Journal of Applied Phycology

, Volume 30, Issue 6, pp 3519–3528 | Cite as

Identification and characterization of the psychrophilic bacterium CidnaK gene in the Antarctic Chlamydomoas sp. ICE-L under freezing conditions

  • Chenlin Liu
  • Xia Zhao
  • Xiuliang Wang


Heat shock protein DnaK (Hsp70) can prevent irreversible protein denaturation by chaperoning the unfolded polypeptides under extremely cold environment, and enhance cold and freezing tolerance of microorganism. From an Antarctic sea-ice green alga Chlamydomonas sp. ICE-L, eight Hsp70 genes were identified and divided into seven subfamilies: four in the cytoplasmic subfamilies, two in the chloroplast subgroups, and one in the mitochondria or endoplasmic reticulum subfamilies. Phylogenetic analyses showed that except CidnaK, each of the other Cihsp70 genes had their homologs in Chlamydomonas reinhardtii and Volvox carteri. CiDnak is a cytoplasmic protein with highly homologous to DnaK proteins from prokaryotes, and it had 100% amino acid sequence identities with the DnaK of Psychroflexus torquis ATCC 700755, a psychrophilic bacterium isolated from Antarctic sea ice. The transcription of CidnaK was significantly induced upon freezing stress in ICE-L, eminently higher than that of other Cihsp70 genes. In addition, the transformed C. reinhardtii 137c with CidnaK gene showed much higher rates of survival and growth than the wild type under freezing or low temperature conditions. Our results suggested that CidnaK was likely originated from the symbiotic bacterium in the sea-ice brine, and is an important candidate freezing inducible gene that confers low temperature tolerance to Chlamydomonas cells, and plays a crucial role in ICE-L adapting the frigid Antarctic environments.


Antarctic sea-ice Chlamydomonas Chlorophyta DnaK/Hsp70 Horizontal gene transfer Freezing tolerance 


Funding information

This work was financially supported by a grant of National Natural Science Foundation (41276203) and an Open Foundation of the State Key Laboratory of Bioactive Seaweed Substances (2017) for Chenlin Liu.


  1. Ahmad S, Selvapandiyan A, Bhatnagar RK (2000) Phylogenetic analysis of Gram-positive bacteria based on grpE, encoded by the dnaK operon. Int J Syst Evol Microbiol 50:1761–1766CrossRefGoogle Scholar
  2. Boorstein WR, Ziegelhoffer T, Craig EA (1994) Molecular evolution of the HSP70 multigene family. J Mol Evol 38:1–17CrossRefGoogle Scholar
  3. Bowman JP, McCammon SA, Lewis T, Skerratt JH, Brown JL, Nichols DS, McMeekin TA (1998) Psychroflexus torquis gen. nov., sp. nov., a psychrophilic species from Antarctic sea ice, and reclassification of Flavobacterium gondwanense (Dobson, et al. 1993) as Psychroflexus gondwanense gen. nov., comb. nov. Microbiology 144:1601–1609CrossRefGoogle Scholar
  4. Céline P, David M, Purificación LG, Céline BA (2012) Horizontal gene transfer of a chloroplast DnaJ-Fer protein to Thaumarchaeota and the evolutionary history of the DnaK chaperone system in Archaea. BMC Evol Biol 12:226CrossRefGoogle Scholar
  5. Chow KC, Tung WL (1998) Overexpression of dnaK/dnaJ and groEL confers freeze tolerance to Escherichia coli. Biochem Biophys Res Commun 253:502–505CrossRefGoogle Scholar
  6. Eardly BD, Nour SM, van Berkum P, Selander RK (2005) Rhizobial 16S rRNA and dnaK genes: Mosaicism and the uncertain phylogenetic placement of Rhizobium galegae. Appl Environ Microbiol 71:1328–1335CrossRefGoogle Scholar
  7. Feng S, Powell SM, Wilson R, Bowman JP (2014) Extensive gene acquisition in the extremely psychrophilic bacterial species Psychroflexus torquis and the link to sea-ice ecosystem specialism. Genome Biol Evol 6:133–148CrossRefGoogle Scholar
  8. Fitzpatrick DA (2012) Horizontal gene transfer in fungi. FEMS Microbiol Lett 329:1–8CrossRefGoogle Scholar
  9. Gorman DS, Levine RP (1965) Cytochrome and plastocyanin: their sequences in the photosynthetic electron transport chain of Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A 54:1665–1669CrossRefGoogle Scholar
  10. Gribaldo S, Lumia V, Creti R, de Macario EC, Sanangelantoni A, Cammarano P (1999) Discontinuous occurrence of the hsp70 (dnaK) gene among Archaea and sequence features of HSP70 suggest a novel outlook on phylogenies inferred from this protein. J Bacteriol 181:434–443PubMedPubMedCentralGoogle Scholar
  11. Hartl FU, Bracher A, Hayer-Hartl M (2011) Molecular chaperones in protein folding and proteostasis. Nature 475:324–332CrossRefGoogle Scholar
  12. Kumar S, Nei M, Dudley J, Tamura K (2008) MEGA: a biologist centric software for evolutionary analysis of DNA and protein sequences. Brief Bioinform 9:299–306CrossRefGoogle Scholar
  13. Lacroix B, Citovsky V (2016) Transfer of DNA from bacteria to eukaryotes. MBio 7:e00863–e00816CrossRefGoogle Scholar
  14. Liang WC, Wang XH, Lin MG, Lin LL (2009) A 70-kda molecular chaperone, dnak, from the industrial bacterium Bacillus licheniformis: gene cloning, purification and molecular characterization of the recombinant protein. Indian J Microbiol 49:151–160CrossRefGoogle Scholar
  15. Liu C, Huang X, Wang X, Li G (2006) Phylogenetic studies on two strains of Antarctic ice algae based on morphological and molecular characteristics. Phycologia 45:190–198CrossRefGoogle Scholar
  16. Liu S, Zhang P, Cong B, Liu C, Lin X, Shen J, Huang X (2010) Molecular cloning and expression analysis of a cytosolic Hsp70 gene from Antarctic ice algae Chlamydomonas sp. ICE-L. Extremophiles 14:329–337CrossRefGoogle Scholar
  17. Liu C, Wu G, Huang X (2012) Validation of housekeeping genes for gene expression studies in an ice alga Chlamydomonas during freezing acclimation. Extremophiles 16:419–425CrossRefGoogle Scholar
  18. Liu C, Wang X, Wang X, Sun C (2016) Acclimation of Antarctic Chlamydomonas, to the sea-ice environment: a transcriptomic analysis. Extremophiles 20:437–450CrossRefGoogle Scholar
  19. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔC T method. Methods 25:402–408CrossRefGoogle Scholar
  20. López-García P, Zivanovic Y, Deschamps P, Moreira D (2015) Bacterial gene import and mesophilic adaptation in archaea. Nat Rev Microbiol 13:447–456CrossRefGoogle Scholar
  21. Lyon BR, Mock T (2014) Polar microalgae: new approaches towards understanding adaptations to an extreme and changing environment. Biology 3:56–80CrossRefGoogle Scholar
  22. Maikova A, Zalutskaya Z, Lapina T, Ermilova E (2016) The HSP70 chaperone machines of Chlamydomonas are induced by cold stress. J Plant Physiol 204:85–91CrossRefGoogle Scholar
  23. Nordhues A, Miller SM, Muhlhaus T, Schroda M (2010) New insights into the roles of molecular chaperones in Chlamydomonas and Volvox. Int Rev Cell Mol Biol 285:75–113CrossRefGoogle Scholar
  24. Park SK, Jin ES, Lee MY (2007) Expression and antioxidant enzymes in Chaetoceros neogracile, an Antarctic alga. Cryo-Letters 29:351–361Google Scholar
  25. Polier S, Dragovic Z, Hartl FU, Bracher A (2008) Structural basis for the cooperation of Hsp70 and Hsp110 chaperones in protein folding. Cell 133:1068–1079CrossRefGoogle Scholar
  26. Powers ET, Balch WE (2013) Diversity in the origins of proteostasis networks—a driver for protein function in evolution. Nat Rev Mol Cell Biol 14:237–248CrossRefGoogle Scholar
  27. Prochnik SE, Umen J, Nedelcu AM, Hallmann A, Miller SM, Nishii I, Ferris P, Kuo A, Mitros T, Fritz-Laylin LK, Hellsten U, Chapman J, Simakov O, Rensing SA, Terry A, Pangilinan J, Kapitonov V, Jurka J, Salamov A, Shapiro H, Schmutz J, Grimwood J, Lindquist E, Lucas S, Grigoriev IV, Schmitt R, Kirk D, Rokhsar DS (2010) Genomic analysis of organismal complexity in the multicellular green alga Volvox carteri. Science 329:223–226CrossRefGoogle Scholar
  28. Proschold T, Harris EH, Coleman AW (2005) Portrait of a species: Chlamydomonas reinhardtii. Genetics 170:1601–1610CrossRefGoogle Scholar
  29. Provasoli L (1968) Media and prospects for the cultivation of marine algae. In: Watanabe A, Hattori R (eds) Culture and collections of algae. Proc. U.S.-Japan Conference, HakoneGoogle Scholar
  30. Renner T, Waters ER (2007) Comparative genomic analysis of the Hsp70s from five diverse photosynthetic eukaryotes. Cell Stress Chaperones 12:172–185CrossRefGoogle Scholar
  31. Sarkar NK, Kundnani P, Grover A (2013) Functional analysis of Hsp70 superfamily proteins of rice (Oryza sativa). Cell Stress Chaperones 18:427–437CrossRefGoogle Scholar
  32. Schönknecht G, Weber APM, Lercher MJ (2014) Horizontal gene acquisitions by eukaryotes as drivers of adaptive evolution. BioEssays 36:9–20CrossRefGoogle Scholar
  33. Soucy SM, Huang J, Gogarten JP (2015) Horizontal gene transfer: building the web of life. Nat Rev Genet 16:472–482CrossRefGoogle Scholar
  34. Sugino M, Hibino T, Tanaka Y, Nii N, Takabe T (1999) Overexpression of DnaK from a halotolerant cyanobacterium Aphanothece halophytica acquires resistance to salt stress in transgenic tobacco plants. Plant Sci 146:81–88CrossRefGoogle Scholar
  35. Sung MS, Im HN, Lee KH (2011) Molecular cloning and chaperone activity of DnaK from cold-adapted bacteria, KOPRI22215. Bull Kor Chem Soc 32:1925–1930CrossRefGoogle Scholar
  36. Takabe T, Uchida A, Shinagawa F, Terada Y, Kajita H, Tanaka Y, Takabe T, Hayashi T, Kawai T, Takabe T (2008) Overexpression of DnaK from a halotolerant cyanobacterium Aphanothece halophytica, enhances growth rate as well as abiotic stress tolerance of poplar plants. Plant Growth Regul 56:265–273CrossRefGoogle Scholar
  37. Varin T, Lovejoy C, Jungblut AD, Vincent WF, Corbeil J (2012) Metagenomic analysis of stress genes in microbial mat communities from Antarctica and the High Arctic. Appl Environ Microbiol 78:549CrossRefGoogle Scholar
  38. Xu X, Jiao L, Feng X, Ran J, Liang X, Zhao R (2017) Heterogeneous expression of DnaK gene from Alicyclobacillus acidoterrestris improves the resistance of Escherichia coli against heat and acid stress. AMB Express 7:36CrossRefGoogle Scholar
  39. Yoshimune K, Galkin A, Kulalova L, Yoshimura T, Esaki N (2005) Cold-active DnaK of an Antarctic psychrotroph Shewanella sp. Ac10 supporting the growth of dnaK-null mutant of Escherichia coli at cold temperatures. Extremophiles 9:145–150CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Key Laboratory of Marine Bioactive Substances, the First Institute of OceanographyState Oceanic AdministrationQingdaoChina
  2. 2.Laboratory for Marine Biology and BiotechnologyQingdao National Laboratory for Marine Science and TechnologyQingdaoChina
  3. 3.College of Environmental Science and EngineeringQingdao UniversityQingdaoChina
  4. 4.Key Lab of Experimental Marine Biology, Institute of OceanologyChinese Academy of SciencesQingdaoChina

Personalised recommendations