Genome-wide analysis of the bHLH gene family in Chinese jujube (Ziziphus jujuba Mill.) and wild jujube

Abstract

Background: The bHLH (basic helix-loop-helix) transcription factor is one of the largest families of transcription factors in plants, containing a large number of members with diverse functions. Chinese jujube (Ziziphus jujuba Mill.) is the species with the highest economic value in the family Rhamnaceae. However, the characteristics of the bHLH family in the jujube genome are still unclear. Hence, ZjbHLHs were first searched at a genome-wide level, their expression levels under various conditions were investigated systematically, and their protein-protein interaction networks were predicted.

Results: We identified 92 ZjbHLHs in the jujube genome, and these genes were classified into 16 classes according to bHLH domains. Ten ZjbHLHs with atypical bHLH domains were found. Seventy ZjbHLHs were mapped to but not evenly distributed on 12 pseudo- chromosomes. The domain sequences among ZjbHLHs were highly conserved, and their conserved residues were also identified. The tissue-specific expression of 37 ZjbHLH genes in jujube and wild jujube showed diverse patterns, revealing that these genes likely perform multiple functions. Many ZjbHLH genes were screened and found to be involved in flower and fruit development, especially in earlier developmental stages. A few genes responsive to phytoplasma invasion were also verified. Based on protein-protein interaction prediction and homology comparison, protein-protein interaction networks composed of 92 ZjbHLHs were also established.

Conclusions: This study provides a comprehensive bioinformatics analysis of 92 identified ZjbHLH genes. We explored their expression patterns in various tissues, the flowering process, and fruit ripening and under phytoplasma stress. The protein-protein interaction networks of ZjbHLHs provide valuable clues toward further studies of their biological functions.

Keywords: Chinese jujube; Flower and fruit development; Phytoplasma; Protein-protein interaction; Tissue-specific expression; ZjbHLHs.

Fig. 1

The phylogenetic analysis of ZjbHLHs. The NJ tree was constructed from the protein sequences of ZjbHLHs using MEGA7 with 1000 bootstrap copies

Fig. 2

Conserved motifs of ZjbHLH proteins. The motifs in ZjbHLHs were identified by using Multiple Em for Motif Elicitation (MEME). In these ZjbHLH proteins, six conserved motifs were identified and displayed in different colors

Fig. 3

The chromosomal location of 70 ZjbHLH genes. Genes are mapped to jujube chromosomes by the Circos tool. The chromosomes of jujube are arranged in a circle

Fig. 4

The gene structure of ZjbHLHs in Chinese jujube. Introns and exons are represented by black lines and red boxes respectively and upstream/downstream are represented by green boxes

Fig. 5

Expression patterns of 37 ZjbHLH genes in seven tissues of jujube and wild type jujube by RT-PCR. ZjACT was used as an internal control. Left: jujube, from left to right: root, young branch, old branch, leaf, flower bud, flower, and fruit. Right: wild jujube, from left to right: root, branch, leaf, flower bud, flower, fruit and seed

Fig. 6

a Heat maps of the relative expression of ZjbHLH genes during flower development. FL1, bud emergence stage; FL2, inflorescence emergence stage; FL3, yellow bud stage; FL4, petal spread stage. Scaled log2 expression values based on qRT-PCR data are shown from blue to red, indicating low to high expression. b The protein-protein interaction analysis of four ZjbHLHs by STRING database

Fig. 7

a Heat maps of the relative expression of ZjbHLH genes during fruit ripening in ‘Lizao’ and ‘Yazao’. Y, young fruit; EWM, early white mature fruit; WM, white mature fruit; HR, half-red fruit; FR, full red fruit. Scaled log2 expression values based on qRT-PCR data are shown from blue to red, indicating low to high expression. b The protein-protein interaction analysis of two ZjbHLHs by STRING database

Fig. 8

Heat maps of the relative expression of ZjbHLH genes under phytoplasma stress. a The gene expression in diseased leaves was significantly lower than that in healthy leaves. b The gene expression in diseased leaves was significantly higher than that in healthy leaves. Scaled log2 expression values based on qRT-PCR data are shown from blue to red, indicating low to high expression, respectively

Fig. 9

A protein-protein interaction network for ZjbHLHs based on their orthologs in Arabidopsis. This network was predicted by online software STRING. ZjbHLH proteins were shown in brackets with Arabidopsis orthologs