Genome-wide analysis of WRKY transcription factors in white pear (Pyrus bretschneideri) reveals evolution and patterns under drought stress

Abstract

Background: WRKY transcription factors (TFs) constitute one of the largest protein families in higher plants, and its members contain one or two conserved WRKY domains, about 60 amino acid residues with the WRKYGQK sequence followed by a C2H2 or C2HC zinc finger motif. WRKY proteins play significant roles in plant development, and in responses to biotic and abiotic stresses. Pear (Pyrus bretschneideri) is one of the most important fruit crops in the world and is frequently threatened by abiotic stress, such as drought, affecting growth, development and productivity. Although the pear genome sequence has been released, little is known about the WRKY TFs in pear, especially in respond to drought stress at the genome-wide level.

Results: We identified a total of 103 WRKY TFs in the pear genome. Based on the structural features of WRKY proteins and topology of the phylogenetic tree, the pear WRKY (PbWRKY) family was classified into seven groups (Groups 1, 2a-e, and 3). The microsyteny analysis indicated that 33 (32%) PbWRKY genes were tandemly duplicated and 57 genes (55.3%) were segmentally duplicated. RNA-seq experiment data and quantitative real-time reverse transcription PCR revealed that PbWRKY genes in different groups were induced by drought stress, and Group 2a and 3 were mainly involved in the biological pathways in response to drought stress. Furthermore, adaptive evolution analysis detected a significant positive selection for Pbr001425 in Group 3, and its expression pattern differed from that of other members in this group. The present study provides a solid foundation for further functional dissection and molecular evolution of WRKY TFs in pear, especially for improving the water-deficient resistance of pear through manipulation of the PbWRKYs.

Fig. 1

Localization and duplication of the WRKY genes in the pear genome. Circular visualization of the 103 WRKY genes was mapped on the different chromosomes in the pear genome using Circos software. Chromosome number is indicated on the chromosome. The microsynteny between each pair of WRKY genes were detected by using the MicroSyn software. The genes with synteny relationship are linked by lines. Red link: ≥30 anchors in a synteny block, blue link: 20–30 anchors, green link: 10–20 anchors, gray link: 5–10 anchors

Fig. 2

Phylogenetic trees of WRKY genes in pear. The un-rooted phylogenetic tree of WRKY domains was constructed with MEGA5.1 program with the NJ method. The numbers beside the branches represent bootstrap values based on 1000 replications. The name of groups (1, 2a-e, and 3) are shown at the inside of the circle. The groups of genes are shown in different colors

Fig. 3

Schematic representations of the conserved motifs and exon–intron compositions. Names of genes are indicated on the left. a Conserved motifs in WRKY proteins. Different motifs are highlighted with different colored boxes with numbers 1 to 20. Lines represent protein regions without detected motif. b Exon–intron compositions in WRKY gene. Exons, represented by gray or red boxes, are drawn to scale. Dashed lines connecting two exons represent an intron. WRKY domain is marked in red

Fig. 4

Heat map of RNA-seq expression of PbWRKY gene in response to drought stress. Color scale of the dendrogram represents log2 ratio value of treated sample to control sample. D1, D3, D6 and D24, dehydrated for 1, 3 and 6 h in an ambient environment and recovered for 24 h in water, respectively. The colors of genes in Cluster 1–4 represent their groups in the phylogenetic tree in Fig. 2

Fig. 5

Quantitative RT-PCR analysis of PbWRKY gene expression in response to drought stress. D0, D1, D3, D6 and D24, dehydrated for 0, 1, 3 and 6 h in an ambient environment and recovered for 24 h in water, respectively