Integrated transcriptomic and metabolomic analysis reveals drivers of protein and oil variation in cottonseed.

The protein and oil content in cottonseed, known for their high quality, exhibits substantial variation across different cotton varieties. This study explored the regulatory mechanisms behind these differences by analyzing protein and oil accumulation patterns, transcriptomics, and metabolomics in two cotton varieties during seed development. Results showed that protein and oil rapidly accumulated between 15 and 30 days post-anthesis (DPA), but significant differences between varieties emerged after 40 DPA. Differentially expressed genes (DEGs) at 40 DPA were enriched in carbon allocation, fatty acid degradation, and nitrogen absorption pathways. Metabolomics identified lipids, lipid-like molecules, and organic acids as key differentially accumulated metabolites (DAMs). Furthermore, the gene , associated with nitrogen source absorption, was identified. Virus-induced gene silencing (VIGS) of this gene in cotton resulted in a significant reduction in protein content in the roots, stems, and leaves. These findings provide insights into protein and oil accumulation and offer genetic resources for improving cottonseed nutritional quality.