The ubiquitin–proteasome system (UPS) is important for protein post-translational modification in plants. E2 (ubiquitin-conjugating enzyme) and E3 (ubiquitin ligases enzyme), key enzymes of UPS, play crucial roles in all aspects of plant development, growth, and environmental stresses. Despite extensive knowledge of UPS roles in crop growth and development, E2-E3 pair functions in potato tuber development and stress responses remain understudied. Here, we describe the role of StUBC18 (a potato E2) in drought stress tolerance. It is determined that StUBC18 (E2)-StPUB40 (E3) pair plays important roles in drought stress tolerance and potato tuber yield. StUBC18 and StPUB40 expression was downregulated under various stresses (drought, salt, polyethylene glycol, and H₂O₂). Overexpression of StUBC18 and StPUB40 in potatoes decreased drought stress tolerance, while interfering with the expression of StUBC18 and StPUB40 increased drought stress tolerance, respectively. The protein interaction test demonstrated that StUBC18 interacts with StPUB40 in the plant cell. Co-overexpression of StUBC18-StPUB40 in potato enhanced reactive oxygen species (ROS) accumulation and induced pleiotropic changes, reducing drought tolerance. Our findings revealed how the StUBC18-StPUB40 pair regulates potato drought stress tolerance by altering leaf anatomy (palisade and spongy tissue thickness) and influences tuber yield.

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