Transcriptome and binding data indicate that citral inhibits single strand DNA-binding proteins.

The mechanism of phytotoxicity of citral was probed in Arabidopsis thaliana using RNA-Seq and in silico binding analyses. Inhibition of growth by 50% by citral downregulated transcription of 9156 and 5541 genes in roots and shoots, respectively, after 1 h. Only 56 and 62 genes in roots and shoots, respectively, were upregulated. In the shoots, the downregulation increased at 3 h (6239 genes downregulated, vs 66 upregulated). Of all genes affected in roots at 1 h (time of greatest effect), 7.69% of affected genes were for nucleic acid binding functions. Genes for single strand DNA binding proteins (SSBP) WHY1, WHY 2 and WHY3 were strongly downregulated in the shoot up until 12 h after citral exposure. Effects were strong in the root at just 1 h after the treatment and then at 12 and 24 h. Similar effects occurred with the transcription factors MYC-2, ANAC and SCR-SHR, which were also significantly downregulated for the first hour of treatment, and downregulation occurred again after 12 and 24 h treatment. Downregulation of ANAC in the first hour of treatment was significantly (P < 0.0001) decreased more than eight times compared to the control. In silico molecular docking analysis suggests binding of citral isomers to the SSBPs WHY1, WHY2, and WHY3, as well as with other transcription factors such as MYC-2, ANAC and SCR-SHR. Such effects could account for the profound and unusual effects of citral on downregulation of gene transcription.