Evolution and Stress Responses of Gossypium hirsutum SWEET Genes

Abstract

The SWEET (sugars will eventually be exported transporters) proteins are sugar efflux transporters containing the MtN3_saliva domain, which affects plant development as well as responses to biotic and abiotic stresses. These proteins have not been functionally characterized in the tetraploid cotton, Gossypium hirsutum, which is a widely cultivated cotton species. In this study, we comprehensively analyzed the cotton SWEET gene family. A total of 55 putative G. hirsutumSWEET genes were identified. The GhSWEET genes were classified into four clades based on a phylogenetic analysis and on the examination of gene structural features. Moreover, chromosomal localization and an analysis of homologous genes in Gossypium arboreum, Gossypium raimondii, and G. hirsutum suggested that a whole-genome duplication, several tandem duplications, and a polyploidy event contributed to the expansion of the cotton SWEET gene family, especially in Clade III and IV. Analyses of cis-acting regulatory elements in the promoter regions, expression profiles, and artificial selection revealed that the GhSWEET genes were likely involved in cotton developmental processes and responses to diverse stresses. These findings may clarify the evolution of G. hirsutum SWEET gene family and may provide a foundation for future functional studies of SWEET proteins regarding cotton development and responses to abiotic stresses.

Keywords: SWEET; artificial selection; cotton; evolution; stress response.

Figure 1

Neighbor-joining phylogenetic tree of SWEET genes from three cotton species, A. thaliana, and rice. The SWEET genes were classified into four clades, which are indicated by different colors.

Figure 2

Phylogenetic relationships and structural features of GhSWEET genes. (A) Neighbor-joining phylogenetic tree of GhSWEET genes. The SWEET genes were classified into four clades, and blue, pink, purple, and orange represent Clades I, II, III, and IV, respectively. (B) Exon-intron organization of GhSWEET genes. Green and black correspond to exons and introns, respectively.

Figure 3

Chromosomal distribution of GhSWEET genes. Blue, orange and black indicate the At subgenome chromosomes, Dt subgeome chromosomes, and scaffolds, respectively. Green lines indicate tandem duplicated genes. The scale is provided in megabases.

Figure 4

Distribution of paralogous SWEET gene pairs between G. hirsutum At and Dt subgenomes. The 12 At subgenome chromosomes, 11 Dt subgenome chromosomes, and six scaffolds are indicated by different colors, with their names on the periphery. Green lines link the homologous genes located on homoeologous chromosomes in the At and Dt subgenomes. Purple lines link the homologous genes located on non-homoeologous chromosomes in the At and Dt subgenomes and scaffolds.

Figure 5

Expression profiles of GhSWEET genes in different tissues. GhSWEET gene expression profiles are divided into four groups. The FPKM values were calculated by RNA-seq data, and shown as a heat map. The colored bar represents the scale of the relative expression levels.

Figure 6

Expression patterns of GhSWEET genes in response to various stresses. The ratios of FPKM between treatments (at 1, 3, 6 and 12 h) and controls (at 0 h) were calculated by RNA-seq data, and shown as a heat map. The colored bar represents the scale of the relative expression levels.

Figure 7

Relative expression levels of the selected GhSWEET genes in response to heat and drought stresses. The GhHis3 was used as an internal control. Relative gene expression levels were calculated based on the 2^−ΔΔCT method. The mean expression values were calculated on the basis of three independent replicates. Error bars indicate the standard deviations of three independent experiments. The relative expression levels of selected GhSWEET genes after treats were compared with the controls at the same time point. Significant differences were determined by t-test (* p < 0.05; ** p < 0.01).

Figure 8

Genetic variations in G. hirsutum SWEET genes. (A) SNP density in 39 GhSWEET genes. (B) Fst values of the SNP loci in 39 GhSWEET genes. The SNPs with a Fst value greater than 0.45 were considered to be loci under selection during domestication.