Evolutionary Analysis of Respiratory Burst Oxidase Homolog (RBOH) Genes in Plants and Characterization of ZmRBOHs

The respiratory burst oxidase homolog (RBOH), as the key producer of reactive oxygen species (ROS), plays an essential role in plant development. In this study, a bioinformatic analysis was performed on 22 plant species, and 181 RBOH homologues were identified. A typical RBOH family was identified only in terrestrial plants, and the number of RBOHs increased from non-angiosperms to angiosperms. Whole genome duplication (WGD)/segmental duplication played a key role in RBOH gene family expansion. Amino acid numbers of 181 RBOHs ranged from 98 to 1461, and the encoded proteins had molecular weights from 11.1 to 163.6 kDa, respectively. All plant RBOHs contained a conserved NADPH_Ox domain, while some of them lacked the FAD_binding_8 domain. Plant RBOHs were classified into five main subgroups by phylogenetic analysis. Most RBOH members in the same subgroup showed conservation in both motif distribution and gene structure composition. Fifteen ZmRBOHs were identified in maize genome and were positioned in eight maize chromosomes. A total of three pairs of orthologous genes were found in maize, including ZmRBOH6/ZmRBOH8, ZmRBOH4/ZmRBOH10 and ZmRBOH15/ZmRBOH2. A Ka/Ks calculation confirmed that purifying selection was the main driving force in their evolution. ZmRBOHs had typical conserved domains and similar protein structures. cis-element analyses together with the expression profiles of the ZmRBOH genes in various tissues and stages of development suggested that ZmRBOH was involved in distinct biological processes and stress responses. Based on the RNA-Seq data and qRT-PCR analysis, the transcriptional response of ZmRBOH genes was examined under various abiotic stresses, and most of ZmRBOH genes were up-regulated by cold stress. These findings provide valuable information for further revealing the biological roles of ZmRBOH genes in plant development and abiotic stress responses.