Advertisement

Journal of Ocean University of China

, Volume 18, Issue 4, pp 977–984 | Cite as

Characterization and Expression Analysis of Protein Kinase C Gene from Dunaliella salina

  • Yuting Cong
  • Yuexin Ma
  • Yuan Wang
  • Yiqiong Liu
  • Xiaojie ChaiEmail author
Article
  • 2 Downloads

Abstract

Protein kinase C (PKC) has a crucial role in signal transduction for a variety of biologically active substances which activate cellular functions and proliferation. We previously isolated the full-length PKC gene from Dunaliella salina (DsPKC) using rapid amplification of cDNA ends (RACE) and RT-PCR methods. And we submitted the mRNA sequence of DsPKC gene to NCBI (Genbank No. JN625213). In the present paper, the DsPKC gene open reading frame obtained by PCR was cloned into pGS-21a vector and transformed into Escherichia coli to generate the fusion protein. Bioinformatics analysis revealed that DsPKC gene was a member of serine/threonine kinase with two conserved domains and highly conserved motifs. The DsPKC was highly expressed upon induction with isopropyl-β-d-thiogalactoside (IPTG) at a final concentration of 0.2mmolL−1 at 37°C. Under salt stress, the fusion protein Green Fluorescent Protein (GFP)-DsPKC was transferred from the cytoplasm to the cell membrane. The expression pattern of DsPKC gene was analyzed using real-time quantitative PCR, and indicated that DsPKC gene was up-regulated by 3.0molL−1 NaCl at 12h, which was significantly higher than in control values (P < 0.05). These results suggest that the DsPKC gene plays an important role in response to salt stress in D. salina.

Key words

Dunaliella salina protein kinase C gene prokaryotic expression subcellular localization salt stress 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Notes

Acknowledgements

This study was supported by the functional analysis of PKC signaling pathway involved in response to salt stress of Dunaliella salina, the National Natural Science Foundation of China (No. 31472260).

References

  1. Arroussi, H. E., Benhima, R., Elbaouchi, A., Sijilmassi, B., Mernissi, N. E., Aafsar, A., Meftah-Kadmiri, I., Bendaou, N., and Smouni, A., 2018. Dunaliella salina exopolysaccharides: A promising biostimulant for salt stress tolerance in tomato (Solanum lycopersicum). Journal of Applied Phycology, 30 (5): 2929–2941.CrossRefGoogle Scholar
  2. Avgay, Y., and Everett, M., 2000. The eukaryotic-like Ser/Thr protein kinases of Mycobacterium tuberculosis. Trends in Microbiology, 8 (5): 238–244.CrossRefGoogle Scholar
  3. Chen, H., Jiang, J. G., and Wu, G. H., 2009. Effects of salinity changes on the growth of Dunaliella salina and its isozyme activities of glycerol-3-phosphate dehydrogenase. Journal of Agricultural and Food Chemistry, 57 (14): 6178–6182.CrossRefGoogle Scholar
  4. Cui, L. Q., Chai, Y. R., Li, J., Liu, H. T., Zhang, L., and Xue, L. X., 2010. Identification of a glucose-6-phosphate isomerase involved in adaptation to salt stress of Dunaliella salina. Journal of Applied Phycology, 22 (5): 563–568.CrossRefGoogle Scholar
  5. Darieva, Z., Webber, A., Warwood, S., and Sharrocks, A. D., 2015. Protein kinase C coordinates histone H3 phosphorylation and acetylation. Elife, 4: e09886.CrossRefGoogle Scholar
  6. Dong, X. F., Cui, N., Wang, L., Zhao, X. C., Qu, B., Li, T. L., and Zhang, G. L., 2012. The SnRK protein kinase family and the function of SnRK1 protein kinase. International Journal of Agriculture and Biology, 14 (4): 575–579.Google Scholar
  7. Emanuelsson, O., 2000. Predicting subcellular localization of proteins based on their amino acid sequence. Journal of Molecular Biology, 300 (4): 1005–1016.CrossRefGoogle Scholar
  8. Fei, X., 2013. Cloning and expression analysis of serine/threonine protein kisnase gene DsSTPK from Dunaliella salina. Doctoral dissertation. Dalian Ocean University, Dalian.Google Scholar
  9. Geraldes, P., and King, G. L., 2010. Activation of protein kinase C isoforms and its impact on diabetic complications. Circulation Research, 106 (8): 1319.CrossRefGoogle Scholar
  10. Goyal, A., 2007. Osmoregulation in Dunaliella, Part II: Photosynthesis and starch contribute carbon for glycerol synthesis during a salt stress in Dunaliella tertiolecta. Plant Physiology and Biochemistry, 45 (9): 705–710.CrossRefGoogle Scholar
  11. Han, J. H., Choi, Y. S., Kim, W. J., Jeon, Y. H., Lee, S. K., Lee, B. J., and Ryu, K. S., 2010. Codon optimization enhances protein expression of human peptide deformylase in E. coli. Protein Expression and Purification, 70 (2): 224–230, DOI:  https://doi.org/10.1016/j.pep.2009.10.005.CrossRefGoogle Scholar
  12. Hopes, A., Nekrasov, V., Kamoun, S., and Mock, T., 2016. Editing of the urease gene by CRISPR-Cas in the diatom Thalassiosira pseudonana. Plant Methods, 12 (1): 49.CrossRefGoogle Scholar
  13. Jahnke, L. S., and White, A. L., 2003. Long-term hyposaline and hypersaline stresses produce distinct antioxidant responses in the marine alga Dunaliella tertiolecta. Journal of Plant Physiology, 160 (10): 1193–1202.CrossRefGoogle Scholar
  14. Kishimoto, A., Takai, Y., and Nishizuka, Y., 1977. Activation of glycogen phosphorylase kinase by a calcium-activated, cyclic nucleotide-independent protein kinase system. Journal of Biological Chemistry, 252 (21): 7449–7452.Google Scholar
  15. Liu, Y., and Yildiz, I., 2018. The effect of salinity concentration on algal biomass production and nutrient removal from municipal wastewater by Dunaliella salina. International Journal of Energy Research, (3): 2997–3006.Google Scholar
  16. Manning, G., Whyte, D. B., Martinez, R., Hunter, T., and Sudarsanam, S., 2002. The protein kinase complement of the human genome. Science, 298 (5600): 1912–1934.CrossRefGoogle Scholar
  17. Mellor, H., and Parker, P. J., 1998. The Extended Protein Kinase C Family. Oxford University Press, New York, 281–292.Google Scholar
  18. Muellerroeber, B., and Pical, C., 2002. Inositol phospholipid metabolism in Arabidopsis. Characterized and putative iso-forms of inositol phospholipid kinase and phosphoinosi-tidespecific phospholipase C. Plant Physiology, 130 (1): 22–46.CrossRefGoogle Scholar
  19. Nishizuka, Y., 1986. Studies and perspectives of protein kinase C. Science, 233 (4761): 305.CrossRefGoogle Scholar
  20. Nishizuka, Y., 1992. Intracellular signaling by hydrolysis of phospholipids and activation of protein kinase C. Science, 258 (5082): 607–614.CrossRefGoogle Scholar
  21. Punta, M., Coggill, P. C., Eberhardt, R. Y., Mistry, J., Tate, J., Boursnell, C., Pang, N., Forslund, K., Ceric, G., and Clements, J., 2012. The Pfam protein families database. Nucleic Acids Research, 32: D138.Google Scholar
  22. Qiang, L., Yong, Z., and Shouyi, C., 2000. Plant protein kinase genes induced by drought, high salt and cold stresses. Science, 45 (13): 1153–1157.Google Scholar
  23. Shinozaki, K., and Yamaguchishinozaki, K., 1997. Gene expression and signal transduction in water-stress response. Plant Physiology, 115 (2): 327.CrossRefGoogle Scholar
  24. Shukla, V., and Mattoo, A. K., 2008. Sucrose non-fermenting 1-related protein kinase 2 (SnRK2): A family of protein kinases involved in hyperosmotic stress signaling. Physiology and Molecular Biology of Plants, 14 (1–2): 91,  https://doi.org/10.1007/s12298-008-0008-0.CrossRefGoogle Scholar
  25. Wang, P., Xue, L., Batelli, G., Lee, S., Hou, Y. J., Oosten, M. J. V., Zhang, H., Tao, W. A., and Zhu, J. K., 2013. Quantitative phosphoproteomics identifies SnRK2 protein kinase substrates and reveals the effectors of abscisic acid action. Proceedings of the National Academy of Sciences of the United States of America, 110 (27): 11205–11210.CrossRefGoogle Scholar
  26. Wei, S., Bian, Y., Zhao, Q., Chen, S., Mao, J., Song, C., Cheng, K., Xiao, Z., Zhang, C., and Ma, W., 2017. Salinity-induced palmella formation mechanism in halotolerant algae Dunaliella salina revealed by quantitative proteomics and phosphoproteomics. Frontiers in Plant Science, 8: 810, DOI:  https://doi.org/10.3389/fpls.2017.00810.CrossRefGoogle Scholar
  27. Xie, H., Xu, P., Jia, Y., Li, J., Lu, Y., and Xue, L., 2007. Cloning and heterologous expression of nitrate reductase genes from Dunaliella salina. Journal of Applied Phycology, 19 (5): 497–504.CrossRefGoogle Scholar
  28. Xue, F., Chai, X. J., Yu, Z. J., Zhang, X. L., and Zhang, T., 2012. Cloning and bioinformatics analysis of a novel gene DsSTPK from Dunaliella salina. Chinese Journal of Bio-chemistry and Molecular Biology, 28 (3): 289–293.Google Scholar
  29. Yang, P., Baciu, P., Kerrigan, B. C., Etheridge, M., Sung, E., Toimil, B. A., Berchuck, J. E., and Jaffe, G. J., 2014. Retinal pigment epithelial cell death by the alternative complement cascade: Role of membrane regulatory proteins, calcium, PKC, and oxidative stress. Investigative Ophthalmology and Visual Science, 55 (5): 3012–3021.CrossRefGoogle Scholar
  30. Zhu, J. K., 2003. Salt and drought stress signal transduction in plants. Annual Review of Plant Biology, 53: 247–273.CrossRefGoogle Scholar

Copyright information

© Ocean University of China, Science Press and Springer-Verlag GmbH Germany 2019

Authors and Affiliations

  • Yuting Cong
    • 1
  • Yuexin Ma
    • 2
  • Yuan Wang
    • 1
  • Yiqiong Liu
    • 1
  • Xiaojie Chai
    • 1
    Email author
  1. 1.Key Laboratory of Hydrobiology in Liaoning Province’s UniversityDalian Ocean UniversityDalianChina
  2. 2.Key Laboratory of Mariculture & Stock Enhancement in North China’s Sea, Ministry of AgricultureDalian Ocean UniversityDalianChina

Personalised recommendations