Genome-wide identification, characterization and evolutionary analysis of the APETALA 2/ethylene responsive element-binding protein (AP2/EREBP), cytochrome c oxidase (COX) and lipid transfer protein (LTP) genes in Zea mays under drought stress

Abstract

Abstract AP2 (APETALA2)/EREBP (ethylene-responsive element-binding protein), cytochrome c oxidase (COX) and nonspecific lipid transfer proteins (LTP) playimportant roles in the response to drought stress. This is the first study to identify the COX gene in Zea mays L. via genome-wide analysis. The qRT‒PCR results indicated that AP2/EREBP, COX and LTPwere downregulated, with fold changes of 0.84, 0.53 and 0.31, respectively, after 12 hours of drought stress. Genome-wide analysis identified 78 AP2/EREBP, 6 COX and 10 LTP genes in Z. mays L.. Domain analysis confirmed the presence of the AP2 domain, Cyt_c_Oxidase_Vb domain and nsLTP1 in the AP2/EREBP, COX and LTP proteins, respectively. Motif and gene structure analyses indicated that genes with closer phylogenetic relationships exhibited more similar genetic structures. The gene structure results revealed that 21 of the 78 AP2/EREBP genes hadintrons. All the COX genes had introns, while 8 LTP genes had introns. Most of the AP2/EREBP and LTP proteins were located in the nucleus and the extracellular space, respectively, whereas COX proteins were predicted to be located in different organelles. Sixteen putative nuclear localization signals (NLSs) were predicted for 21 AP2/EREBP proteins. A total of 187 microRNAs were predicted to target the AP2/EREBP genes, 48 microRNAs were predicted to target COX proteins, and 30 microRNAs werepredicted to target LTP genes. The AP2/EREBP, COX and LTP genes contained defense and stress response elements, abscisic acid-responsive elements, methyl jasmonate (MeJA)-responsive elements, salylic acid and the MYB binding site (MBS) element, which are involved in the drought response. AP2/EREBP genes were found on the 10 chromosomes of Z. mays L.. COX genes were found on chromosomes 1, 3, 4, 5, 7 and 8. LTP genes were found on chromosomes 1, 3, 6, 8, 9 and 10. In the present study, the Ka/Ks ratios of the AP2/EREBPparalogous pairs indicated that the AP2/EREBP genes were influenced primarily by purifying selection, which indicated that the AP2/EREBP genes received strong environmental pressure during evolution. The Ka/Ks ratios of the COX-3/COX-4paralogous pairs indicate that the COX-3/COX-4 genes were influenced primarily by Darwinian selection (driving change). For the LTP genes, the Ka/Ks ratios of the LTP-1/LTP-10, LTP-5/LTP-3 and LTP-4/LTP-8 paralogous pairs indicate that these genes were influenced primarily by purifying selection, while the Ka/Ks ratios of the LTP-2/LTP-6 paralogous pairs indicate that thesegenes were influenced primarily by Darwinian selection. The duplication time of the AP2/EREBPparalogous gene pairs in Z. mays L.ranged fromapproximately 9.364 to 100.935 Mya. The duplication time of the COX-3/COX-4paralogous gene pair wasapproximately 5.217 Mya. The duplication time of the LTP paralogous gene pairs ranged from approximately 19.064 to 96.477 Mya. In the resulting phylogenetic tree, the AP2/EREBP proteins were classified into three distinct clades. The AP2/EREBP protein family (AP2) includes five different domain types according to the Phytozome-13 website: the AP2/ERF domain, the EREBP-like factor (EREBP), the ethylene responsive factor (ERF), the dehydration responsive element binding protein (DREB) and the SHN SHINE. COX and LTP proteins were classified into three distinct clades. Synteny analysis of the AP2/EREBP, COX and LTP genes revealed collinearity orthologous relationships in O. sativa, H. vulgare and A. thaliana. Gene Ontology enrichment analysis confirmed the functional role of the AP2/EREBP, COX and LTP proteins in stress responses. The results of the present study will improve the understanding of the functions of the AP2/EREBP, COX and LTP genes in response to drought stress.