Identification of Sequences Encoding P-type ATPases

  • L. G. Popova
  • D. V. Belyaev
  • A. V. Shuvalov
  • A. A. Yurchenko
  • D. A. Matalin
  • D. E. Khramov
  • Y. V. Orlova
  • Y. V. Balnokin
Genomics. Transcriptomics


De novo assembled transcriptomes of the marine microalga Dunaliella tertiolecta (Chlorophyta) were analyzed. Transcriptome assemblies were performed using short-read RNA-seq data deposited in the SRA database (DNA and RNA Sequence Read Archive, NCBI). A merged transcriptome was assembled using a pooled RNA-seq data set. The goal of the study was in silico identification of nucleotide sequences encoding P-type ATPases in D. tertiolecta transcriptomes. P-type ATPases play a considerable role in the adaptation of an organism to a variable environment, and this problem is particularly significant for microalgae inhabiting an environment with an unstable ionic composition. Particular emphasis was given to searching for a sequence coding Na+-ATPase. This enzyme is expected to function in the plasma membrane of D. tertiolecta like in some marine algae, in particular, in the closely related alga Dunaliella maritima. An ensemble of 12 P-type ATPases consisting of members belonging to the five main subfamilies of the P-type ATPase family was revealed in the assembled transcriptomes. The genes of the following P-type ATPases were found: (1) heavy metal ATPases (subfamily PIB); (2) Ca2+-ATPases of SERCA type (subfamily P2A); (3) H+-ATPases (subfamily P3); (4) phospholipid-transporting ATPases (flippases) (subfamily P4); (5) cation- transporting ATPases of uncertain specificities (subfamily P5). The presence of functional Na+-ATPases in marine algae is presently undoubted. However, contrary to expectations, we failed to find a nucleotide sequence encoding a protein that could unequivocally be considered a Na+-ATPase. Further study is necessary to elucidate the roles of in silico revealed D. tertiolecta ATPases in Na+ transport.


Dunaliella tertiolecta P-type ATPases marine microalgae de novo transcriptome assembly 


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  1. 1.
    Scarborough G.A. 1999. Structure and function of the P-type ATPases. Curr. Opin. Cell Biol. 11, 517–522.CrossRefPubMedGoogle Scholar
  2. 2.
    Palmgren M.G., Harper J.F. 1999. Pumping with plant P-type ATPases. J. Exp. Bot. 50, 883–893.CrossRefGoogle Scholar
  3. 3.
    Axelsen K., Palmgren M.G. 1998. Evolution and substrate specificities in the P-type ATPase superfamily. J. Mol. Evol. 46, 84–101.CrossRefPubMedGoogle Scholar
  4. 4.
    Axelsen K.B., Palmgren M.G. 2001. Inventory of the superfamily of P-type ion pumps in Arabidopsis. Plant Physiol. 126, 696–706.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Derelle E., Ferraz C., Rombauts S. 2006. Genome analysis of the smallest free-living eukaryote Ostreococus tauri unveils many unique features. Proc. Natl. Acad. Sci USA. 103, 11647–11652.CrossRefPubMedGoogle Scholar
  6. 6.
    Merchant S.S., Prochnik S.E., Vallon O., Harris E.H., Karpowicz S.J., Witman G.B., Terry A., Salamov A., Fritz-Laylin L.K., Marechal-Drouard L., et al. 2007. The Chlamydomonas genome reveals the evolution of key animal and plant functions. Science. 318, 245–250.CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Foflonker F., Price D.C., Qiu H., Palenik B., Wang S., Bhattacharya D. 2014. Genome of the halotolerant green alga Picochlorum sp. reveals strategies for thriving under fluctuating environmental conditions. Environ. Microbiol. 17 (2), 412–426. doi 10.1111/1462-2920.12541PubMedGoogle Scholar
  8. 8.
    Ward J.A., Ponnala L., Weber C.A. 2012. Strategies for transcriptome analysis in nonmodel plants. Am. J. Bot. 99, 267–276.CrossRefPubMedGoogle Scholar
  9. 9.
    Jain M. 2012. Next generation sequencing technologies for gene expression profiling in plants. Brief. Funct. Genomics. 2, 63–70.CrossRefGoogle Scholar
  10. 10.
    Zhao Q.-Y., Wang Y., Kong Y.-M., Luo D., Li X., Hao P. 2011. Optimizing de novo transcriptome assembly from short-read RNA-Seq data: A comparative study. BMC Bioinformatics. 12 (Suppl. 14), S2. CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Dunaliella salina Genome Sequencing Project. US Department of Energy Join Genome Institute.
  12. 12.
    Massyuk N.P. 1973. Morphology, Taxonomy, Ecology and Geographic Distribution of the Genus Dunaliella Teod. and Prospects for its Potential Utilization. Kiev: Naukova Dumka.Google Scholar
  13. 13.
    Oren A. 2005. A hundred years of Dunaliella research: 1905–2005. Saline Systems. 1, 1–14.CrossRefGoogle Scholar
  14. 14.
    Katz A., Avron M. 1985. Determination of intracellular osmotic volume and sodium concentration in Dunaliella. Plant Physiol. 78, 817–820.CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Shumkova G.A., Popova L.G., Balnokin Y.V. 2000. Export of Na+ from cells of the halotolerant microalga Dunaliella maritima: Na+/H+ antiporter or primary Na+-pump? Biochemistry (Moscow). 65, 917–923.PubMedGoogle Scholar
  16. 16.
    Rodriguez-Navarro A., Benito B. 2010. Sodium or potassium efflux ATPase. A fungal, briophyte, and protozoal ATPase. Biochim. Biophys. Acta. 1798, 1841–1853.CrossRefGoogle Scholar
  17. 17.
    Balnokin Yu.V, 2012. Ionnyi gomeostaz i soleustoichivost' rastenii: 70-e Timiryazevskoe chtenie (Plant Ion Homeostasis and Salt Tolerance. The 70th Timiryazev’s Lectures). Moscow: Nauka.Google Scholar
  18. 18.
    Yamaguchi T., Hamamoto S., Uozumi N. 2013. Sodium transport system in plant cells. Front. Plant Sci. 4, 410. doi 10.3389/fpls.2013.00410CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Balnokin Yu., Popova L., Gimmler H. 1997. Further evidence for an ATP-driven sodium pump in the marine alga Tetraselmis (Platymonas) viridis. J. Plant Physiol. 150, 264–270.CrossRefGoogle Scholar
  20. 20.
    Popova L.G., Balnokin Y.V. 2013. Na+-ATPases of halotolerant microalgae. Russian J. Plant Physiol. 60, 472–482.CrossRefGoogle Scholar
  21. 21.
    Balnokin Y.V., Popova L.G. 1994. The ATP-driven Na+-pump in the plasma membrane of the marine unicellular alga Platymonas viridis. FEBS Lett. 343, 61–64.CrossRefPubMedGoogle Scholar
  22. 22.
    Popova L.G., Shumkova G.A., Andreev I.M., Balnokin Y.V. 2005. Functional identification of electrogenic Na+-translocating ATPase in the plasma membrane of the halotolerant microalga Dunaliella maritima. FEBS Lett. 579, 5002–5006.CrossRefPubMedGoogle Scholar
  23. 23.
    Popova L.G., Shumkova G.A., Andreev I.M., Balnokin Y.V. 2000. Na+-dependent electrogenic ATPase from the plasma membrane of the halotolerant microalga Dunaliella maritima. Dokl. Biochem. Biophys 375, 235–238.CrossRefGoogle Scholar
  24. 24.
    Shono M., Wada M., Hara Y., Fujii T. 2001. Molecular cloning of Na+-ATPase cDNA from a marine alga Heterosigma akashiwo. Biochim. Biophys. Acta. 1511, 193–199.CrossRefPubMedGoogle Scholar
  25. 25.
    Uji T., Hirata R., Mikami K., Mizuta H., Saga N. 2012. Molecular characterization and expression analysis of sodium pump genes in the marine red alga Porphyra yezoensis. Mol. Biol. Rep. 39, 7973–7980.CrossRefPubMedGoogle Scholar
  26. 26.
    Radakovits R., Jinkerson R.E., Darzins A., Posewitz M.C. 2010. Genetic engineering of algae for enhanced biofuel production. Eukaryot. Cell. 9, 486–501.CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Pagis L.Y., Popova L.G., Andreev I.M., Balnokin Y.V. 2003. Comparative characterization of the two primary pumps, H+-ATPase and Na+-ATPase, in the plasma membrane of the marine alga Tetraselmis viridis. Physiol. Plant. 118, 514–522.CrossRefGoogle Scholar
  28. 28.
    Pedersen C.N.S., Axelsen K.B., Harper J.F., Palmgren M.G. 2012. Evolution of plant P-type ATPases. Front. Plant Sci. 3, 31. doi www.frontiersin. org doi 10.3389/fpls.2012.00031Google Scholar
  29. 29.
    Andrews S. 2010. FastQC: A quality control tool for high throughput sequence data. Google Scholar
  30. 30.
    Bolger A.M., Lohse M., Usadel B. 2014. Trimmomatic: A flexible trimmer for Illumina sequence data. Bioinformatics. 30 (15), 2114–2120.CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Haas B.J., Papanicolaou A., Yassour M., Grabherr M., Blood P.D., et al. 2013. De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nat. Protoc. 8, 1494–1512.CrossRefPubMedGoogle Scholar
  32. 32.
    Langmead B., Salzberg S.L. 2012. Fast gapped-read alignment with Bowtie 2. Nat. Methods. 9, 357–359.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Altschul S. F., Madden T.L., Schäffer A.A., Zhang J., Zhang Z., Miller W., Lipman D.J. 1997. Gapped BLAST and PSI-BLAST: A new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402.CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Mitchell A., Chang H.Y., Daugherty L., Frazer M., Hunter S., Lopez R., McAnulla C., McMenamin C., Nuka G., Pesseat S., Sangrador-Vegas A., Scheremetjew M., Rato C., Yong S.Y., Bateman A., et al. 2015. The InterPro protein families database: The classification resource after 15 years. Nucleic Acids Res. 43, D213–D221.CrossRefPubMedGoogle Scholar
  35. 35.
    Thever M.D., Milton H.S., Jr. 2009. Bioinformatic characterization of P-type ATPases encoded within the fully sequenced genomes of 26 eukaryotes. J. Membr. Biol. 229, 115–130.CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Nakamura Y., Sasaci N., Kobayashi M., Ojima N., Yasuike M., Shigenobu Y., Satomi M., Fukuma Y., Shiwaku K., Tsujimoto A., Kobayashi T., Nakayama I., Ito F., Nakajima K., Sano M., et al. 2013. The first symbiont-free genome sequence of marine red alga, susabi-nori (Pyropia yezoensis). PLoS One. 8 (3), e57122. Scholar
  37. 37.
    Raschke B.C., Wolf A.H. 1996. Molecular cloning of a P-type Ca (2+)-ATPase from the halotolerant alga Dunaliella bioculata. Planta. 200, 78–84.CrossRefPubMedGoogle Scholar
  38. 38.
    Li X., Chanroj S., Wu Z., Romanovsky S.M., Harper J.F., Sze H. 2008. A distinct endosomal Ca2+/Mn2+ pump affects root growth through the secretory process. Plant Physiol. 147, 1675–1689.CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Geisler M., Axelsen K.B., Harper J.F., Palmgren M.G. 2000. Molecular aspects of higher plant P-type Ca2+- ATPases. Biochim. Biophys. Acta. 1465, 52–78.CrossRefPubMedGoogle Scholar
  40. 40.
    Duby G., Boutry M. 2009. The plant plasma membrane proton pump ATPase: A highly regulated P-type ATPase with multiple physiological roles. Eur. J. Physiol. 457, 645–655.CrossRefGoogle Scholar
  41. 41.
    Wolf A.H., Slayman C.W., Gradmann D. 1995. Primary structure of the plasma membrane H+-ATPase from the halotolerant alga Dunaliella bioculata. Plant Mol. Biol. 28, 657–666.CrossRefPubMedGoogle Scholar
  42. 42.
    Campbell A.M., Coble A.J., Cohen L.D., Ch’ng T.H., Russo K.M., Armburst E.V. 2001. Identification and DNA sequence of a new H+-ATPase in the unicellular green alga Chlamydomonas reinhardtii (Chlorophyceae). J. Phycol. 37, 536–542.CrossRefGoogle Scholar
  43. 43.
    Poulsen L.R., Lopez-Marques R.L., Palmgren M.G. 2008. Flippases: Still more questions than answers. Cell. Mol. Life Sci. 65, 3119–3125.CrossRefPubMedGoogle Scholar
  44. 44.
    Poulsen L.R, Lopez-Marques R.L., Pedas P.R., McDowell S.C., Brown E., Kunze R., Harper J.F., Pomorski T.G., Palmgren M. 2015. A phospholipid uptake system in the model plant Arabidopsis thaliana. Nat. Commun. 6, 7649. doi doi 10.1038/ncomms8649CrossRefPubMedGoogle Scholar
  45. 45.
    Andersen J.P., Vestergaard A.L., Mikkelsen S.A., Mogensen L.S., Chalat M., Molday R. 2016. P4-ATPases as phospholipid flippases: Structure, function, and enigmas. Front. Physiol. 7, 275. doi 10.3389/fphys.2016.00275CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Møller A.B., Asp T., Holm P.B., Palmgren M.G. 2008. Phylogenetic analysis of P5 P-type ATPases, a eukaryotic lineage of secretory pathway pumps. Mol. Phylogenet. Evol. 46, 619–634.CrossRefPubMedGoogle Scholar
  47. 47.
    Oren-Shamir M., Avron M., Degani H. 1988. In vivo NMR studies of the alga Dunaliella salina embedded in beads. FEBS Lett. 233, 124–127.CrossRefGoogle Scholar
  48. 48.
    Chan C.X., Zäuner S., Wheeler G., Grossman A.R., Prochnik S.E., Blouin N.A., Zhuang Y.Z., Benning C., Berg G.M., Yarich C., Eriksen R.L., Klein A.S., Lin S., Levine I., Brawley S.H., Bhattacharya D. 2012. Analysis of Porphyra membrane transporters demonstrated gene transfer among photosynthetic eukaryotes and numerous sodium-coupled transport systems. Plant Physiol. 158, 2001–2012.CrossRefPubMedPubMedCentralGoogle Scholar
  49. 49.
    Ristani-Yazdi H., Haznedaroglu B.Z., Bibby K., Peccia J. 2011. Transcriptome sequencing and annotation of the microalga Dunaliella tertiolecta: Pathway description and gene discovery for production of next-generation biofuels. BMC Genomics. 12, 148. doi 10.1186/1471-2164-12-148CrossRefGoogle Scholar
  50. 50.
    Gimmler H., Weiss U., Weiss C., Kugel H., Treffny B. 1989. Dunaliella acidophila (Kalina) Masyuk: An alga with a positive membrane potential. New Phytol. 113d, 175–184.CrossRefGoogle Scholar
  51. 51.
    Kaim G., Dimroth P. 1994. Construction, expression and characterization of a plasmid-encoded Na+-specific ATPase hybrid consisting of Propionigenium modestum Fo-ATPase and Escherichia coli F1-ATPase. Eur. J. Biochem. 222, 615–623.CrossRefPubMedGoogle Scholar
  52. 52.
    Kholodenko B.N. 1993. Kinetic models of coupling between H+ and Na+-translocation and ATP synthesis/ hydrolysis by FoF1-ATPases: Can a cell utilize both delta mu H+ and delta mu Na+ for ATP synthesis under in vivo conditions using the same enzyme? J. Bioenerg. Biomembr. 25, 285–295.CrossRefPubMedGoogle Scholar

Copyright information

© Pleiades Publishing, Inc. 2018

Authors and Affiliations

  • L. G. Popova
    • 1
  • D. V. Belyaev
    • 1
  • A. V. Shuvalov
    • 1
  • A. A. Yurchenko
    • 2
  • D. A. Matalin
    • 1
  • D. E. Khramov
    • 1
    • 3
  • Y. V. Orlova
    • 1
  • Y. V. Balnokin
    • 1
    • 3
  1. 1.Timiryazev Institute of Plant PhysiologyRussian Academy of SciencesMoscowRussia
  2. 2.Theodosius Dobzhansky Center for Genome BioinformaticsSt. Petersburg State UniversitySt. PetersburgRussia
  3. 3.Department of BiologyMoscow State UniversityMoscowRussia

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