In-planta analyses of the three Arabidopsis isoforms of the bifunctional RIBA with enzymatic domains for GTP cyclohydrolase II and 3,4-dihydroxy-2-butanone 4-phosphate synthase

• Applicant: Professor Dr. Bernhard Grimm
• Subject Area: Plant Physiology
• Term: from 2014 to 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 258667322

Riboflavin serves as precursor for flavocoenzymes (FMN and FAD) and is essential for all living organisms. The initial two committed enzymatic steps of riboflavin biosynthesis are performed in plants by bifunctional RIBA enzymes comprising GTP cyclohydrolase II (GCHII) and 3,4-dihydroxy-2-butanone-4-phosphate synthase (DHBPS). In Angiosperms the RIBA gene family consists of three members. Starting our studies in plant riboflavin biosynthesis, we found that strongly reduced AtRIBA1 expression in an Arabidopsis thaliana rfd1 mutant (T-DNA insertion in RIBA1) and in RIBA1 antisense lines cannot be complemented by the simultaneously expressed AtRIBA2 AtRIBA3. Only little information is available on biochemistry and control of plant riboflavin metabolism. By means of the proposed working program it is aimed to verify the function of all three RIBA isoforms in riboflavin biosynthesis. It is intended to examine in planta the enzymatic properties and the physiological importance of the three bipartite wild-type and multiple engineered RIBA variants, which will be expressed in a lethal riba mutant. In addition, transgenic Arabidopsis plants with silenced and overexpressed RIBA1 genes will be analysed on the modified content of flavins and the flavocoenzyme-dependent proteins and their cellular processes. Screens for RIBA-interaction partners are aimed at to decode the function, organisation and control of riboflavin biosynthesis and the interrelation to other subcellular processes. In preliminary experiments RIBA1 and RIBA2 phosphorylation was achieved with plastid extracts. In continuation it is planned to identify the phosphorylation site of the RIBA proteins and unravel the physiological consequences of RIBA phosphorylation in Arabidopsis plants.

• DFG Programme: Research Grants
• Participating Persons: Professor Alisdair Fernie, Ph.D.; Professor Dr. Markus Fischer; Dr. Michael Melzer