Figure 5.

The Canonical LPIAR Degron Promotes JAZ8 Binding to COI1, Destabilizes JAZ8, and Restores JA Responsiveness.

(A) JAZ8^LPIAR interacts with COI1 in a JA-Ile–dependent manner. Pull-down reactions were performed in the presence (+) or absence (−) of 2.5 μM JA-Ile as described in the legend for Figure 1A. Protein bound to the indicted JAZ-His fusion protein was separated by SDS-PAGE and analyzed by immunoblotting with anti-Myc antibody for the presence of COI1-Myc.

(B) Transgenic lines overexpressing JAZ8^LPIAR are sensitive to JA. Root growth assays were performed with wild-type (WT), 35S:JAZ10 (JAZ10.1), 35S:JAZ8 (JAZ8), and 35S:JAZ8^LPIAR (JAZ8^LPIAR) seedlings. Data show the mean ± se for each genotype (n = 20). Asterisk denotes significant differences (P < 0.05, Student’s t test) in comparisons between the indicated transgenic line and the wild type.

(C) JAZ8^LPIAR-YFP is degraded in vivo in response to JA treatment. Six-day-old transgenic seedlings expressing JAZ8-YFP (left) or JAZ8^LPIAR-YFP (right) were pretreated with water or the 26S proteasome inhibitor MG132 (100 μM) for 75 min, at which time seedlings were treated with either MeJA (+MeJA, 50 μM) or water (mock control) for an additional 15 min. YFP signal in root tissue was visualized by fluorescence microscopy. Microscope settings were identical for all images. Bars = 50 μm.

[See online article for color version of this figure.]