Genome-wide identification of the cucumber 2-OXOGLUTARATE-DEPENDENT DIOXYGENASE (Cs2OGD) superfamily and functional analysis of CsF3H2 in salt stress response

Soil salinization threatens sustainable agriculture. Cucumber (Cucumis sativus L.), a globally important vegetable, is highly salt-sensitive. The 2-OXOGLUTARATE-DEPENDENT DIOXYGENASE (2-OGD) superfamily is the second largest superfamily in plants, playing a crucial role in growth, development and stress tolerance. However, its functions in cucumber remain largely unknown. Here, we identified 89 Cs2OGD genes in cucumber genome, classifying them into 3 subfamilies across 28 clades. Collinearity analysis revealed that segmental duplication is the primary driver of family expansion, with the highest homology and conserved in gibberellin metabolic observed in Cucumis melo. Promoter analysis revealed 2,686 cis-acting elements, 53% of which are stress-responsive. Transcription factor (TF) binding analysis indicated that CsDofs were the main TF regulators, targeting 85% members of this family. RNA-seq analysis revealed that five distinct expression patterns (Group I–V) for Cs2OGD genes across cucumber tissues. Further investigation of constitutively highly expressed genes (Group I and II) under abiotic stress (heat, cold, salt, drought) and hormone treatments (SA, ABA, GA, MeJA) delineated their response profiles and identified CsF3H2 as a key member regulating salt tolerance. Subcellular localization confirmed CsF3H2 resides in the nucleus and endoplasmic reticulum. Overexpression of CsF3H2 significantly enhanced flavonoid/anthocyanin content, boosted antioxidant enzyme (SOD, CAT, APX) activity, and upregulated expression of antioxidant enzymes (CsCSD, CsFSD, CsMSD, CsCAT) and ion homeostasis genes (CsSOS1, CsNHX, CsHKT), thereby improving salt tolerance. Taken together, this work provides a comprehensive evolutionary and functional analysis of the Cs2OGD superfamily and identifies a valuable genetic resource for improving crop resilience.