Genome-Wide Identification of LRR-RLK Family in Saccharum and Expression Analysis in Response to Biotic and Abiotic Stress

Abstract

The leucine-rich repeat receptor-like protein kinase (LRR-RLK) gene family is the largest family of the receptor-like protein kinases (RLKs) superfamily in higher plants, which is involved in regulating the plant growth and development, stress responses, signal transduction and so on. However, no comprehensive analyses of LRR-RLKs have been reported in sugarcane. Here, we performed a comprehensive analysis of the LRR-RLK gene family in sugarcane ancestor species Saccharum spontaneum. A total of 437 LRR-RLK genes were identified and categorized into 14 groups based on a maximum likelihood phylogenetic tree. The chromosome location showed an uneven distribution on all 32 chromosomes in sugarcane. Subsequently, the exon-intron organization structure and conserved motif arrangement were relatively conserved among the same groups or subgroups and between Arabidopsis and S. spontaneum genomes. Furthermore, the promoter sequences analyses showed that sugarcane LRR-RLK genes (SsLRR-RLKs) were strongly regulated by various environmental stimuli, phytohormonal factors and transcription factors (TFs). Eventually, the expression profiles of SsLRR-RLK genes at different stresses were analyzed based on RNA-seq data, suggesting their potential roles in the regulation of sugarcane responses to diverse abiotic and biotic stress. Overall, the findings provide insight into the potential functional roles and lay the foundation for further functional study.

Keywords: LRR-RLK gene family; Saccharum; bioinformatics; gene expression analysis; molecular evolution; stress response.

Figure 1

Chromosome distribution of LRR-RLK family in S. spontaneum. 437 SsLRR-RLK alleles were located in all 32 chromosomes. The figure was drawn using MG2C v2.0.

Figure 2

Phylogenetic tree of the LRR-RLK genes between Arabidopsis and S. spontaneum. The phylogenetic tree was generated using FastTree 2.0 using the maximum likelihood method. All sugarcane LRR-RLKs were classified into 14 distinct groups and different groups were shown by different color.

Figure 3

Representative exon–intron (A) and conserved motifs structure (B) of each S. spontaneum LRR-RLK subfamily. (A) Exon–intron structures of SsLRR-RLKs. Untranslated regions (UTRs), exons and introns are represented by blue boxes, red boxes and black lines, respectively. (B) The motif distribution of SsLRR-RLK proteins. The 15 predicted motifs are represented by distinct color boxes using the MEME program.

Figure 4

Heatmaps of the expression analysis of SsLRR-RLKs in response to diverse abiotic stress. The differential expression profiles in two different sugarcane varieties is shown by heatmap under (A,B) low nitrogen (leaf) stress, (C,D) low nitrogen (root) stress and (E,F) cold stress. Heatmap was generated based on FPKM values. The color scale represents expression values of each sample, with red showing high levels and green showing low levels of transcript abundance.

Figure 5

Heatmaps of the expression analysis of SsLRR-RLKs under different biotic stresses. The differential expression patterns in two different sugarcane varieties were shown by heatmap under (A,B) infection by sugarcane leaf blight disease and (C,D) infection by sugarcane smut. Heatmap was generated based on FPKM values. The color scale represents expression values of each sample, with red showing high levels and green showing low levels of transcript abundance.

Figure 6

Relative expression levels of nine selected LRR-RLK genes were analyzed using quantitative real-time PCR (qRT-PCR). Among them, three obvious periods were discovered, namely the early, medium and late periods. (A) contains three highly expressed genes that respond to early period (SsLRR-RLK88/146/346) that were identified. (B) contains three highly expressed genes that respond to medium period (SsLRR-RLK19/24/391) that were identified. (C) contains three highly expressed genes that respond to late period (SsLRR-RLK203/249/359) that were identified. X-axes show different stages and y-axes show relative expression level of each LRR-RLK gene. Bars represent the standard deviation (SD) (n = 3) of three technical replicates. Different lowercase letters above the bars in the figure indicate a significant difference (p < 0.05). Lowercase letters reflect the 5% significance level. As shown in the chart, a, b, c, d, e and f are different from each other, and they are all significantly different from each other. In addition, the same lowercase letter means no significantly different.