Ubiquitin-conjugating enzyme gene SgUBC2 confers manganese tolerance in Stylosanthes guianensis through antioxidant defense augmentation and manganese-responsive gene regulation.

Stylo (Stylosanthes guianensis) is an important tropical legume with superior tolerance to manganese (Mn) toxicity, which severely constrains plant growth, especially in tropical acid soils. However, the mechanisms underlying Mn tolerance in stylo remain largely unknown. In this study, excess Mn-caused growth inhibition was observed in stylo shoots but not roots, accompanied by increased activities of superoxide dismutase and catalase, as well as elevated concentrations of ascorbate and glutathione, which may contribute to oxidative stress scavenging under Mn toxicity. SgUBC2, encoding a ubiquitin-conjugating enzyme, was functionally characterized. The results showed that the expression of SgUBC2 was significantly induced by excess Mn in stylo roots. Subcellular localization analysis revealed that SgUBC2 localized to both the cytosol and nucleus in Arabidopsis mesophyll protoplasts. Moreover, compared to the control plants, overexpression of SgUBC2 increased the shoot and root dry weights of the transgenic composite stylo plants under excess Mn conditions, likely due to reduced Mn accumulation and increased superoxide dismutase activity in roots. RNA-seq analysis showed that SgUBC2 overexpression altered the expression of multiple Mn-responsive genes, including those associated with antioxidant defense response, ion transporters and organic acid metabolism. Collectively, this study indicates that SgUBC2 plays critical roles in enhancing Mn tolerance in stylo, potentially through an elevated antioxidant defense system and coordinated regulation of Mn-responsive gene expression.