Functional characterization of citrus SOS genes reveals CsSOS2 as a key regulator of salinity tolerance in Carrizo...

Salt Overly Sensitive 2 (SOS2) modulates citrus salinity tolerance by coordinating ion homeostasis and antioxidant defense, thereby enhancing growth and stress resilience in Carrizo citrange under salt stress. Citrus productivity is increasingly threatened by environmental stressors, including salinity, which reduces fruit yield and quality. Salt stress activates molecular networks such as the Salt Overly Sensitive (SOS) pathway, which maintains ion balance by promoting Na⁺ efflux and regulating stress-responsive genes. Tolerance depends on restoring ionic homeostasis, reducing osmotic stress, and limiting ROS accumulation. In this study, we identified three SOS genes in citrus SOS1, SOS2, and SOS3, and analyzed their expression under salt stress in 'Hamlin' sweet orange grafted onto Carrizo citrange. Subcellular localization of EGFP fusion proteins in Nicotiana benthamiana showed plasma membrane localization for CsSOS1, cytoplasmic and nuclear localization for CsSOS2, and a cytosolic/nuclear distribution for CsSOS3, aligning with their roles in ion transport and stress signaling. Under NaCl treatment, CsSOS2 transcript levels were downregulated in 'Hamlin' trees. To further investigate its functional role in detail, Carrizo citrange lines overexpressing CsSOS2 were developed using Agrobacterium-mediated transformation. Under salt stress, transgenic lines exhibited improved growth, higher chlorophyll content, increased antioxidant capacity, and reduced lipid peroxidation compared to wild-type plants. Ion analysis revealed moderate Na⁺ accumulation, while expression profiling showed stress-specific upregulation of antioxidant genes (CsAPX2, CsPOD1, CsPOD2) and ion transporter genes (CsSOS1, CsSOS2, CsSOS3, CsNHX1). These results suggest that CsSOS2 overexpression enhances citrus salt tolerance by modulating antioxidant defenses and maintaining ionic homeostasis, offering a promising target for developing salt-tolerant citrus cultivars and advancing understanding of stress adaptation in woody perennials.