Genome-Wide Identification and Expression Analysis of HSF Transcription Factors in Alfalfa (Medicago sativa) under Abiotic Stress

Abstract

Alfalfa (Medicago sativa) is one of the most important legume forage species in the world. It is often affected by several abiotic stressors that result in reduced yields and poor growth. Therefore, it is crucial to study the resistance of M. sativa to abiotic stresses. Heat shock transcription factors (HSF) are key players in a number of transcriptional regulatory pathways. These pathways play an essential role in controlling how plants react to different abiotic stressors. Studies on the HSF gene family have been reported in many species but have not yet undergone a thorough analysis in M. sativa. Therefore, in order to identify a more comprehensive set of HSF genes, from the genomic data, we identified 16 members of the MsHSF gene, which were unevenly distributed over six chromosomes. We also looked at their gene architectures and protein motifs, and phylogenetic analysis allowed us to divide them into 3 groups with a total of 15 subgroups. Along with these aspects, we then examined the physicochemical properties, subcellular localization, synteny analysis, GO annotation and enrichment, and protein interaction networks of amino acids. Finally, the analysis of 16 MsHSF genes' expression levels across all tissues and under four abiotic stresses using publicly available RNA-Seq data revealed that these genes had significant tissue-specific expression. Moreover, the expression of most MsHSF genes increased dramatically under abiotic stress, further validating the critical function played by the MsHSF gene family in abiotic stress. These results provided basic information about MsHSF gene family and laid a foundation for further study on the biological role of MsHSF gene in response to stress in M. sativa.

Keywords: HSF gene family; Medicago sativa; abiotic stress; expression profile.

Figure 1

Model for phylogenetic analysis of HSF in M. sativa. Each subgroup is distinguished by a different color.

Figure 2

Conserved domain alignment of MsHSF members.

Figure 3

Three-dimensional model of the protein of MsHSF family members, with the starting position of the α1 DBD structural domain indicated by 1. (A–P) in the figure represent the 16 proteins of the M. sativa HSF family, respectively.

Figure 4

Phylogenetic relationship tree (A), gene structure (B) and conserved patterns (C) of HSF in M. sativa.

Figure 5

Distribution of MsHSF genes on the chromosomal scaffolds of M. sativa.

Figure 6

Chromosome distribution and interchromosomal relationships of MsHSF genes. Gray lines indicate synthetic blocks within the M. sativa genome, and red lines indicate duplicated MsHSF gene pairs.

Figure 7

Synthetic analysis of the M. sativa genome with the genomes of one monocotyledon and three dicotyledon plants. Gray lines represent alignment blocks between paired genomes, and blue lines indicate synthetic HSF gene pairs.

Figure 8

A schematic representation of the cis-acting elements identified in the 2000 bp promoter region upstream of the MsHSF gene. The different colors represent the number of cis-acting elements contained (A). The number of MsHSF genes corresponding to the cis-acting element (B).

Figure 9

A heat map representation of MsHSF expression between different tissues. The values in the rectangle represent the magnitude of the gene expression.

Figure 10

Expression of 16 MsHSF genes in cold (A), drought (B), ABA (C), and salt (D) treatments. The values in the rectangle represent the magnitude of the gene expression.

Figure 11

GO enrichment analysis of the MsHSF proteins relative to the GO database. The horizontal axis indicates the enrichment factor, and the size of the circle indicates the number of genes annotated with a given GO term.

Figure 12

Interaction network of HSF proteins in M. sativa. Nodes represent proteins; central nodes are indicated in blue, and black lines indicate interactions between nodes.