Metabolome and Transcriptome Unveil the Correlated Metabolites and Transcripts with 2-acetyl-1-pyrroline in Fragrant Rice

Fragrance is a valuable trait in rice varieties, with its aroma significantly influencing consumer preference. In this study, we conducted comprehensive metabolome and transcriptome analyses to elucidate the genetic and biochemical basis of fragrance in the Shangsixiangnuo (SSXN) variety, a fragrant indica rice cultivated in Guangxi, China. Through sensory evaluation and genetic analysis, we confirmed SSXN as strongly fragrant, with an 806 bp deletion in the BADH2 gene associated with fragrance production. In the metabolome analysis, a total of 238, 233, 105 and 60 metabolic compounds exhibited significant changes at the seedling (S), reproductive (R), filling (F), and maturation (M) stages, respectively. We identified four compounds that exhibited significant changes in SSXN across all four development stages. Our analyses revealed a significant upregulation of 2-acetyl-1-pyrroline (2AP), the well-studied aromatic compound, in SSXN compared to the non-fragrant variety. Additionally, correlation analysis identified several metabolites strongly associated with 2AP, including ethanone, 1-(1H-pyrrol-2-yl)-, 1H-pyrrole, and pyrrole. Furthermore, Weighted Gene Co-expression Network Analysis (WGCNA) analysis highlighted the magenta and yellow modules as particularly enriched in aroma-related metabolites, providing insights into the complex aromatic compounds underlying the fragrance of rice. In the transcriptome analysis, a total of 5582, 5506, 4965, and 4599 differential expressed genes (DEGs) were identified across the four developmental stages, with a notable enrichment of the common pathway amino sugar and nucleotide sugar metabolism in all stages. In our correlation analysis between metabolome and transcriptome data, the top three connected metabolites, phenol-, 3-amino-, and 2AP, along with ethanone, 1-(1H-pyrrol-2-yl)-, exhibited strong associations with transcripts, highlighting their potential roles in fragrance biosynthesis. Additionally, the downregulated expression of the P4H4 gene, encoding a procollagen-proline dioxygenase that specifically targets proline, in SSXN suggests its involvement in proline metabolism and potentially in aroma formation pathways. Overall, our study provides comprehensive insights into the genetic and biochemical mechanisms underlying fragrance production in rice, laying the foundation for further research aimed at enhancing fragrance quality in rice breeding programs.