Molecular insights into a non-lethal yellow bud mutant in Ilex × ‘Nellie R. Stevens’

•The key highlights of our study include:.•The mutant had less pigment, abnormal chloroplasts, and reduced photosynthesis.•The chlorophyll and heme metabolism exhibited imbalance in the mutant.•Metabolite and gene levels were reduced in the mutant carotenoid metabolism.•Nine differentially expressed genes in porphyrin metabolism were detected. While the mechanisms behind the yellow phenotype have been widely investigated in model plant seedlings, research on leaf mutations in woody species has been restricted. The purpose of this research is to characterize a holly bud mutant of Ilex × ‘Nellie R. Stevens’ with a non-lethal yellow phenotype. Through cytological, physiological, metabolomic, and transcriptomic analyses, we identified remarkable differences between the mutant and its wild-type. The mutant exhibited abnormal chloroplasts, reduced chlorophyll levels, and elevated carotenoid/chlorophyll ratios. Metabolomic and transcriptomic analyses identified differentially expressed genes (DEGs) associated with chlorophyll and heme biosynthesis disruption, contributing to the chlorophyll-deficient phenotype. Moreover, genes associated with carotenoid metabolism were significantly down-regulated, linking to decreased carotenoid content and impacting photoprotection. Down-regulated DEGs in photosynthesis-related underscored the mutant's diminished photosynthetic activity. Additionally, the study identified 382 DEGs encoding putative transcription factors such as bHLHs, NACs, and GLKs. The study suggests an intricate interplay between chlorophyll, heme and carotenoid metabolism as major regulators of leaf mutation in I. × ‘Nellie R. Stevens.’ These insights contribute to the understanding of leaf color variation, enabling targeted strategies for developing desirable ornamental cultivars.