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UV-mediated Chlamydomonas mutants with enhanced nuclear transgene expression by disruption of DNA methylation-dependent and independent silencing systems

Plant Molecular Biology volume 92, pages 629–641 (2016)Cite this article

Abstract

Key message

In this investigation, we succeeded to generate Chlamydomonas mutants that bear dramatically enhanced ability for transgene expression. To yield these mutants, we utilized DNA methyltransferase deficient strain. These mutants must be useful as a plant cell factory.

Abstract

Chlamydomonas reinhardtii (hereafter Chlamydomonas) is a green freshwater microalga. It is a promising cell factory for the production of recombinant proteins because it rapidly grows in simple salt-based media. However, expression of transgenes integrated into the nuclear genome of Chlamydomonas is very poor, probably because of severe transcriptional silencing irrespective of the genomic position. In this study, we generated Chlamydomonas mutants by ultraviolet (UV)-mediated mutagenesis of maintenance-type DNA methyltransferase gene (MET1)-null mutants to overcome this disadvantage. We obtained several mutants with an enhanced ability to overexpress various transgenes irrespective of their integrated genomic positions. In addition, transformation efficiencies were significantly elevated. Our findings indicate that in addition to mechanisms involving MET1, transgene expression is regulated by a DNA methylation-independent transgene silencing system in Chlamydomonas. This is in agreement with the fact that DNA methylation occurs rarely in this organism. The generated mutants may be useful for the low-cost production of therapeutic proteins and eukaryotic enzymes based on their rapid growth in simple salt-based media.

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Acknowledgments

Special thanks to Ms. Miyuki Nakajima for her encouragement.

Funding:

This research was partly supported by the Japan Science and Technology Agency (JST) and Core Research for Evolutional Science and Technology (CREST).

Author conrtibutions

T.Y. and T.O. designed the research, S.D.K., F.K., and D.W. performed experiments, T.O. wrote the manuscript with feedback from S.O.

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Authors and Affiliations

  1. School of Environmental Science and Engineering, Kochi University of Technology, Tosayamada, Kochi, 782-8502, Japan

    Sari Dewi Kurniasih, Tomohito Yamasaki, Fantao Kong, Dwiyantari Widyaningrum & Takeshi Ohama

  2. Laboratory of Aquatic Natural Products Chemistry, Graduate School of Agricultural & Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo, 113-8657, Japan

    Sigeru Okada

Authors
  1. Sari Dewi Kurniasih
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  2. Tomohito Yamasaki
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  3. Fantao Kong
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  4. Sigeru Okada
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  5. Dwiyantari Widyaningrum
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  6. Takeshi Ohama
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Correspondence to Takeshi Ohama.

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Kurniasih, S.D., Yamasaki, T., Kong, F. et al. UV-mediated Chlamydomonas mutants with enhanced nuclear transgene expression by disruption of DNA methylation-dependent and independent silencing systems. Plant Mol Biol 92, 629–641 (2016). https://doi.org/10.1007/s11103-016-0529-9

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  • DOI: https://doi.org/10.1007/s11103-016-0529-9

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