Identification of the Glyceraldehyde-3-Phosphate Dehydrogenase (GeGAPDH) Gene Family in Gastrodia elata Revealing Its Response Characteristics to Low-Temperature and Pathogen Stress

The glyceraldehyde-3-phosphate dehydrogenase (<i>GAPDH</i>) gene plays a pivotal role in the glycolysis/gluconeogenesis process, contributing significantly to glycosyl donor synthesis, plant growth and development, and stress responses. <i>Gastrodia elata</i> Bl., a heterotrophic plant in the Orchidaceae family, has its dried tubers used as the traditional Chinese medicine. This study identified three <i>GeGAPDH</i> genes in <i>G. elata</i>, all encoding basic, stable, hydrophilic proteins. Phylogenetic analysis and subcellular localization predictions categorized <i>GeGAPDH1</i> as a plastid subtype, while <i>GeGAPDH2</i> and <i>GeGAPDH3</i> were classified as cytoplasmic subtypes. Prokaryotic expression experiments demonstrated successful expression of the GeGAPDH1 protein in <i>Escherichia coli</i>, which exhibited significant GAPDH enzymatic activity. Subcellular localization experiments showed that GeGAPDH1 was localized in the plastid. Expression analysis indicated that the three <i>GeGAPDH</i> genes were predominantly expressed in tubers. Under low-temperature stress, although the total GAPDH enzyme activity in tubers did not change significantly, the expression of <i>GeGAPDH1</i> was significantly up-regulated, while <i>GeGAPDH2</i> and <i>GeGAPDH3</i> were significantly down-regulated. This suggests that different subtypes of <i>GeGAPDH</i> may regulate cold resistance through different pathways. Upon pathogen infection, the <i>GeGAPDH</i> gene family exhibited pathogen-specific regulatory patterns. During infection by <i>Fusarium oxysporum</i>, both the expression levels of all three <i>GeGAPDH</i> genes and the total GAPDH enzyme activity in tubers increased significantly; however, <i>F. solani</i> infection induced a significant increase in total GAPDH enzyme activity without significant changes in gene expression. These results suggest that the <i>GeGAPDH</i> gene family may respond to different pathogen infections through transcriptional or translational regulation mechanisms. This study systematically identified and characterized the <i>GeGAPDH</i> gene family in <i>G. elata</i>, providing a theoretical foundation for understanding the functional differentiation of <i>GAPDH</i> in heterotrophic plants.