Genome-Wide Identification, Characterization and Expression Analysis of the Chalcone Synthase Family in Maize

Abstract

Members of the chalcone synthase (CHS) family participate in the synthesis of a series of secondary metabolites in plants, fungi and bacteria. The metabolites play important roles in protecting land plants against various environmental stresses during the evolutionary process. Our research was conducted on comprehensive investigation of CHS genes in maize (Zea mays L.), including their phylogenetic relationships, gene structures, chromosomal locations and expression analysis. Fourteen CHS genes (ZmCHS01-14) were identified in the genome of maize, representing one of the largest numbers of CHS family members identified in one organism to date. The gene family was classified into four major classes (classes I-IV) based on their phylogenetic relationships. Most of them contained two exons and one intron. The 14 genes were unevenly located on six chromosomes. Two segmental duplication events were identified, which might contribute to the expansion of the maize CHS gene family to some extent. In addition, quantitative real-time PCR and microarray data analyses suggested that ZmCHS genes exhibited various expression patterns, indicating functional diversification of the ZmCHS genes. Our results will contribute to future studies of the complexity of the CHS gene family in maize and provide valuable information for the systematic analysis of the functions of the CHS gene family.

Keywords: chalcone synthase; evolution; expression; genome-wide analysis; maize.

Figure 1

Phylogenetic tree and gene structure of the 14 predicted maize chalcone synthases (CHSs). (A) The unrooted neighbor-joining tree was constructed using the MEGA4.0 program. The bootstrap values, which were produced using 1000 replicates with the pairwise deletion option, are noted at each node; (B) Exon-intron structure was generated using Gene Structure Display Server (GSDS). The exons and introns are indicated by green boxes and gray lines, respectively. The scale at the bottom is in kilobases.

Figure 2

Distribution of 20 conserved motifs in the putative CHS proteins of maize. Motifs in the putative ZmCHS proteins were obtained using the Multiple Em for Motif Elicitation (MEME) web server. The length of each box does not represent the actual size of each motif.

Figure 3

Sequence alignment of ZmCHSs against the other plant CHSs. The first line represents the secondary structure of Medicago sativa chalcone synthase (MsCHS). The blue boxes and red letters represent conserved residues. The red regions represent sequences of strict sequence conservation. The black wave lines and black arrows represent α-helix and β-pleated sheet. The catalytic triad of CHSs, residues connected with CoA-binding and CHS family-specific Pro375 are labeled with blue points, green pentagrams and black quadrangles at the bottom of the sequences, respectively. MsCHS = Medicago sativa (alfalfa) chalcone synthase (P30074); AtCHS = Arabidopsis chalcone synthase (AAB35812.1); PpCHS = Physcomitrella patens chalcone synthase (ABB84527.1).

Figure 4

Location of 14 CHS genes on maize chromosomes. The “Chr.” at the top of each bar represent the chromosome number of maize. The location of each CHS gene is indicated by corresponding gene name at the left of corresponding chromosome. The segmental duplicated genes are connected by dashed lines. The scale on the left is in megabases.

Figure 5

Sliding window plots of duplicated CHS genes. The gray blocks, from dark to light, indicate the positions of the Chal_sti_synt_N domain and Chal_sti_synt_C domain, respectively. The window size was 150 bp and the step size was 9 bp.

Figure 6

Clustering of expression profiles of 14 ZmCHS genes. Different organs/tissues are exhibited below each column. 24H, 24 h after imbibition; DAS, days after sowing; GH, growth hormone; SAM, Shoot apical meristem; V3, Vegetative 3, three fully extended leaves; V5, Vegetative 5, five fully extended leaves; VT, Vegetative tasseling, last branch of the tassel fully emerged; R2, Reproductive 2, 10–14 days after silk emergence; DAP, days after pollination. The names of ZmCHS genes are displayed at the right side of each row. The colour box from blue to orange indicate an increased expression level.

Figure 7

Expression patterns of 10 ZmCHS genes after salicylic acid treatment. Relative expression levels of ZmCHS genes in response to salicylic acid were examined by qPCR and normalized by the expression of Maize Actin 1 (NM_001136991.1). The y-axis represents the relative expression level and the x-axis represents the time course of stress treatment. Seedlings were sampled at 0, 1, 3, 6, 12, 24, 36 and 48 h after salicylic acid treatment. There were three technical replicates for each of the three biological replicates.