EIN3 is a transcriptional activator of <i>YUC9</i>.

<p><b>(A)</b> Left panel: Schematic diagrams of effector and reporter constructs used in the transient dual-luciferase assays. <i>CaMV 35S</i> promoter driving <i>EIN3</i> (<i>35S</i>:<i>EIN3</i>) was used as effector, and empty vector as a negative control. Promoter fragments of <i>YUC9</i> were used to make the <i>YUC9p-LUC</i> reporter. Right panel: Transient dual-luciferase assay shows that <i>EIN3</i> transactivates the promoter of <i>YUC9</i> in <i>Arabidopsis</i> protoplasts. Data represent the means of three biological replicates. Error bars represent Student’s <i>t</i> test confidence intervals (n ≥ 9). Statistical significant difference indicated by asterisks (Fisher’s exact test, **P < 0.01). <b>(B)</b> Physical interactions of EIN3 with <i>YUC9</i> promoter in Y1H assays. Yeast expression plasmids pGADT7-EIN3 were reintroduced into yeast strain Y1H Gold carrying the reporter gene <i>AbAr</i> under the control of the <i>YUC9</i> promoter. The transformants were screened for their growth on the yeast synthetic defined media (SD/-Leu) in the presence of 50 or 100 ng ml^-1 Aureobasidin A (AbA) for stringent selection. The empty vector pGADT7 was included as a negative control. Yeast cultures were diluted (1:10 successive dilution series) spotted onto plates. <b>(C)</b> EIN3 associated with the promoter of <i>YUC9</i> in ChIP-qPCR assay. Up panel: Schematic diagrams of <i>YUC9</i> promoter of showing the potential EIN3 binding site (black triangles). The translational start sites (ATG) are shown as +1. Numbers above the diagram indicated the distance away from ATG. DNA fragments (P1 and P2) were used for ChIP. Chromatins isolated from <i>35S</i>:<i>EIN3-GFP</i> transgenic line and <i>35S</i>:<i>GFP</i> control were immunoprecipitated with anti-GFP antibody followed by qPCR to amplify P1 and P2 regions. Segment C located in the coding region was used as negative control. Input sample was used to normalize the qPCR results in each ChIP. Fold enrichment was presented as a ratio of normalized results from <i>35S</i>:<i>EIN3-GFP</i> plants and <i>35S</i>:<i>GFP</i>. Data are mean ± SD.