Identification of circadian-regulated genes for enhancing acidic soil tolerance in rice through integrative transcriptomic and systems analysis

The plant circadian clock orchestrates key physiological processes and stress responses by regulating time-dependent gene expression. However, its role in mediating responses to soil acidity, one of the major abiotic constraints affecting rice productivity, remains poorly understood. In this study, we performed a comprehensive transcriptomic and systems analysis using publicly available RNA-Seq data to investigate the role of circadian-regulated genes in adapting to acidic stress in Oryza sativa. A total of 2,285 differentially expressed genes (DEGs) were identified under various pH conditions (4.5 and 6.5), of which 37 were associated with the circadian clock. Weighted Gene Co-expression Network Analysis (WGCNA) classified the DEGs into five modules, three of which were strongly correlated with acidic stress. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses revealed key roles in redox homeostasis, hormone signaling, and photosynthetic regulation. Hub genes within stress-associated modules were further analyzed for protein–protein interactions, hydrophobicity profiles, and 3D structures using AlphaFold. Notably, several circadian-related genes were downregulated under acidic stress, suggesting clock suppression as an adaptive mechanism. These findings reveal the connections between circadian regulation and pH stress tolerance, providing promising genetic targets for developing acidity-resilient rice cultivars.