Integrative Multi-Scale Analysis Identifies the Hub Gene TRE1 Associating Trehalose Metabolism With Salinity...

Salinity is a major constraint on rice (Oryza sativa L.) production, yet the regulatory hubs that distinguish tolerant from susceptible genotypes remain largely unknown. Using a computational and experimental pipeline combining time-series meta-transcriptomics, weighted gene co-expression network analysis (WGCNA), fuzzy clustering, network centrality metrics, promoter analysis, and molecular and physicochemical evaluations, we identified genotype-specific hub genes associated with salinity tolerance in rice. Analysis of RNA-Seq data from tolerant (Pokkali) and susceptible (IR29) genotypes across five time points after salinity treatment yielded 3369 meta-differentially expressed genes. WGCNA revealed a salinity-responsive co-expression module, within which fuzzy clustering uncovered a cluster of 48 genes showing striking opposite temporal patterns between genotypes. Multi-metric centrality ranking prioritized Trehalase1 (TRE1), encoding a trehalase enzyme, among the identified hub genes. Promoter analysis of TRE1 revealed the presence of stress-related cis-elements (ABRE, TGACG-motif, MYB/MYC, DRE). Independent greenhouse validation confirmed the opposite expression pattern of TRE1 between Pokkali and IR29 under salinity, correlating with increased trehalose accumulation in Pokkali and trehalose depletion in IR29. Physicochemical assessments showed that Pokkali exhibited lower oxidative stress markers, higher relative water content, better photosynthetic performance, and a more favorable K⁺/Na⁺ ratio than IR29. Principal component analysis distinctly separated the two genotypes, with TRE1 expression clustering with susceptibility-associated traits, while trehalose clustered with tolerance-associated traits. Our network-based discovery and experimental validation together indicate that the contrasting regulation of the hub gene TRE1 links trehalose metabolism to salinity tolerance in rice, highlighting TRE1 as a possible target for further functional analysis to enhance salinity tolerance.