Cold Shock Protein (EpCSP) from Exiguobacterium profundum Confers Enhanced Stress Tolerance to Drought and Cold in a Heterologous System

Abstract Extremophile-derived genes represent vital genetic resources for enhancing abiotic stress tolerance in crops. In this study, the cold shock protein gene EpCSP from the deep-sea bacterium Exiguobacterium profundum was heterologously expressed in tobacco (Nicotiana tabacum). Molecular identification confirmed the stable integration and transcription of EpCSP in transgenic lines. Bioinformatic analysis indicated that EpCSP is a hydrophilic protein lacking transmembrane domains, strongly suggesting its cytosolic localization. Phenotypic evaluations demonstrated that EpCSP-overexpressing (OE) plants exhibited significantly enhanced resilience to both drought and cold stresses, characterized by markedly reduced wilting, delayed chlorosis, and superior biomass preservation compared to the wild type (WT). Physiological analyses showed that OE plants maintained higher relative water content (RWC) and chlorophyll levels, accumulated more proline, and exhibited elevated superoxide dismutase (SOD) activity, which collectively mitigated thiobarbituric acid reactive substances (TBARS) accumulation. Correlation heatmap analysis further identified a physiological “damage decoupling” effect, where EpCSP attenuated the strict linkage between water deficit and membrane lipid peroxidation. These results suggest that EpCSP functions as a putative RNA chaperone that coordinates a multi-tiered defense network involving osmotic adjustment, antioxidant defense, and photosynthetic preservation, thereby conferring dual tolerance to drought and cold stresses. This study highlights EpCSP as a promising candidate gene for engineering climate-resilient crops.