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A 14-bp stretch plays a critical role in regulating gene expression from β1-tubulin promoters of basidiomycetes

  • Dong Xuan Nguyen
  • Taku Sakaguchi
  • Takehito Nakazawa
  • Masahiro Sakamoto
  • Yoichi HondaEmail author
Original Article

Abstract

Cis-acting elements play a vital role in regulation of transcription initiation. Several cis-acting elements have been identified in filamentous fungi; however, the fundamental requirements for basic promoter function in basidiomycetes are obscure. In this study, core elements in β1-tubulin promoters of basidiomycetes were functionally characterized. Using transient transformation in Ceriporiopsis subvermispora as a promoter assay, we found that a 14-bp region (β1-tubulin core promoter element, BCE), as well as CT-rich stretch, in the β1-tubulin promoter of the species played a critical role in the expression of a recombinant hph as a reporter gene. In addition, in silico analysis revealed other members of basidiomycetes also harboured the BCE motif as well as CT-rich stretch in the β1-tubulin promoter region, suggesting their functional conservation among the species of basidiomycetes. To confirm the function of BCE, we investigated the effects of BCE motif deletion in the Pleurotus ostreatus β1-tubulin promoter on expression levels of a recombinant luminous shrimp luciferase reporter gene, which was targeted into the Pofcy1 locus. Intriguingly, luciferase activity was abolished when the BCE motif was deleted in the β1-tubulin promoter, strongly demonstrating its essential function in transcription from this promoter on the chromosome. This study clearly demonstrates the crucial role of the BCE as well as the CT-rich stretch regions in the β1-tubulin promoter among basidiomycetes and provides new insights into the fundamental mechanism of transcription initiation in this group.

Keywords

Basidiomycetes Ceriporiopsis subvermispora Pleurotus ostreatus β1-tubulin gene Transient transformation Core promoter element 

Notes

Acknowledgements

This work was supported in part by the Noda Institute for Scientific Research (to Y. H.), and JSPS KAKENHI [15K14771 and 18H02254 to Y. H.]. We would like to thank Prof. Yitzhak Hadar (Hebrew University of Jerusalem, Israel) for providing P. ostreatus strains 20b, Profs. Kazumitsu Ueda and Noriyuki Kioka (Kyoto University, Japan) for help in measurement of luciferase activity, Prof. Keishi Osakabe (Tokushima University, Japan) for providing the plasmid containing the luciferase reporter gene.

Supplementary material

294_2019_1014_MOESM1_ESM.pdf (1.1 mb)
Supplementary material 1 (PDF 1146 kb)
294_2019_1014_MOESM2_ESM.docx (4.7 mb)
Supplementary material 2 (DOCX 4782 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Graduate School of AgricultureKyoto UniversityKyotoJapan
  2. 2.Biotechnology Center of Ho Chi Minh CityHo Chi Minh CityVietnam

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