Abstract
Larvae of the African midge Polypedilum vanderplanki show extreme desiccation tolerance, known as anhydrobiosis. Recently, the cultured cell line Pv11 was derived from this species; Pv11 cells can be preserved in the dry state for over 6 months and retain their proliferation potential. Here, we attempted to expand the use of Pv11 cells as a model to investigate the mechanisms underlying anhydrobiosis in P. vanderplanki. A newly developed vector comprising a constitutive promoter for the PvGapdh gene allowed the expression of exogenous proteins in Pv11 cells. Using this vector, a stable Pv11 cell line expressing green fluorescence protein (GFP) was established and retained desiccation tolerance. Gene silencing with GFP-specific siRNAs significantly suppressed GFP expression to approximately 7.5–34.6% of that in the non-siRNA-transfected GFP stable line. Establishment of these functional assays will enable Pv11 cells to be utilized as an effective tool to investigate the molecular mechanisms underlying anhydrobiosis.
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Acknowledgements
We are grateful to R. Hatanaka and T. Furusawa for helping perform plasmid construction and cell-sorting, respectively. We also thank T. Shirotori and Y. Kikuzato for technical assistance. This work was supported in part by the Grants-in-Aid from Ministry of Education, Culture, Sports, Science and Technology (MEXT)/Japan Society for the Promotion of Science (JSPS) KAKENHI (Grant Numbers 16K18827, 16J09151, 16K07308, 15H02378, 25252060, and 16K15073); research fellowship of JSPS for Young Scientists (#13J08784); a Grant for Basic Science Research Projects from the Sumitomo Foundation (140890); a Grant for the Narishige Zoological Science Award 2016; construction of siRNA and plasmids were supported by Russian Science Foundation grant for international groups (No. 14-44-00022).
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Communicated by H. Atomi.
This article is part of a special feature based on the 11th International Congress on Extremophiles held in Kyoto, Japan, September 12–16, 2016.
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Sogame, Y., Okada, J., Kikuta, S. et al. Establishment of gene transfer and gene silencing methods in a desiccation-tolerant cell line, Pv11. Extremophiles 21, 65–72 (2017). https://doi.org/10.1007/s00792-016-0880-4
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DOI: https://doi.org/10.1007/s00792-016-0880-4