Figure 3.

A subset of JAZs interact with FAMA. Y2H assay to detect interactions of FAMA with the 12 JAZ proteins. Yeast cells cotransformed with pGADT7-FAMA, or pGADT7-AP2 (preys) and pGBKT7-JAZ1-12 (bait) were grown on yeast synthetic dropout lacking Leu and Trp (SD/-2) as transformation control or on selective media lacking Ade, His, Leu, and Trp (SD/-4) to test protein interactions. pGADT7-FAMA, and pGADT7-AP2 cotransformed with pGBDT7 vector were included as a control. Split-LUC assays showing that JAZ1 and JAZ9 can interact with FAMA in N. benthamiana leaves. Representative images of N. benthamiana leaves 72 h after infiltration are shown. The images were digitally extracted for comparison. The right part of pseudocolor bar shows the range of luminescence intensity in each image. The bottom part indicates the infiltrated constructs. Bars = 10 mm. Three biological replicates were performed, and similar results were obtained. C, Co-IP assays to verify the interaction of JAZ1 with FAMA in vivo. Protein extracts from transgenic plants carrying both 35S_pro:JAZ1-GUS and ProFAMA:FAMA-GFP (FAMA-GFP JAZ1-GUS) or from transgenic plants harboring 35S_pro:JAZ1-GUS (JAZ-GUS) were immunoprecipitated with GFP antibody, and immunoprecipitated proteins were analyzed by immunoblotting using anti-GUS and anti-GFP antibodies. The experiments were repeated 3 times with similar results.