Biotechnological application of potyvirus-based vectors in cucurbit plants
- Autores: Fakhreddine Houhou
- Directores de la Tesis: José Antonio Daròs Arnau (dir. tes.), Carmelo López del Rincón (tut. tes.)
- Lectura: En la Universitat Politècnica de València ( España ) en 2021
- Idioma: español
- Tribunal Calificador de la Tesis: Francisco Tenllado Peralo (presid.), Frederic Aparicio Herrero (secret.), Inmaculada Ferriol Safont (voc.)
- Programa de doctorado: Programa de Doctorado en Biotecnología por la Universitat Politècnica de València
- Materias:
- Enlaces
- Tesis en acceso abierto en: RiuNet
- Resumen
Nowadays, plant viruses are not only perceived as pathogens, but also able to build a beneficial partnership with their hosts and co-work together from a biotechnological view. The virus, as a vector, can be a tool to introduce heterologous genes into the plant, which will process together with the viral information and produce valuable recombinant proteins, metabolites or nanoparticles. Viral vectors are also able to interfere with plant silencing machinery giving priority to the viral genes.
Potyviruses are plant RNA viruses, mainly encoding a polyprotein of about ten mature proteins with different functions, from which the responsible of silencing suppression are helper-component proteinase (HC-Pro) and the viral protein genome-linked (VPg). In the first part of this work, we used a mild isolate of Watermelon mosaic virus (WMV; genus Potyvirus, family Potyviridae) to build a vector for virus induced gene silencing (VIGS) in melon plants. Using this virus as a vector, we expressed a fragment of melon Phytoene desaturase (PDS) mRNA in sense, antisense, and hairpin modalities to investigate the effect of the viral construct on gene silencing. The results showed a stable expression of the inserted sequence fragment in the plant in both sense and antisense orientations, whereas in the hairpin modality the insert was soon lost. Yet, all three constructs induced silencing of the endogenous PDS gene. The usefulness of the WMV for reverse genetic analysis in melon was confirmed expressing a fragment of Magnesium chelatase subunit I (CHLI). Overall, our results supported that the WMV vector is useful to apply the VIGS technology in melon and, possibly, other cucurbits.
In the second part of this work, with the aim to fortify edible fruits with health promoting metabolites, a viral RNA vector derived from Zucchini yellow mosaic virus (ZYMV; genus Potyvirus, family Potyviridae) was used to express a bacterial phytoene synthase (crtB) in zucchini (Cucurbita pepo L.) fruits. This enzyme catalyzes the first committed step of carotenoid biosynthesis. The crtB expression mediated by ZYMV resulted in the overaccumulation of a range of carotenoids and tocopherols metabolites, namely ¿- and ß-carotene (pro-vitamin A), lutein and phytoene, as well as ¿- and ¿-tocopherol (vitamin E), in both zucchini rind and flesh. This result illustrates how edible fruits can be metabolically fortified using viral vectors without plant genome modification.
