Integrated Transcriptomics and Metabolomics with Machine Learning Identify Flavonoids as Key Effectors in Wheat Root Thermotolerance.

Root plasticity is vital for crop survival amid global warming. Yet, the molecular mechanisms governing wheat root thermotolerance remain largely unknown. In this study, we combined phenomics, transcriptomics, and metabolomics with machine learning to analyze the performance of heat-tolerant cultivar YM158 and heat-sensitive cultivar YM15 under varying heat stress. While high temperatures (35 °C) severely inhibited root growth and caused oxidative damage in YM15, YM158 maintained robust root architecture and redox balance. Using weighted gene co-expression network analysis (WGCNA) alongside the random forest feature selection algorithm, we identified the flavonoid biosynthesis pathway as central to thermotolerance. Protein-protein interaction network analysis revealed that wheat root adaptability to high temperatures involves maintaining protein homeostasis via the endoplasmic reticulum protein processing system, specifically activating the flavonoid biosynthesis pathway and enhancing the antioxidant enzyme system. Furthermore, we identified a potential regulatory hub involving the cell wall sensor FERONIA (FER) and heat shock factors (HSFs), highlighting a complex interaction between hormonal signaling and secondary metabolism. Our study offers a detailed map of root heat adaptation and positions the flavonoid-mediated antioxidant system as a promising target for breeding climate-resilient crops.