figshare
Browse
Fig 6.TIF (2.81 MB)

CLCuMuB βC1 represses JA responses though interfering with the integrity of SCFCOI1.

Download (2.81 MB)
figure
posted on 2016-06-17, 03:30 authored by Qi Jia, Na Liu, Ke Xie, Yanwan Dai, Shaojie Han, Xijuan Zhao, Lichao Qian, Yunjing Wang, Jinping Zhao, Rena Gorovits, Daoxin Xie, Yiguo Hong, Yule Liu

(A) Total root length of HA-βC1 transgenic (#2 HA-βC1 and #3 HA-βC1) and wild-type (#2 WT and #3 WT) N. benthamiana seedlings was measured every 24 h beginning at the 6th day after sowing (n ≥11). Bars represent SEM. #2 HA-βC1 and #2 WT were presented on same plates, while #3 HA-βC1 and #3 WT were presented on same plates. These experiments were repeated 3 times. (B) Jasmonate sensitivity was measured as root growth inhibition. Six-day-old seedlings (n ≥10) were grown on MS contained with 50 μM MeJA for additional 4 days. Bars represent SEM. The raw data were analysed by Mann-Whitney rank sum test to show the significance level at 0.05 (*). (C) Relative expression level of marker genes of jasmonate responses in mock- or MeJA-treated HA-βC1 transgenic and wild-type N. benthamiana seedlings determined by quantitative real-time PCR. #2 HA-βC1 and #2 WT were presented on same plates, while #3 HA-βC1 and #3 WT were presented on same plates. HA-βC1-expressing lines are compared with their corresponding control in each condition. Actin was used as the internal control. Bars represent SEM. The raw data were analysed by two-sample t-test to show the significance level at 0.05 (*), 0.01 (**) and 0.001 (***). These experiments were repeated at least twice. (D) CLCuMB βC1 enhanced degradation of COI1 in vitro. The purified Myc-COI1 protein was added to total protein extracts from N.benthamiana which expressed HA-nLUC or HA-βC1, incubated at 25°C for the indicated time periods, and subjected to immunoblot analysis with the anti-Myc antibody. Intensity was detected through Total Lab TL120. The PVDF membrane was stained with Ponceaux to visualize the large subunit of ribulose-1,5-bisphosphate as the loading control. (E) Quantitative analysis of the relative abundance of COI1 in the presence of HA-nLUC or HA-βC1 for the time periods indicated. The abundance of COI1 at the start point (0-h) was set to 100% as a reference for calculating its relative abundance after different incubation periods. Error bars represent SD. The experiment was repeated three times.

History