Characterization of the late embryogenesis abundant (LEA) proteins family and their role in drought stress tolerance in upland cotton

Abstract

Background: Late embryogenesis abundant (LEA) proteins are large groups of hydrophilic proteins with major role in drought and other abiotic stresses tolerance in plants. In-depth study and characterization of LEA protein families have been carried out in other plants, but not in upland cotton. The main aim of this research work was to characterize the late embryogenesis abundant (LEA) protein families and to carry out gene expression analysis to determine their potential role in drought stress tolerance in upland cotton. Increased cotton production in the face of declining precipitation and availability of fresh water for agriculture use is the focus for breeders, cotton being the backbone of textile industries and a cash crop for many countries globally.

Results: In this work, a total of 242, 136 and 142 LEA genes were identified in G. hirsutum, G. arboreum and G. raimondii respectively. The identified genes were classified into eight groups based on their conserved domain and phylogenetic tree analysis. LEA 2 were the most abundant, this could be attributed to their hydrophobic character. Upland cotton LEA genes have fewer introns and are distributed in all chromosomes. Majority of the duplicated LEA genes were segmental. Syntenic analysis showed that greater percentages of LEA genes are conserved. Segmental gene duplication played a key role in the expansion of LEA genes. Sixty three miRNAs were found to target 89 genes, such as miR164, ghr-miR394 among others. Gene ontology analysis revealed that LEA genes are involved in desiccation and defense responses. Almost all the LEA genes in their promoters contained ABRE, MBS, W-Box and TAC-elements, functionally known to be involved in drought stress and other stress responses. Majority of the LEA genes were involved in secretory pathways. Expression profile analysis indicated that most of the LEA genes were highly expressed in drought tolerant cultivars Gossypium tomentosum as opposed to drought susceptible, G. hirsutum. The tolerant genotypes have a greater ability to modulate genes under drought stress than the more susceptible upland cotton cultivars.

Conclusion: The finding provides comprehensive information on LEA genes in upland cotton, G. hirsutum and possible function in plants under drought stress.

Keywords: Cotton (Gossypium spp); Drought; Gene expression; Gene ontology; Genome; Identification; LEA proteins; miRNAs.

Fig. 1

Phylogenetic tree, gene structure and motif compositions of LEA genes in upland cotton. The phylogenetic tree was constructed using MEGA 6.0. Exon/intron structures of LEA genes in upland cotton, exons introns and up / down-stream were represented by yellow boxes, black lines and blue boxes, respectively. Protein motif analysis represented by different colours, and each motif represented by number

Fig. 2

Phylogenetic relationship of LEA genes in three cotton species with Arabidopsis and Pinus tabuliformis. Neighbor-joining phylogeny of 242 genes for G. hirsutum, 136 genes for G. arboreum, 146 genes for G. raimondii, 30 genes for Pinus tabuliformis and 51 Arabidopsis LEA protein sequences, as constructed by MEGA 6.0. The difference colours mark the various LEA gene types

Fig. 3

LEA genes distribution in tetraploid upland cotton, Gossypium hirsutum chromosomes. Chromosomal position of each LEA genes was mapped according to the upland cotton genome. The chromosome number is indicated at the top of each chromosome

Fig. 4

LEA genes distribution in A and D cotton chromosomes: Chromosomal position of each LEA genes was mapped according to the upland cotton genome. The chromosome number is indicated at the top of each chromosome. a: chromosomes of the diploid cotton of A genome, G. arboreum; b: chromosomes of the diploid cotton of D genome, G. raimondii

Fig. 5

Syntenic relationships among LEA genes from G. hirsutum, G. raimondii and G. arboretum. G. hirsutum, G. raimondii and G. arboretum chromosomes are indicated in different colours. The putative orthologous LEA genes between G. hirsutum and G. raimondii, G. hirsutum and G. arboretum, and G. raimondii and G. arboretum by different colours

Fig. 6

Gene Ontology (GO) annotation results for upland cotton LEA genes. GO analysis of 242 LEA protein sequences predicted for their involvement in biological processes (BP), molecular functions (MF) and cellular components (CC). For the results presented as detailed bar diagrams, as illustrated in Additional file 5: Table S4

Fig. 7

Average number of the cis-promoters ABRELATERD1 (ACGTG), DRECRTCOREAT (G/ACCGAC), MYBCORE (TAACTG), LTRE1HVBLT49 (CCGAC) and others in promoter region of Gossypium hirsutum LEA genes from each LEA families. The promoter regions were analyzed in the 1 kb upstream promoter region of translation start site using the PLACE database

Fig. 8

Differential expression of upland cotton LEA genes under drought stress. The heat map was visualized using Mev.exe program. (Showed by log₂ values) in control, and in treated samples 7 and 14 days after drought treatment. a – BC₂F₁ (offspring), b – Gossypium tomentosum and c – Gossypium hirsutum. (i) Yellow – up regulated, blue – down regulated and black- no expression. (i). Percentage of genes exhibiting different responses to dehydration in leaf, root and stem of BC₂F₁; (ii). Percentage of genes exhibiting different responses to dehydration in leaf, root and stem of Gossypium tomentosum (iii). Percentage of genes exhibiting different responses to dehydration in leaf, root and stem of Gossypium hirsutum

Fig. 9

Quantitative PCR analysis of the selected LEA genes. Abbreviations: Rt (root), Sm (stem) and Lf (leaf). 0, 7 and 14 days of stress. Gh - Gossypium hirsutum, Gt - Gossypium tomentosum and BC-BC₂F₁ offspring. Y-axis: relative expression (2^−ΔΔCT)