Molecular characterization, expression and functional analysis of acyl-CoA-binding protein gene family in maize (Zea mays)

Abstract

Background: Acyl-CoA-binding proteins (ACBPs) possess a conserved acyl-CoA-binding (ACB) domain that facilitates binding to acyl-CoA esters and trafficking in eukaryotic cells. Although the various functions of ACBP have been characterized in several plant species, their structure, molecular evolution, expression profile, and function have not been fully elucidated in Zea mays L.

Results: Genome-wide analysis identified nine ZmACBP genes in Z. mays, which could be divided into four distinct classes (class I, class II, class III, and class IV) via construction of a phylogenetic tree that included 48 ACBP genes from six different plant species. Transient expression of a ZmACBP-GFP fusion protein in tobacco (Nicotiana tabacum) epidermal cells revealed that ZmACBPs localized to multiple different locations. Analyses of expression profiles revealed that ZmACBPs exhibited temporal and spatial expression changes during abiotic and biotic stresses. Eight of the nine ZmACBP genes were also found to have significant association with agronomic traits in a panel of 500 maize inbred lines. The heterologous constitutive expression of ZmACBP1 and ZmACBP3 in Arabidopsis enhanced the resistance of these plants to salinity and drought stress, possibly through alterations in the level of lipid metabolic and stress-responsive genes.

Conclusion: The ACBP gene family was highly conserved across different plant species. ZmACBP genes had clear tissue and organ expression specificity and were responsive to both biotic and abiotic stresses, suggesting their roles in plant growth and stress resistance.

Keywords: Acyl-CoA binding protein (ACBP); Evolution; Expression profiles; Stress; Subcellular localization; Zea mays.

Fig. 1

The distribution of the ACBP family genes in different plant species. The left is the phylogenetic tree ACBP from 22 plant species; the middle is the genome size of 22 plant species; the right is the number of ACBP genes in 22 plant species. Dots of different colors represent the different species (Red, Eudicots; light green, Chlorophyta; yellow, Monocots; Green, Moss)

Fig. 2

The phylogenic relationships, gene structures, and conserved motifs of maize ACBP family genes. The phylogenetic tree was constructed using MEGA 7.1 software by the Neighbor-Joining method with 1000 bootstrap replicates. The different ACBP classes were marked with dots of different colors: green, class I; yellow, class II; blue, class III; red, class IV. The yellow boxes and grey lines represent exons and introns, respectively. The wide boxes of different colors represent different conserved motifs. The scales were referred to the lengths of exons, introns and motifs, respectively

Fig. 3

Putative regulatory cis-elements in maize ACBP family gene promoters. Boxes of different colors represent different cis-elements

Fig. 4

Subcellular localizations of ZmACBPs in tobacco leaf epidermal cells by confocal laser-scanning microscopy. a-f ZmACBPs-GFP fusion protein; g-k bright field; l-p Merged image of bright field with GFP fluorescence; d ER marker, ZmBiP-RFP. GFP control, 35S::GFP from pBI-eGFP vector control

Fig. 5

Tissue expression patterns of ACBP family genes in Zea mays. a Hierarchical clustering and classification of the nine ZmACBP genes based on RNA-seq data during development stages. b Tissue-specific expression patterns of the nine ZmACBP genes were in maize root, stem, leaf, silk, immature cob, anther, and kernel (10 DAP) by using Quantitative real-time PCR. The expression levels were normalized to expression levels of maize TUB-ribosylation factor. Data represent mean ± SE of three replicates. Different letters indicate values which departed significantly at the 0.01 among tissues

Fig. 6

Regional associations of ZmACBP genes associated with agronomic traits. a-c Three ZmACBP genes, ZmACBP3/4/7, were significantly associated with ear height (EH), kernel width (KW), and heading date (HD), respectively. The circles of different colors represent different SNPs in each plot, and the purple diamond represents the most significantly associated SNP. The x axis indicates the SNP location, and the y axis shows the log10(p) (P-value). d Heat map showing the correlation among ZmACBP genes and the oil content in maize kernel. Blue and red colors on the heat map indicate positive and negative correlations, respectively. Asterisk indicates a significant correlation at ≤0.01 level between gene expression and oil accumulation

Fig. 7

Quantitative real-time PCR analysis of ZmACBPs expression under stress treatments. a 200 mM NaCl, b 20% PEG6000, c wounding, d 4 °C, e 20 μmol/L Cu²⁺, and (f) infection with fungus infection (Ustilago maydis). The expression levels were normalized to that of TUB-ribosylation factor. Data represent mean ± SE of three replicates. Different letters indicate differences of the treatment compared to control at P < 0.01

Fig. 8

ZmACBP1 and ZmACBP3 positively regulated plant tolerance in A. thaliana. Phenotypes of 10-day-old vector control (VC) and ZmACBP1 transgenic line (OE1 and OE2) seedlings under without stress (a), 100 mM NaCl (b), and 200 mM mannitol (c). d Root lengths of VC and ZmACBP1 transgenic plants. Phenotypes of 10-day-old VC and ZmACBP3 transgenic line (OE1 and OE2) seedlings under without stress (e), 100 mM NaCl (f), and 200 mM mannitol (g). h Root lengths of VC and ZmACBP3 transgenic plants. Data represent mean ± SE of three replicates. Different letters indicate differences of transgenic lines compared to VC at P < 0.01

Fig. 9

Expression patterns of lipid metabolism-related genes (a) and stress-responsive genes (b) in vector control (VC) and ZmACBP3 transgenic Arabidopsis seedlings in response to 100 mM NaCl or 200 mM mannitol. Data represent mean ± SE of three replicates. Different letters indicate differences of transgenic lines compared to VC at P < 0.01