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Trojan Horse Strategy for Non-invasive Interference of Clock Gene in the Oyster Crassostrea gigas

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Abstract

RNA interference is a powerful method to inhibit specific gene expression. Recently, silencing target genes by feeding has been successfully carried out in nematodes, insects, and small aquatic organisms. A non-invasive feeding-based RNA interference is reported here for the first time in a mollusk bivalve, the pacific oyster Crassostrea gigas. In this Trojan horse strategy, the unicellular alga Heterocapsa triquetra is the food supply used as a vector to feed oysters with Escherichia coli strain HT115 engineered to express the double-stranded RNA targeting gene. To test the efficacy of the method, the Clock gene, a central gene of the circadian clock, was targeted for knockout. Results demonstrated specific and systemic efficiency of the Trojan horse strategy in reducing Clock mRNA abundance. Consequences of Clock disruption were observed in Clock-related genes (Bmal, Tim1, Per, Cry1, Cry2, Rev.-erb, and Ror) and triploid oysters were more sensitive than diploid to the interference. This non-invasive approach shows an involvement of the circadian clock in oyster bioaccumulation of toxins produced by the harmful alga Alexandrium minutum.

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Acknowledgements

This work was supported by the financial support of the National Research Agency (ANR), ACCUTOX project 13-CESA-0019 (2013-2017) and a ministerial PhD scholarship for L. Payton. Authors thank Dr. Denis Dupuy (IECB, ARNA-INSERM, U869, Pessac, France) for providing HT115 bacteria, Dr. Hélène Hegaret and Dr. Malwenn Lassudrie (LEMAR, UMR 6539, Plouzané, France) for providing A. minutum (strain AM89BM). Authors also thank Dr. Katherine Flynn for English corrections.

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Authors and Affiliations

  1. EPOC, UMR 5805, University of Bordeaux, F-33120, Arcachon, France

    Laura Payton, Mickael Perrigault, Anjara Marcel, Jean-Charles Massabuau & Damien Tran

  2. CNRS, EPOC, UMR 5805, Place du Dr Peyneau, 33120, Arcachon, France

    Laura Payton, Mickael Perrigault, Jean-Charles Massabuau & Damien Tran

  3. CNRS, UMR 5234, Fundamental Microbiology and Pathogenicity Laboratory, European Institute of Chemistry and Biology, University of Bordeaux, 2, rue Robert Escarpit, 33607, Pessac, France

    Jean-Paul Bourdineaud

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  1. Laura Payton
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  2. Mickael Perrigault
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  3. Jean-Paul Bourdineaud
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  5. Jean-Charles Massabuau
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Correspondence to Damien Tran.

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Payton, L., Perrigault, M., Bourdineaud, JP. et al. Trojan Horse Strategy for Non-invasive Interference of Clock Gene in the Oyster Crassostrea gigas . Mar Biotechnol 19, 361–371 (2017). https://doi.org/10.1007/s10126-017-9761-9

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