Leaky mutations in the zeaxanthin epoxidase in Capsicum annuum result in bright‐red fruit containing a high amount of zeaxanthin

• Published in: The Plant journal : for cell and molecular biology 2024-04, Vol.118 (2), p.469-487
• Main Authors: Lee, Seo‐Young, Jang, So‐Jeong, Jeong, Hyo‐Bong, Lee, Joung‐Ho, Kim, Geon Woo, Venkatesh, Jelli, Back, Seungki, Kwon, Jin‐Kyung, Choi, Da‐Min, Kim, Jeong‐Il, Kim, Geun‐Joong, Kang, Byoung‐Cheorl
• Format: Article
• Language: English
• Subjects: Abscisic acid; aesthetics; Amino acid sequence; amino acid sequences; Amino acids; biosynthesis; capsanthin; Capsicum - genetics; Capsicum - metabolism; Capsicum annuum; carotenoid; Carotenoids; Carotenoids - metabolism; Chromosome 2; Color; Complementation; Cultivars; E coli; Enzymatic activity; enzyme activity; Escherichia coli; ethyl methanesulfonate; Fruit - metabolism; fruit color; Fruits; Gene silencing; Genes; Germination; Germplasm; Hormones; Liquid chromatography; loci; Loss of Function Mutation; mature fruit color; Missense mutation; mutants; Mutation; MutMap; Oxidoreductases; pepper; Peppers; Plant Breeding; Plant hormones; salt stress; Seeds; surveys; Thylakoid membranes; thylakoids; Xanthophylls; Zeaxanthin; Zeaxanthin epoxidase; Zeaxanthins - metabolism
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1111/tpj.16619

SUMMARY Fruit color is one of the most important traits in peppers due to its esthetic value and nutritional benefits and is determined by carotenoid composition, resulting from diverse mutations of carotenoid biosynthetic genes. The EMS204 line, derived from an EMS mutant population, presents bright‐red color, compared with the wild type Yuwolcho cultivar. HPLC analysis indicates that EMS204 fruit contains more zeaxanthin and less capsanthin and capsorubin than Yuwolcho. MutMap was used to reveal the color variation of EMS204 using an F3 population derived from a cross of EMS204 and Yuwolcho, and the locus was mapped to a 2.5‐Mbp region on chromosome 2. Among the genes in the region, a missense mutation was found in ZEP (zeaxanthin epoxidase) that results in an amino acid sequence alteration (V291 → I). A color complementation experiment with Escherichia coli and ZEP in vitro assay using thylakoid membranes revealed decreased enzymatic activity of EMS204 ZEP. Analysis of endogenous plant hormones revealed a significant reduction in abscisic acid content in EMS204. Germination assays and salinity stress experiments corroborated the lower ABA levels in the seeds. Virus‐induced gene silencing showed that ZEP silencing also results in bright‐red fruit containing less capsanthin but more zeaxanthin than control. A germplasm survey of red color accessions revealed no similar carotenoid profiles to EMS204. However, a breeding line containing a ZEP mutation showed a very similar carotenoid profile to EMS204. Our results provide a novel breeding strategy to develop red pepper cultivars containing high zeaxanthin contents using ZEP mutations. Significance Statement Despite health benefits, such as macular protection and antioxidant effects, most crops contain low levels of zeaxanthin. Orange pepper is an important source of zeaxanthin which is scarce in our diet. We demonstrated that a leaky mutation in zeaxanthin epoxidase leads to the development of bright‐red fruit, elevated zeaxanthin levels, and affects endogenous plant hormones in pepper. Our findings suggest a novel breeding strategy for developing red pepper cultivars with high levels of zeaxanthin.