RE3DB: A multi-omics phylogenomics platform for rice E3 ubiquitin ligases identifies novel regulators of pollen germination

The ubiquitin proteasome system (UPS) shapes rice development and stress responses through E3 ubiquitin ligases, yet roles of E3 ligases in male gametophyte function remain undiscovered and plant-focused functional informatics resource is absent. We developed the Rice E3 Ubiquitin Ligase Database (RE3DB; https://re3db.khu.ac.kr/) and used it to nominate F-box ligases involved in rice (Oryza sativa) pollen germination. RE3DB combines 15 published datasets comprising 1,602 rice E3-ligase genes; RNA-seq across 25 tissues and stress conditions; protein-to-transcript ratios from 14 tissues; six high-confidence protein-protein interaction resources; and family level phylogenies. Integrated modules provide (i) classification/annotation, (ii) E3-substrate prediction from interaction evidence, co-expression and coordinated proteolytic turnover, and (iii) functional-redundancy assessment using phylogenetic heatmaps. Applying RE3DB, we found extensive reproductive enrichment: 284 F-box genes are preferentially expressed in mature anthers. Comparative transcriptomics across nine male-sterile rice mutants identified 111 F-box genes significantly downregulated in at least one mutant (log2 fold change < -1), resolving distinct co-regulation clusters. Integration with a literature-curated regulatory network positions previously uncharacterized F-box genes within modules governing pollen-tube initiation and elongation, consistent with regulation by the RUPO receptor-like kinase and the OsMADS62/63/68 transcription-factor complex. RE3DB is the first comprehensive multi-omics web platform dedicated to functional characterization of rice E3 ligases. By unifying classification, interaction and expression-based inference, and redundancy analysis, it accelerates hypothesis-driven discovery and guides targeted mutagenesis in molecular plant science. The prioritized F-box cohort provides testable candidates for UPS-mediated control of pollen germination and tube growth, and the framework is readily extensible to other crops, holding significant potential for agricultural biotechnology.