Integrated metabolomic and transcriptomic analysis reveals the key gene CitF3‘H-like involving in the biosynthesis of nobiletin in citrus

Bitterness is regarded as one of the crucial characteristics influencing citrus fruit flavor and post-harvest processing. The novel citrus cultivar ‘Yangguang No.1’ (YG) exhibits a relatively reduced level of bitterness, particularly in the peel, compared to its parent (‘Chunxiang’, CX). In this study, integrated metabolomic and transcriptomic analyses were employed to investigate the disparities in bitterness between YG and CX. Metabolomic profiling, conducted using a widely targeted ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) approachd, revealed that flavonoid compounds associated with bitterness, particularly nobiletin and tangeretin, accumulated at significantly lower levels in YG. Transcriptome analysis indicated that differentially expressed genes in YG relative to CX were markedly enriched in metabolic pathways including phenylpropanoid biosynthesis and flavonoid biosynthesis. Weighted gene co-expression network analysis (WGCNA) revealed that the expression of CitF3’H , CitF3’H-like and CitOMT2/3/5/10/11/12 was closely correlated with the accumulation of bitter flavonoids. Functional validation showed that overexpression of CitF3’H-like in YG peel resulted in increased nobiletin content, whereas virus-induced gene silencing (VIGS) of CitF3’H-like in CX seedlings led to a significant reduction in nobiletin levels. These findings demonstrated that CitF3’H-like played a functional role in the biosynthesis of nobiletin in citrus. Collectively, this study identifies CitF3’H-like as a key candidate gene involved in the accumulation of bitter compound, and provides valuable insights for breeding citrus varieties suitable for post-harvest processing through reduced bitterness.