Extrachromosomal genetic engineering of the marine diatom Phaeodactylum tricornutum enables the heterologous production of monoterpenoids
Fabris, M.; George, J.; Kuzhiumparambil, U.; Lawson, C.A.; Jaramillo-Madrid, A.C.; Abbriano, R.; Vickers, C.E.; Ralph, P.J. (2020). Extrachromosomal genetic engineering of the marine diatom Phaeodactylum tricornutum enables the heterologous production of monoterpenoids. ACS Synthetic Biology 9(3): 598-612. https://dx.doi.org/10.1021/acssynbio.9b00455 In: ACS Synthetic Biology. AMER CHEMICAL SOC: Washington. ISSN 2161-5063, more |  |
| Keywords | Phaeodactylum tricornutum Bohlin, 1897 [WoRMS]
Marine/Coastal | | Author keywords | Microalgae, Phaeodactylum tricornutum, genetic engineering, monoterpenoids, extrachromosomal expression, episome |
| Authors | | Top | - Fabris, M., more
- George, J.
- Kuzhiumparambil, U.
- Lawson, C.A.
| - Jaramillo-Madrid, A.C.
- Abbriano, R.
- Vickers, C.E.
- Ralph, P.J.
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| Abstract | Geraniol is a commercially relevant plant-derived monoterpenoid that is a main component of rose essential oil and used as insect repellent. Geraniol is also a key intermediate compound in the biosynthesis of the monoterpenoid indole alkaloids (MIAs), a group of over 2000 compounds that include high-value pharmaceuticals. As plants naturally produce extremely small amounts of these molecules and their chemical synthesis is complex, industrially sourcing these compounds is costly and inefficient. Hence, microbial hosts suitable to produce MIA precursors through synthetic biology and metabolic engineering are currently being sought. Here, we evaluated the suitability of a eukaryotic microalga, the marine diatom Phaeodactylum tricornutum, for the heterologous production of monoterpenoids. Profiling of endogenous metabolism revealed that P. tricornutum, unlike other microbes employed for industrial production of terpenoids, accumulates free pools of the precursor geranyl diphosphate. To evaluate the potential for larger synthetic biology applications, we engineered P. tricornutum through extrachromosomal, episome-based expression, for the heterologous biosynthesis of the MIA intermediate geraniol. By profiling the production of geraniol resulting from various genetic and cultivation arrangements, P. tricornutum reached the maximum geraniol titer of 0.309 mg/L in phototrophic conditions. This work provides (i) a detailed analysis of P. tricornutum endogenous terpenoid metabolism, (ii) a successful demonstration of extrachromosomal expression for metabolic pathway engineering with potential gene-stacking applications, and (iii) a convincing proof-of-concept of the suitability of P. tricornutum as a novel production platform for heterologous monoterpenoids, with potential for complex pathway engineering aimed at the heterologous production of MIAs. |
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