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The Ovate Family Proteins (OFPs) gene family houses a class of proteins that are involved in regulating plant growth and development. To date, there is no report of the simultaneous functional characterization of this gene family in all members of U's Triangle of Brassica. Here, we retrieved a combined total of 256 OFP protein sequences and analyzed their chromosomal localization, gene structure, conserved protein motif domains, and the pattern of cis-acting regulatory elements. The abundance of light-responsive elements like G-box, MRE, and GT1 motif suggests that OFPs are sensitive to the stimuli of light. The protein-protein interaction network analysis revealed that OFP05 and its orthologous genes were involved in regulating the process of transcriptional repression through their interaction with homeodomain transcription factors like KNAT and BLH. The presence of domains like DNA binding 2 and its superfamily speculated the involvement of OFPs in regulating gene expression. The biotic and abiotic stress, and the tissue-specific expression analysis of the RNA-seq datasets revealed that some of the genes such as BjuOFP30, and BnaOFP27, BolOFP11, and BolOFP10 were highly upregulated in seed coat at the mature stage and roots under various chemical stress conditions respectively which suggests their crucial role in plant growth and development processes. Experimental validation of prominent BnaOFPs such as BnaOFP27 confirmed their involvement in regulating gene expression under salinity, heavy metal, drought, heat, and cold stress. The GO and KEGG pathway enrichment analysis also sheds light on the involvement of OFPs in regulating plant growth and development. These findings have the potential to serve as a forerunner for future studies in terms of functionally diverse analysis of the OFP gene family in Brassica and other plant species.

Abstract Very long-chain fatty acids (VLCFAs) possess more than twenty carbon atoms and are the major components of seed storage oil, wax, and lipids. FAE (Fatty Acid Elongation) like genes take part in the biosynthesis of VLCFAs, growth regulation, and stress responses, and are further comprised of KCS (Ketoacyl-CoA synthase) and ELO (Elongation Defective Elongase) sub-gene families. The comparative genome-wide analysis and mode of evolution of KCS and ELO gene families have not been investigated in tetraploid Brassica carinata and its diploid progenitors. In this study, 53 KCS genes were identified in B. carinata compared to 32 and 33 KCS genes in B. nigra and B. oleracea respectively, which suggests that polyploidization might has impacted the fatty acid elongation process during Brassica evolution. Polyploidization has also increased the number of ELO genes in B. carinata (17) over its progenitors B. nigra (7) and B. oleracea (6). Based on comparative phylogenetics, KCS, and ELO proteins can be classified into eight and four major groups, respectively. The approximate date of divergence for duplicated KCS and ELO genes varied from 0.03 to 3.20 million years ago (MYA). Gene structure analysis indicated that the maximum number of genes were intron-less and remained conserved during evolution. The neutral type of selection seemed to be predominant in both KCS and ELO genes evolution. String-based protein-protein interaction analysis suggested that bZIP53, a transcription factor might be involved in the activation of transcription of ELO/KCS genes. The presence of biotic and abiotic stress-related cis-regulatory elements in the promoter region suggests that both KCS and ELO genes might also play their role in stress tolerance. The expression analysis of both gene family members reflect their preferential seed-specific expression, especially during the mature embryo development stage. Furthermore, some KCS and ELO genes were found to be specifically expressed under heat stress, phosphorus starvation, and Xanthomonas campestris infection. The current study provides a basis to understand the evolution of both KCS and ELO genes in fatty acid elongation and their role in stress tolerance.