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Successful reversal of transgene silencing in Chlamydomonas reinhardtii
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  • Remy Beauchemin,
  • Natacha Mérindol,
  • Elisa I. Fantino,
  • Pamela Lavoie,
  • Serge Basile Nouemssi,
  • Fatma Meddeb-Mouelhi,
  • Isabel Desgagné-Penix
Remy Beauchemin
Université du Québec à Trois-Rivières
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Natacha Mérindol
Université du Québec à Trois-Rivières
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Elisa I. Fantino
Université du Québec à Trois-Rivières
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Pamela Lavoie
Université du Québec à Trois-Rivières
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Serge Basile Nouemssi
Université du Québec à Trois-Rivières
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Fatma Meddeb-Mouelhi
Université du Québec à Trois-Rivières
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Isabel Desgagné-Penix
Universite du Quebec a Trois-Rivieres

Corresponding Author:[email protected]

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Chlamydomonas reinhardtii has been successfully engineered to produce compounds of interest following transgene integration and heterologous protein expression. The advantages of this model include the availability of validated tools for bioengineering, its photosynthetic ability and its potential use as biofuel. Despite this, breakthroughs have been hindered by its ability to silence transgene expression through epigenetic changes. Histone deacetylases (HDAC) are main players in gene expression. We hypothesized that transgene silencing can be reverted with chemical treatments using HDAC inhibitors. To analyze this, we transformed C. reinhardtii, integrating into its genome the mVenus reporter gene under the HSP70-rbcs2 promoter. From 384 transformed clones, 88 (22.9 %) displayed mVenus positive (mVenus+) cells upon flow-cytometry analysis. Five clones with different fluorescence intensities were selected. The number of integrated copies was measured by qPCR. Transgene expression levels were followed over the growth cycle and upon SAHA treatment, using a microplate reader, flow cytometry, RT-qPCR, and western blot analysis. First, we observed that expression varies with the cell cycle, reaching a maximum level just before the stationary phase in all clones. Second, we uncovered that supplementation with HDAC inhibitors of the hydroxamate family, such as vorinostat (suberoylanilide-hydroxamic-acid, SAHA) at the initiation of culture increases the frequency (% of mVenus+ cells) and the level of transgene expression per cell over the whole growth cycle, through histone deacetylase inhibition. Thus, we propose a new tool to successfully trigger the expression of heterologous proteins in the green algae C. reinhardtii, overcoming its main handicap as an expression platform.
22 May 2023Submitted to Biotechnology Journal
24 May 2023Submission Checks Completed
24 May 2023Assigned to Editor
25 May 2023Reviewer(s) Assigned
26 Jun 2023Review(s) Completed, Editorial Evaluation Pending
27 Jun 2023Editorial Decision: Revise Major
13 Sep 20231st Revision Received
14 Sep 2023Submission Checks Completed
14 Sep 2023Assigned to Editor
14 Sep 2023Reviewer(s) Assigned
15 Nov 2023Review(s) Completed, Editorial Evaluation Pending
15 Nov 2023Editorial Decision: Accept