A strong NF-κB p65 responsive cis-regulatory sequence from Arabidopsis thaliana interacts with WRKY40.

KEY MESSAGE: Transcription factors from mammals and plants, which play a role in innate immunity, interact with the same microbe-associated molecular pattern (MAMP)-responsive sequences from Arabidopsis thaliana. The interaction of mouse NF-κB p65 with MAMP-responsive sequences containing the core motif GACTTT of the WT-box was investigated. This revealed one sequence, derived from the promoter of the A. thaliana gene At1g76960, a gene with unknown function, to activate NF-κB p65 dependent reporter gene expression in plant cells very strongly. A bioinformatic analysis predicts three putative NF-κB p65 binding sites in this sequence and all three sites are required for reporter gene activation and binding. The sequence is one of the weakest MAMP-responsive sequences previously isolated, but the introduction of a GCC-box increases its MAMP responsivity in combination with upstream WT-box sequences. Although a bioinformatic analysis of the unmutated cis-sequence only predicts NF-κB p65 binding, A. thaliana WRKY40 was selected in a yeast one-hybrid screen. WRKY40, which is a transcriptional repressor, requires the sequence TTTTCTA for direct binding. This sequence is similar to the WK-box TTTTCCAC, previously shown to interact with tobacco NtWRKY12. In summary, this work supports the similarity in binding site recognition between NF-κB and WRKY factors.

Transcription factors involved in basal immunity in mammals and plants interact with the same MAMP-responsive cis-sequence from Arabidopsis thaliana.

KEY MESSAGE: WRKY and NF-κB transcription factors, involved in innate immunity in plants and mammals, interact with the same cis-sequence. Novel microbe-associated molecular pattern (MAMP)-responsive cis-sequences, designated type II WT-boxes, are required for flg22-responsive gene expression in Arabidopsis thaliana protoplasts. While type I WT-boxes like TGACTTTT and CGACTTTT interact with WRKY transcription factors (TFs), the question remained which TFs bind to the type II WT-boxes GGACTTTC, GGACTTTT, and GGACTTTG. Surprisingly, a bioinformatic analysis predicts mouse (Mus musculus) NF-κB p65 as a TF interacting with type II WT-boxes. NF-κB p65, like WRKY factors in plants, plays a role in innate immunity in mammals. Therefore, the interaction of NF-κB p65 with type II WT-boxes was tested experimentally. NF-κB p65 requires the WT-boxes GGACTTTC, GGACTTTT, and GGACTTTG for activating reporter gene expression in plant cells. NF-κB p65 directly binds to WT-box containing synthetic promoters in vitro and requires the WT-box for binding. Earlier studies indicate that the sequence GGACTTTC is also required for WRKY26 mediated reporter gene activation. Here it is shown that WRKY26, like NF-κB p65, binds to the sequence GGACTTTC. Consistent with other recent studies, type II WT boxes are WRKY binding sites and the distinction between type I and type II no longer applies.