Genetic Dissection and Functional Validation of DXS2 Controlling Antheraxanthin Content in Maize Kernels

Antheraxanthin (ANT) is an important xanthophyll carotenoid in maize, but its genetic basis in maize kernels remains poorly understood. Here, we investigated the genetic architecture of ANT content using a teosinte-maize BC₂F₅ population evaluated across multiple environments. ANT showed abundant phenotypic variation and high broad-sense heritability (H² = 0.86), indicating strong genetic control. A total of 11 QTLs were identified on chromosomes 1, 4, 5, 7, and 8, among which the major-effect locus L4 on chromosome 7 was consistently detected across all environments and explained up to 37.46% of phenotypic variation. Fine mapping and candidate gene analysis identified DXS2, encoding 1-deoxy-D-xylulose 5-phosphate synthase 2, which is a key rate-limiting enzyme in the MEP pathway, as the strongest causal gene underlying L4. Functional validation using a MuDR insertion mutant demonstrated that loss of DXS2 function significantly reduced ANT accumulation in maize kernels. Transcriptomic analysis further revealed that DXS2 influences the MEP pathway, carotenoid biosynthesis, and downstream metabolic networks. These findings elucidate the genetic architecture of ANT accumulation, support DXS2 as a key regulator connecting the MEP pathway with downstream carotenoid and ANT metabolism, and provide valuable genetic resources and theoretical support for carotenoid biofortification in maize breeding.