Genome-wide association identifies a missing hydrolase for tocopherol synthesis in plants

Area of Science:

• Plant biochemistry
• Molecular genetics
• Nutrient biosynthesis

Background:

• The tocopherol biosynthetic pathway is encoded by VTE genes 1-6 but many quantitative trait loci for seed tocopherols do not contain VTE genes.
• This suggests additional enzymes are involved in tocopherol synthesis.
• The precise mechanisms providing precursors for tocopherol synthesis remain incompletely understood.

Purpose of the Study:

• To identify novel genes involved in tocopherol biosynthesis.
• To characterize the function of a newly identified alpha/beta hydrolase, AtVTE7, in Arabidopsis thaliana seed tocopherol production.
• To elucidate the role of VTE7 in the context of chlorophyll and tocopherol metabolic pathways.

Main Methods:

• Genome-wide association study (GWAS) in Arabidopsis to identify genetic loci associated with seed tocopherol levels.
• Creation and analysis of null and leaky mutants for AtVTE7 and its maize ortholog, ZmVTE7.
• Complementation studies involving overexpression of AtVTE7 and ZmVTE7.
• Measurement of tocopherol and chlorophyll levels in wild-type, mutant, and overexpressing lines.
• Analysis of chlorophyll biosynthetic intermediates.

Main Results:

• A novel gene, AtVTE7, encoding a seed-specific alpha/beta hydrolase, was identified as a major quantitative trait locus for seed tocopherols.
• Atvte7 null mutants showed a 55% decrease in seed total tocopherols, and ZmVTE7 mutants exhibited reduced tocopherol levels in maize kernels and leaves.
• Overexpression of VTE7 partially or fully complemented the mutant phenotype and significantly increased leaf tocopherol levels.
• VTE7's function is linked to chlorophyll biosynthetic intermediates, not direct chlorophyll degradation, suggesting a role in releasing prenyl alcohols for tocopherol synthesis.

Conclusions:

• VTE7 is a key enzyme in tocopherol biosynthesis, acting as an esterase to release prenyl alcohols from chlorophyll pathway intermediates.
• This pathway involves a membrane-associated complex required for both chlorophyll and tocopherol synthesis.
• The findings reveal a novel link between chlorophyll biosynthesis and tocopherol production, expanding the known tocopherol pathway.