C3P3-G1: first generation of a eukaryotic artificial cytoplasmic expression system

Philippe Jaïs 1, * Etienne Decroly 2 Eric Jacquet 3 Marine Le boulch 1 Aurélien Jaïs 1 Olivier Jean-Jean 4 Heather Eaton 5 Prishila Ponien 3 Frédérique Verdier 6 Bruno Canard 2 Sergio Gonçalves 1 Stéphane Chiron 1 Maude Le Gall 7 Patrick Mayeux 6 Maya Shmulevitz 5
* Corresponding author
1 Génopole Campus - Eukarÿs SAS [Evry, France]
2 AFMB - Architecture et fonction des macromolécules biologiques
3 ICSN - Institut de Chimie des Substances Naturelles
4 B2A - Adaptation Biologique et Vieillissement = Biological Adaptation and Ageing
5 Medical Microbiology and Immunology [Alberta, Canada]
6 IC UM3 (UMR 8104 / U1016) - Institut Cochin
7 Centre de recherche sur l'inflamation - UMR 1149
Abstract : Most eukaryotic expression systems make use of host-cell nuclear transcriptional and post-transcriptional machineries. Here, we present the first generation of the chimeric cytoplasmic capping-prone phage polymerase (C3P3-G1) expression system developed by biological engineering, which generates capped and polyadenylated transcripts in host-cell cytoplasm by means of two components. First, an artificial single-unit chimeric enzyme made by fusing an mRNA capping enzyme and a DNA-dependent RNA polymerase. Second, specific DNA templates designed to operate with the C3P3-G1 enzyme , which encode for the transcripts and their artificial polyadenylation. This system, which can potentially be adapted to any in cellulo or in vivo eu-karyotic expression applications, was optimized for transient expression in mammalian cells. C3P3-G1 shows promising results for protein production in Chinese Hamster Ovary (CHO-K1) cells. This work also provides avenues for enhancing the performances for next generation C3P3 systems.
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Philippe Jaïs, Etienne Decroly, Eric Jacquet, Marine Le boulch, Aurélien Jaïs, et al.. C3P3-G1: first generation of a eukaryotic artificial cytoplasmic expression system. Nucleic Acids Research, Oxford University Press, 2019, 47 (5), pp.2681-2698. ⟨10.1093/nar/gkz069⟩. ⟨inserm-02016614⟩

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