Identification and Analysis of the Plasma Membrane H+-ATPase Gene Family in Cotton and Its Roles in Response to Salt Stress

Abstract

Plant plasma membrane (PM) H⁺-ATPase functions as a proton-motive force by exporting cellular protons to establish a transmembrane chemical gradient of H⁺ ions and an accompanying electrical gradient. These gradients are crucial for plant growth and development and for plant responses to abiotic and biotic stresses. In this study, a comprehensive identification of the PM H⁺-ATPase gene family was conducted across four cotton species. Specifically, 14 genes were identified in the diploid species Gossypium arboreum and Gossypium raimondii, whereas 39 and 43 genes were identified in the tetraploid species Gossypium hirsutum and Gossypium barbadense, respectively. The characteristics of this gene family were subsequently compared and analyzed using bioinformatics. Chromosomal localization and collinearity analyses elucidated the distribution characteristics of this gene family within the cotton genomes. Gene structure and phylogenetic analyses demonstrated the conservation of this family across cotton species, whereas the examination of cis-acting elements in gene promoters highlighted their involvement in environmental stress and hormone response categories. An expression profile analysis revealed eight genes whose expression was upregulated under salt stress conditions, and quantitative real-time PCR results suggested that the cotton PM H⁺-ATPase genes may play crucial roles in conferring resistance to salt stress. These findings establish a robust foundation for subsequent investigations into the functions of cotton PM H⁺-ATPase genes and may offer valuable insights for selecting genes for resistance breeding programs.

Keywords: PM H+-ATPase gene family; bioinformatics; cotton species; qRT-PCR; salt stress.

Figure 1

Chromosome distribution of the cotton PM H⁺-ATPase gene family. The cotton AHA genes were distributed in the genomes of G. barbadense, G. hirsutum, G. arboreum, and G. raimondii. The AHA gene name is highlighted with a red font, and the heatmap within the genome bar shows the gene density of the chromosome. The scale on the left of the genome represents the length of the chromosome.

Figure 2

Collinearity analysis of cotton AHA genes across multiple genomes. (A) Collinearity analysis of the AHA duplicated gene pairs in G. hirsutum, G. barbadense, G. arboretum, and G. raimondii. The syntenic AHA gene pairs within the cotton genomes are shown with different colored lines, and those between the cotton genomes are shown as grey lines. (B) Collinearity analysis of the AHA gene pairs of cotton with Arabidopsis.

Figure 3

The structure of the cotton AHA genes. Clustering was conducted via MEGA, and the genes identified from different cultivars are shown in different colors. The roman numerals represent the type that genes belong to. The conserved motifs on the left were obtained via MEME, which corresponds to Figure S1, and the bar below refers to the amino acid length of the AHA protein. The exon–intron patterns on the right are summarized from the genome annotation file, and the bar below shows the length of the nucleic acid fragment where the gene is located.

Figure 4

The phylogenetic tree of the cotton AHA gene family. The color under the genes and the roman numerals represent the type to which they belong, and the round dots before the gene name represent the plant species. The phylogenetic tree was constructed via MEGA with the neighbor–joining method. The substitution model was the Tamura 3-parameter model, and the bootstrap value was 2000.

Figure 5

The cis-acting element analysis of the cotton AHA gene promoter.

Figure 6

The expression patterns of cotton AHA genes in multiple tissues (A) and under NaCl stress (B). The genes that respond to stress are highlighted in red in the heatmap.

Figure 7

Relative expression levels of GhAHA genes in cotton leaves subjected to different durations of NaCl stress. “*” indicates two-tailed significance of p ≤ 0.05, “**” indicates two-tailed significance of p ≤ 0.01.