Genome-Wide Identification and Expression Profiling of Glutathione S-Transferase Gene Family in Foxtail Millet (Setaria italica L.)

Abstract

Glutathione S-transferases (GSTs) are a critical superfamily of multifunctional enzymes in plants. As a ligand or binding protein, GSTs regulate plant growth and development and detoxification. Foxtail millet (Setaria italica (L.) P. Beauv) could respond to abiotic stresses through a highly complex multi-gene regulatory network in which the GST family is also involved. However, GST genes have been scarcely studied in foxtail millet. Genome-wide identification and expression characteristics analysis of the foxtail millet GST gene family were conducted by biological information technology. The results showed that 73 GST genes (SiGSTs) were identified in the foxtail millet genome and were divided into seven classes. The chromosome localization results showed uneven distribution of GSTs on the seven chromosomes. There were 30 tandem duplication gene pairs belonging to 11 clusters. Only one pair of SiGSTU1 and SiGSTU23 were identified as fragment duplication genes. A total of ten conserved motifs were identified in the GST family of foxtail millet. The gene structure of SiGSTs is relatively conservative, but the number and length of exons of each gene are still different. The cis-acting elements in the promoter region of 73 SiGST genes showed that 94.5% of SiGST genes possessed defense and stress-responsive elements. The expression profiles of 37 SiGST genes covering 21 tissues suggested that most SiGST genes were expressed in multiple organs and were highly expressed in roots and leaves. By qPCR analysis, we found that 21 SiGST genes were responsive to abiotic stresses and abscisic acid (ABA). Taken together, this study provides a theoretical basis for identifying foxtail millet GST family information and improving their responses to different stresses.

Keywords: expression analysis; foxtail millet; glutathione S-transferase (GSTs); stress response.

Figure 1

Phylogenetic tree of GST proteins among foxtail millet and A. thaliana. The GST protein sequences of 73 foxtail millet and 55 A. thaliana are divided into eight classes. Different subfamilies are marked as different colors.

Figure 2

Chromosome mapping of GST genes in foxtail millet. A total of 30 tandem duplication gene pairs belonging to 11 clusters were present (highlighted by full red lines). Only one pair of gene segmental duplication event was found (highlighted by dashed black line).

Figure 3

Gene structure and conserved motif of foxtail millet GSTs. (a) Neighbor-joining (NJ) phylogenetic tree based on SiGST protein sequences. (b) Conserved motif of SiGSTs. (c) Gene structure of SiGSTs.

Figure 4

Predicted cis-acting element in SiGST promoter of foxtail millet.

Figure 5

Relative expression patterns (FPKM value) of GST genes involved in 21 tissues of foxtail millet. The FPKM values of SiGST genes in different tissues were obtained from the Setrari italica Multiomics database (http://foxtail-millet.biocloud.net/home, accessed on 8 June 2022). The color bar represents log₂ expression levels (FPKM), with red indicating high gene expression levels and green indicating low gene expression levels.

Figure 6

Relative expression patterns of 21 SiGSTs in leaves of foxtail millet under abiotic stresses and ABA treatments. Relative expression patterns of 21 SiGST genes were analyzed with qPCR under osmotic (20% PEG 6000) (a), salt (200 mM NaCl) (b), cold stress (4 °C temperature) (c), and 100 μM ABA (d). The relative expression levels of each gene were calculated by the 2^−∆∆Ct method. The heat map of SiGSTs expression condition was constructed by TBtools, expression value was standardized by Log2. The expression level of target gene at 0 h was used as control.