Functional and regulatory diversity of homeobox-leucine zipper transcription factors BnaHB6 under dehydration and salt stress in Brassica napus L.

The plant-specific homeodomain-leucine zipper I subfamily is involved in the regulation of various biological processes, particularly growth, development and stress response. In the present study, we characterized four BnaHB6 homologues from Brassica napus. All BnaHB6 proteins have transcriptional activation activity. Structural and functional data indicate the complex role of BnaHB6 genes in regulating biological processes, with some functions conserved and others diverged. Transcriptional analyzes revealed that they are induced in a similar manner in different tissues but show different expression patterns in response to stress and circadian rhythm. Only the BnaA09HB6 and BnaC08HB6 genes are expressed under dehydration and salt stress, and in darkness. The partial transcriptional overlap of BnaHB6s with the evolutionarily related genes BnaHB5 and BnaHB16 was also observed. Transgenic Arabidopsis thaliana plants expressing a single proBnaHB6::GUS partially confirmed the expression results. Bioinformatic analysis allowed the identification of TF-binding sites in the BnaHB6 promoters that may control their expression under stress and circadian rhythm. ChIP-qPCR analysis revealed that BnaA09HB6 and BnaC08HB6 bind directly to the promoters of the target genes BnaABF4 and BnaDREB2A. Comparison of their expression patterns in the WT plants and the bnac08hb6 mutant showed that BnaC08HB6 positively regulates the expression of the BnaABF4 and BnaDREB2A genes under dehydration and salt stress. We conclude that four BnaHB6 homologues have distinct functions in response to stress despite high sequence similarity, possibly indicating different binding preferences with BnaABF4 and BnaDREB2A. We hypothesize that BnaC08HB6 and BnaA09HB6 function in a complex regulatory network under stress.

Evolutionary Consequences of Functional and Regulatory Divergence of HD-Zip I Transcription Factors as a Source of Diversity in Protein Interaction Networks in Plants.

The HD superfamily has been studied in detail for several decades. The plant-specific HD-Zip I subfamily attracts the most attention because of its involvement in plant development and stress responses. In this review, we provide a comprehensive insight into the evolutionary events responsible for the functional redundancy and diversification of the HD-Zip I genes in regulating various biological processes. We summarized the evolutionary history of the HD-Zip family, highlighting the important role of WGDs in its expansion and divergence of retained duplicates in the genome. To determine the relationship between the evolutionary origin and functional conservation of HD-Zip I in different species, we performed a phylogenetic analysis, compared their expression profiles in different tissues and under stress and traced the role of orthologs and paralogs in regulating developmental processes. We found that HD-Zip I from different species have similar gene structures with a highly conserved HD and Zip, bind to the same DNA sequences and are involved in similar biological processes. However, they exhibit a functional diversity, which is manifested in altered expression patterns. Some of them are involved in the regulation of species-specific leaf morphology and phenotypes. Here, we discuss the role of changes in functional domains involved in DNA binding and protein interaction of HD-Zip I and in cis-regulated regions of its target genes in promoting adaptive innovations through the formation of de novo regulatory systems. Understanding the role of the HD-Zip I subfamily in organism-environment interactions remains a challenge for evolutionary developmental biology (evo-devo).