Background: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is the central enzyme of glycolysis and plays important regulatory roles in plant growth and development and responses to adverse stress conditions. However, studies on the characteristics and functions of cotton GAPDH family genes are still lacking.
Methods: In this study, genome-wide identification of the cotton GAPDH gene family was performed, and the phylogeny, gene structures, promoter progenitors and expression profiles of upland cotton GAPDH gene family members were explored by bioinformatics analysis to highlight potential functions. The functions of GhGAPDH9 in response to drought stress were initially validated based on RNA-seq, qRT‒PCR, VIGS techniques and overexpression laying a foundation for further studies on the functions of GAPDH genes.
Results: This study is the first systematic analysis of the cotton GAPDH gene family, which contains a total of 84 GAPDH genes, among which upland cotton contains 27 members. Quantitative, phylogenetic and covariance analyses of the genes revealed that the GAPDH gene family has been conserved during the evolution of cotton. Promoter analysis revealed that most cis-acting elements were related to MeJA and ABA. Based on the identified promoter cis-acting elements and RNA-seq data, it was hypothesized that Gh_GAPDH9, Gh_GAPDH11, Gh_GAPDH19 and Gh_GAPDH21 are involved in the response of cotton to abiotic stress. The expression levels of the Gh_GAPDH9 gene in two drought-resistant and two drought-sensitive materials were analyzed by qRT‒PCR and found to be high early in the treatment period in the drought-resistant material. The silencing of Gh_GAPDH9 based on virus-induced gene silencing (VIGS) technology resulted in significant leaf wilting or whole-plant dieback in silenced plants after drought stress compared to the control. The content of-malondialdehyde (MDA) in cotton leaves was significantly increased, and the content of proline (Pro) and chlorophyll (Chl) was reduced. In addition, the leaf wilting and dryness of transgenic lines under drought stress were lower than those of wild-type Arabidopsis, indicating that Gh_GAPDH9 is a positive regulator of drought resistance. In conclusion, our results demonstrate that GAPDH genes play an important role in the response of cotton to abiotic stresses and provide preliminary validation of the function of the Gh_GAPDH9 gene under drought stress. These findings provide an important theoretical basis for further studies on the function of the Gh_GAPDH9 gene and the molecular mechanism of the drought response in cotton.
Keywords: Drought stress; G. hirsutum; Gh_GAPDH9; Glyceraldehyde-3-phosphate dehydrogenase; Phylogenetic analysis.
© 2023 Geng et al.