Genome-Wide Analysis and Characterization of the Proline-Rich Extensin-like Receptor Kinases (PERKs) Gene Family Reveals Their Role in Different Developmental Stages and Stress Conditions in Wheat (Triticum aestivum L.)

Abstract

Proline-rich extensin-like receptor kinases (PERKs) are a class of receptor kinases implicated in multiple cellular processes in plants. However, there is a lack of information on the PERK gene family in wheat. Therefore, we identified 37 PERK genes in wheat to understand their role in various developmental processes and stress conditions. Phylogenetic analysis of PERK genes from Arabidopsis thaliana, Oryza sativa, Glycine max, and T. aestivum grouped them into eight well-defined classes. Furthermore, synteny analysis revealed 275 orthologous gene pairs in B. distachyon, Ae. tauschii, T. dicoccoides, O. sativa and A. thaliana. Ka/Ks values showed that most TaPERK genes, except TaPERK1, TaPERK2, TaPERK17, and TaPERK26, underwent strong purifying selection during evolutionary processes. Several cis-acting regulatory elements, essential for plant growth and development and the response to light, phytohormones, and diverse biotic and abiotic stresses, were predicted in the promoter regions of TaPERK genes. In addition, the expression profile of the TaPERK gene family revealed differential expression of TaPERK genes in various tissues and developmental stages. Furthermore, TaPERK gene expression was induced by various biotic and abiotic stresses. The RT-qPCR analysis also revealed similar results with slight variation. Therefore, this study's outcome provides valuable information for elucidating the precise functions of TaPERK in developmental processes and diverse stress conditions in wheat.

Keywords: PERK; RT-qPCR; drought; heat stress; kinase; promoter.

Figure 1

Phylogenetic analysis of TaPERK proteins with Arabidopsis (15), rice (8), and soybean (16). The phylogenetic analysis was executed using the ClustalW program as well as MEGAX software by the neighbor-joining method and bootstrap values of 1000 replicates. The numbers on the nodes indicate the bootstrap values. Distinct groups are represented by the different colors.

Figure 2

Genomic distribution of identified PERK genes on the 21 chromosomes of wheat and within the three sub-genomes. (A) Schematic representations of the chromosomal distribution of PERK genes on the 21 chromosomes of wheat and the name of the gene on the right. The colored circles on the chromosomes indicate the position of the PERK genes. The chromosome numbers of the three sub-genomes are indicated at the top of each bar. (B) Distribution of PERK genes in the three sub-genomes. (C) Distribution of PERK genes across 21 chromosomes, Un: unaligned contig.

Figure 2

Genomic distribution of identified PERK genes on the 21 chromosomes of wheat and within the three sub-genomes. (A) Schematic representations of the chromosomal distribution of PERK genes on the 21 chromosomes of wheat and the name of the gene on the right. The colored circles on the chromosomes indicate the position of the PERK genes. The chromosome numbers of the three sub-genomes are indicated at the top of each bar. (B) Distribution of PERK genes in the three sub-genomes. (C) Distribution of PERK genes across 21 chromosomes, Un: unaligned contig.

Figure 3

Syntenic relationships of TaPERK genes between Aegilops tauschii, Brachypodium distachyon, and Oryza sativa. The gray lines in the background represent the collinear blocks within Triticum aestivum and other plant genomes, while the red lines highlight the syntenic PERK gene pairs.

Figure 4

Diagrammatic representation of the exon–intron organization of the TaPERK genes. Yellow boxes represent exons, untranslated regions (UTRs) are indicated by blue boxes, and black lines represent introns. The lengths of the boxes and lines are scaled based on gene length. The exon and intron sizes can be estimated using the scale at the bottom.

Figure 5

Conserved motifs of TaPERK genes elucidated by MEME. Up to 10 motifs were shown in different colors. (A) Colored boxes representing different conserved motifs with different sequences and sizes. (B) Sequence logo conserved motif of the wheat PERK proteins. The overall height of each stack represents the degree of conservation at this position, while the height of individual letters within each stack indicates the relative frequency of the corresponding amino acids. The sequence of each motif, combined p-value, and length are shown on the left side of the figure. MEME Parameters: number of repetitions, any; maximum number of motifs, 10; optimum motif width, between 6 and 50.

Figure 5

Conserved motifs of TaPERK genes elucidated by MEME. Up to 10 motifs were shown in different colors. (A) Colored boxes representing different conserved motifs with different sequences and sizes. (B) Sequence logo conserved motif of the wheat PERK proteins. The overall height of each stack represents the degree of conservation at this position, while the height of individual letters within each stack indicates the relative frequency of the corresponding amino acids. The sequence of each motif, combined p-value, and length are shown on the left side of the figure. MEME Parameters: number of repetitions, any; maximum number of motifs, 10; optimum motif width, between 6 and 50.

Figure 6

Multiple sequence alignment of the TaPERK protein sequences. The conserved protein tyrosine kinase domain is boxed in red. Colored and shaded amino acids are chemically similar residues. Dashes indicate gaps introduced to maximize the alignment of the homologous region. * indicates positions which have a single, fully conserved residue.

Figure 6

Multiple sequence alignment of the TaPERK protein sequences. The conserved protein tyrosine kinase domain is boxed in red. Colored and shaded amino acids are chemically similar residues. Dashes indicate gaps introduced to maximize the alignment of the homologous region. * indicates positions which have a single, fully conserved residue.

Figure 7

Cis-acting regulatory elements (CAREs) in the promoter region of the TaPERK genes family. The CAREs analysis was performed with a 2kb upstream region using PlantCARE online server. The different numbers of cis-regulatory elements represent different colors. (A) Hormone-responsive elements, stress-responsive elements, growth and development-related elements, light-responsive elements, and other elements with unknown functions are differentiated by color. (B) Most commonly occurring CAREs in TaPERKs.

Figure 8

Heatmap representing expression profile of the TaPERK genes at various developmental stages. Columns represent genes, and rows represent different developmental stages. TPM values were used directly to create the heatmaps. The “z” nomenclature refers to Zadok’s growth stage.

Figure 9

Quantitative real-time PCR analysis of selected TaPERK genes in response to drought stress (DS), heat stress (HS), and cold stress to verify RNA seq data. The wheat actin gene was used as the internal control to standardize the RNA samples for each reaction. Asterisks indicate significant differences compared with control. Bars represent results of Tukey’s HSD test at the <0.05 and <0.001 level (* p < 0.05, ** p lies in between the values of 0.05 and 0.001, and *** p < 0.001). Error bars show standard deviation. Data are mean ± SD (n = 3).

Figure 10

Protein–protein interaction analysis of TaPERKs proteins. Protein–protein interaction network produced by STRINGV9.1, each node represents a protein, and each edge represents an interaction, colored by evidence type. The figure highlights the connections between differentially represented proteins.