Genome-wide identification and expression analysis of GRAS gene family in Eucalyptus grandis

Abstract

Background: The GRAS gene family is a class of plant-specific transcription factors with important roles in many biological processes, such as signal transduction, disease resistance and stress tolerance, plant growth and development. So far, no information available describes the functions of the GRAS genes in Eucalyptus grandis.

Results: A total of 82 GRAS genes were identified with amino acid lengths ranging from 267 to 817 aa, and most EgrGRAS genes had one exon. Members of the GRAS gene family of Eucalyptus grandis are divided into 9 subfamilies with different protein structures, while members of the same subfamily have similar gene structures and conserved motifs. Moreover, these EgrGRAS genes expanded primarily due to segmental duplication. In addition, cis-acting element analysis showed that this family of genes was involved involved in the signal transduction of various plant hormones, growth and development, and stress response. The qRT-PCR data indicated that 18 EgrGRAS genes significantly responded to hormonal and abiotic stresses. Among them, the expression of EgrGRAS13, EgrGRAS68 and EgrGRAS55 genes was significantly up-regulated during the treatment period, and it was hypothesised that members of the EgrGRAS family play an important role in stress tolerance.

Conclusions: In this study, the phylogenetic relationship, conserved domains, cis-elements and expression patterns of GRAS gene family of Eucalyptus grandis were analyzed, which filled the gap in the identification of GRAS gene family of Eucalyptus grandis and laid the foundation for analyzing the function of EgrGRAS gene in hormone and stress response.

Keywords: Abiotic stress; Eucalyptus grandis; GRAS family; Genome-wide analysis; Phytohormone.

Fig. 1

Phylogenetic tree of GRAS genes from E. grandis, Arabidopsis, rice and soybean. 83 EgrGRAS genes, 34 AtGRAS genes, 40 OsGRAS genes and 61 GmGRAS genes are clustered into 9 subfamilies. GRAS genes from E. grandis, Arabidopsis, rice and soybean are denote by red, blue, yellow and green shape, respectively. Details of the GRAS genes from four species are listed in Table S3. The tree was generated with the Clustal X 2.0 software using the neighbor-joining (N-J) method

Fig. 2

Analysis of the motif and gene structure of the GRAS gene family in E. grandis. (A) Gene structure of GRAS genes in E. grandis. Exons are indicated by green rectangles. Gray lines connecting two exons represent introns. (B) Conserved motifs of GRAS genes in E. grandis. Distribution of the 20 conserved motifs in the EgrGRAS genes following analysis by MEME tool. The different-colored boxes represent different motifs and their position in each protein sequence of GRAS. (C) Domain analysis of GRAS proteins in E. grandis

Fig. 3

Chromosomal location of GRAS genes in E. grandis. (A) The 82 GRAS genes are widely mapped to 11 chromosomes of E. grandis. The blue boxes in front of the genes on behalf of these genes belonging to a gene cluster. (B) The number distribution of GRAS gene family in 10 chromosomes

Fig. 4

Collinearity analysis. (A) Collinearity analysis of GRAS gene in E. grandis. (B) GRAS gene collinearity between E. grandis and other species (Arabidopsis, rice and soybean) genomes

Fig. 5

Cis-acting elements analysis of EgrGRAS genes in promoter region of E. grandis. Left panel: Number of each cis-acting element in the promoter region (2000 bp) of EgrGRAS genes. Right panel: Statistics for the total number of EgrGRAS genes

Fig. 6

Expression analysis of 18 EgrGRAS genes following ABA and GA₃ treatments by qRT-PCR. The Y-axis and X-axis indicates relative expression levels and the time courses of stress treatments, respectively. Statistical significance was performed using a paired Student’s t test. Mean values and standard deviations (SDs) were obtained from three biological and three technical replicates, and significant differences relative to controls were indicated at ^∗P ≤ 0.05 and ^∗∗P ≤ 0.01. The error bars indicate standard deviation

Fig. 7

Expression analysis of 18 EgrGRAS genes following cold, salt and drought treatments by qRT-PCR. The Y-axis and X-axis indicates relative expression levels and the time courses of stress treatments, respectively. Statistical significance was performed using a paired Student’s t test. Mean values and standard deviations (SDs) were obtained from three biological and three technical replicates, and significant differences relative to controls were indicated at ^*P ≤ 0.05 and ^**P ≤ 0.01. The error bars indicate standard deviation

Fig. 8

Correlations and co-regulatory networks of 18 EgrGRAS genes under stress treatments. (A, B, C, D, E) Correlation analysis using the R package program. Each correlation is shown by the shades of blue and red and the size of the circle shape. * and ** represent correlations with Pvalue≤0.05 and Pvalue≤0.01, respectively. (a, b, c, d, e) Co-regulatory networks. The co-regulatory networks of 18 EgrGRAS genes under stress treatments were established based on the Pearson correlation coefficients (PCCs) of these gene pairs using transformed qPCR data