Characterization of Glycerol-3-phosphate Acyltransferase Gene (AnGPAT) and Its Promoter Involved in Cold Tolerance from Ammopiptanthus nanus

Abstract Chloroplast glycerol-3-phosphate acyltransferase (GPAT), the primary enzyme influencing phosphatidylglycerol unsaturation in thylakoid membranes, plays a crucial role in regulating plant cold tolerance. Ammopiptanthus nanus exhibits significant cold stress tolerance due to its unique growth environment. The characterization of the GPAT gene in A. nanus remains largely unexplored. This study isolated and analyzed the sequence structure of the AnGPAT gene and its promoter in A. nanus. Phylogenetic analysis indicated that AnGPAT is closely related to GPATs from Glycine max, Lupinus albus, and Glycine soja, all members of the Fabaceae family. The subcellular localization indicated that AnGPAT is localized in the chloroplast membrane. AnGPAT transcription levels increased in response to cold stress and ABA treatment. Overexpression of AnGPAT in Arabidopsis thaliana enhanced seed germination and transgenic seedling growth under cold stress conditions. In transgenic plants under cold stress, proline content increased, while malondialdehyde content and relative electrolyte leakage decreased. The content of GPAT enzyme and unsaturated fatty acids in transgenic plants were significantly higher than in WT plants under cold stress. In addition, the promoter of AnGPAT includes MYB/MYC elements responsive to cold and ABRE elements responsive to ABA, which trigger GUS protein expression under cold stress and ABA treatment. The findings demonstrated that AnGPAT improved cold tolerance in transgenic plants. This study provides evidence for further understanding the regulation mechanism related to ABA and cold stress in A. nanus.