Abstract
The plant defense responses to pests results in the synchronized change of a complex network of interconnected genes and signaling pathways. An essential part of this process is mediated by the binding of transcription factors to the specific responsive cis-elements within in the promoters of phytophagous-responsive genes. In this work, it is reported the identification and characterization of a bidirectional promoter that simultaneously co-regulate two divergent genes, At5g10300 and At5g10290, upon arthropod feeding. Computational analysis identified the presence of cis-elements within the intergenic region between two loci, mainly from the DOF but also from the AP2/ERF, Golden 2-like and bHLH families. The function of the bidirectional promoter was analyzed using two enhanced variants of the GFP and CherryFP reporter genes, in both orientations, in transient tobacco and stably transformed Arabidopsis plants. Promoter activity was tested in response to feeding of Tetranychus urticae and Pieris brassicae, as well as wounding, flagellin and chitin treatments. Using RT-qPCR assays and confocal microscopy, it was shown that all treatments resulted in the induction of both reporter genes. Furthermore, our findings revealed the asymmetric character of the promoter with stronger activity in the forward than in the reverse orientation. This study provides an example of a bidirectional promoter with a strong potential to be used in plant biotechnology in pest control that requires stacking of the defense genes.
Highlights
A bidirectional promoter is the intergenic region between two divergent genes located on complementary strands of the DNA which drives their coordinated transcription in opposite directions
Many binding sequences for transcription factors (TFs) were identified in this region. It is remarkable the presence of several conserved DOF-binding motifs (AAAG/CTTT), as well as some cis-elements potentially recognized by defense-related TFs from the AP2/ethylene response factors (ERF) (GCC box), Golden 2-like (GAATCT) and basic Helix-Loop-Helix, basic helix-loop-helix (bHLH) (CACGTG) TF families
Gene pairs oriented in divergent transcriptional configuration and controlled by bidirectional promoters are abundant in plant genomes
Summary
A bidirectional promoter is the intergenic region between two divergent genes located on complementary strands of the DNA which drives their coordinated transcription in opposite directions. These promoters control of the expression of gene pairs usually involved in the same or in related physiological processes (Mitra et al, 2009; Singh et al, 2009; Oropeza-Aburto et al, 2017). In a first search of bidirectional promoters in the Arabidopsis genome, Wang et al (2009) identified near 2,500 gene shared intergenic regions, enriched in regulatory elements essential for transcription. Comparative analysis of bidirectional promoters, performed in rice, Arabidopsis and Populus (Dhadi et al, 2009) and in three grass species such as maize, sorghum, and Brachypodium (Krom and Ramakrishna, 2010) provided very useful information about common cis-elements, regulatory mechanisms, co-expression and evolutionary dynamics
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