Genome-wide identification, structural analysis and new insights into late embryogenesis abundant (LEA) gene family formation pattern in Brassica napus

Abstract

Late embryogenesis abundant (LEA) proteins are a diverse and large group of polypeptides that play important roles in desiccation and freezing tolerance in plants. The LEA family has been systematically characterized in some plants but not Brassica napus. In this study, 108 BnLEA genes were identified in the B. napus genome and classified into eight families based on their conserved domains. Protein sequence alignments revealed an abundance of alanine, lysine and glutamic acid residues in BnLEA proteins. The BnLEA gene structure has few introns (<3), and they are distributed unevenly across all 19 chromosomes in B. napus, occurring as gene clusters in chromosomes A9, C2, C4 and C5. More than two-thirds of the BnLEA genes are associated with segmental duplication. Synteny analysis revealed that most LEA genes are conserved, although gene losses or gains were also identified. These results suggest that segmental duplication and whole-genome duplication played a major role in the expansion of the BnLEA gene family. Expression profiles analysis indicated that expression of most BnLEAs was increased in leaves and late stage seeds. This study presents a comprehensive overview of the LEA gene family in B. napus and provides new insights into the formation of this family.

Figure 1. Phylogenetic analysis of the B. napus LEA genes.

LEA gene families are distinguished by different colors. The unrooted tree was generated using ClustalW in MEGA6 using the full-length amino acid sequences of the 108 B. napus LEA proteins.

Figure 2. Exon–intron organization of the BnLEA genes.

Double-sided wedge boxes represent exons, and different colors indicate different LEA gene families. Black lines represent introns, and untranslated regions (UTRs) are indicated by light-gray purple boxes. Numbered marks represent the splicing phases. Phase-0 is not marked. The exon and intron sizes can be estimated using the scale at the bottom.

Figure 3. Motif patterns of different BnLEA families and WebLogo plot of consensus motifs in each BnLEA gene family.

Representative B. napus LEA proteins were selected for alignment, and LEA motifs are shown as motif 1 (light blue box). The lengths of the proteins and motifs can be estimated using the scale at the bottom. The Pfam codes of the LEA motifs of each family are shown.

Figure 4. Distribution of BnLEA gene family members on B. napus chromosomes.

The 96 BnLEA genes for which exact chromosomal information was available in the database were mapped to the 19 B. napus chromosomes. The color of each gene indicates the corresponding family.

Figure 5. Synteny analysis map of gene clusters in B. napus chromosomes.

(A) Genes located on B. napus chromosome C4 are syntenic with genes of B. oleracea and A. thaliana. (B) Genes located on B. napus chromosome A9 are syntenic with genes of B. rapa and A. thaliana. The different colors of the gene IDs indicate their individual LEA families (brown: dehydrin; light green: LEA_4;bottle green: LEA_3; red: LEA_1; purple: LEA_6; blue: SMP).

Figure 6. Phylogenetic relationships and hypothetical evolutionary progress of the clustering of BnLEA genes in B. napus chromosome C5.

(A) Phylogenetic relationships of selected BnLEA genes in the cluster. (B) Hypothetical mechanism of BnLEA gene cluster formation. The letters T, S, and W in the schematic diagram of the hypothetical origins of BnLEA genes indicate putative tandem duplication, segmental duplication and whole-genome duplication, respectively.

Figure 7. Hierarchical clustering of the expression profiles of BnLEA genes in different tissues.

The log-transformed values of the relative expression levels of BnLEA genes were used for hierarchical cluster analysis (original data shown in Table S4). The color scale represents relative expression levels with increased transcript (yellow) or decreased transcript (purple). Early_stage seeds were got 19 weeks after seeding, late_stage seeds were 40 weeks after seeding.