StMAPKK1 Enhances Thermotolerance in Potato (Solanum tuberosum L.) by Enhancing Antioxidant Defense and Photosynthetic Efficiency Under Heat Stress

The functional role of MAPKK genes in potato (<i>Solanum tuberosum</i> L.) under high-temperature stress remains unexplored, despite their critical importance in stress signaling and yield protection. We characterized StMAPKK1, a novel group D MAPKK localized to plasma membrane/cytoplasm. Quantitative real-time polymerase chain reaction (qRT-PCR) revealed cultivar-specific upregulation in potato ('<i>Atlantic</i>' and '<i>Desiree</i>') leaves under heat stress (25 °C, 30 °C, and 35 °C). Transgenic lines overexpressing (OE) <i>StMAPKK1</i> exhibited elevated antioxidant enzyme activity, including ascorbate peroxidase (APX), catalase (CAT), superoxide dismutase (SOD), and peroxidase (POD), mitigating oxidative damage. Increased proline and chlorophyll accumulation and reduced oxidative stress markers, hydrogen peroxide (H₂O₂) and malondialdehyde (MDA), indicate improved cellular redox homeostasis. The upregulation of key antioxidant and heat stress-responsive genes (<i>StAPX</i>, <i>StCAT1/2</i>, <i>StPOD12/47</i>, <i>StFeSOD2/3</i>, <i>StMnSOD</i>, <i>StCuZnSOD1/2</i>, <i>StHSFA3</i> and <i>StHSP20</i>/<i>70</i>/<i>90</i>) strengthened the enzymatic defense system, enhanced thermotolerance, and improved photosynthetic efficiency, with significant improvements in net photosynthetic rate (Pn), transpiration rate (E), and stomatal conductance (Gs) under heat stress (35 °C) in <i>StMAPKK1</i>-OE plants. Superior growth and biomass (plant height, plant and its root fresh and dry weights, and tuber yield) accumulation, confirming the positive role of <i>StMAPKK1</i> in thermotolerance. Conversely, RNA interference (RNAi)-mediated suppression of <i>StMAPKK1</i> led to a reduction in enzymatic activity, proline content, and chlorophyll levels, exacerbating oxidative stress. Downregulation of antioxidant-related genes impaired ROS scavenging capacity and declines in photosynthetic efficiency, growth, and biomass, accompanied by elevated H₂O₂ and MDA accumulation, highlighting the essential role of <i>StMAPKK1</i> in heat stress adaptation. These findings highlight <i>StMAPKK1</i>'s potential as a key genetic target for breeding heat-tolerant potato varieties, offering a foundation for improving crop resilience in warming climates.