Dissection and modulation of isoprenoid biosynthesis in higher plants

• Autores: Xin Huang
• Directores de la Tesis: Paul Christou (dir. tes.), Teresa Capell Capell (codir. tes.)
• Lectura: En la Universitat de Lleida ( España ) en 2023
• Idioma: español
• Tribunal Calificador de la Tesis: Albert Ferrer Prats (presid.), Ludovic Bassie (secret.), Isabel Díaz Rodríguez (voc.)
• Programa de doctorado: Programa de Doctorado en Ciencia y Tecnología Agraria y Alimentaria por la Universidad de Lleida
• Enlaces
  • Tesis en acceso abierto en: TDX
• Resumen

  • My thesis focuses on identifying and characterizing the regulatory mechanisms of isoprenoid biosynthesis in higher plants by multigene engineering and gene editing. The major aim of my thesis was to better understand the isoprenoid biosynthesis pathway's bottlenecks by analyzing genes involved in isoprenoid accumulation and their functionality for future use in metabolic engineering applications. The overall objectives were to: a) enhance the production of crocins in two Nicotiana species and compare the accumulation of crocins in these species; b) investigate the regulation of the transcription factor OsBZ8 on isoprenoid biosynthetic pathway genes in rice embryo and endosperm; c) explore the potential of CRISPR/Cas9 to generate a series of maize mutants with altered strigolactone contents.

    I focused on the biosynthetic enzymes of two crocin-producing plants, (Crocus sativus) CsCCD2L and (Buddleja davidii) BdCCD4.1 and transferred them to two different Nicotiana species. I created transgenic lines expressing different combinations of transgenes and investigated the resulting crocin profiles to determine how expression of the integrated transgene complement influenced the accumulation of crocin. Engineered N. glauca plants expressing CsCCD2L alone accumulated the highest levels of crocins.

    My results indicated that N. glauca is a better host for crocin production than N. tabacum.

    I demonstrated that the transcription factor OsBZ8 and five associated genes, AACT3, HMGS1, HMGR1, DXS2, IPPI1, which are involved in the rice MVA and MEP pathways, exhibited similar expression patterns in rice endosperm and embryo. Moreover, I confirmed that OsBZ8 specifically binds to a G-box or a hybrid G/C-box in the promoters of HMGR1, DXS2 and IPPI1.

    I used CRISPR/Cas9 technology to modulate the expression of ZmCCD7 and ZmCCD8, which are involved in Strigolactones (SL) biosynthesis in maize. I was able to recover SpCas9-induced mutations in two independent plant lines in the coding region of ZmCCD8. In depth analysis of the ZmCCD8 mutants I recovered will contribute towards a better understanding of the molecular basis controlling structural traits in maize. Future genome sequencing of subsequent generations as well as metabolomic analysis is needed to assess fully the impact of these mutations. Collectively my thesis provides novel insights of the regulation of isoprenoid biosynthesis in higher plants at different levels, which will make it easier to predict the effect of metabolic engineering in plants for nutritional improvement or the production of valuable metabolites.