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. 2018 Jul 20;178(1):428–440. doi: 10.1104/pp.18.00683

Figure 1.

Figure 1.

The ubiquitin E3 ligase RHA2b interacts with MYB30 in vivo. A, ABA induced the degradation of MYB30. Eight-day-old Col-0/35S:FLAG-MYB30 transgenic seedlings were treated without or with 100 μm ABA for 24 h and then incubated in the same medium with 100 μm CHX. Samples were collected 0, 6, and 12 h after treatment. MYB30 protein was detected with anti-FLAG antibody, and ACTIN was used as a loading control. The protein level compared with ACTIN at the initial time 0 was identified as 100%. Data represent mean values of three independent experiments ± sd. B, ABA-induced degradation of MYB30 was mediated by the 26S proteasome. Eight-day-old Col-0/35S:FLAG-MYB30 transgenic seedlings were treated with 100 μm ABA for 24 h and then incubated in the same medium with 100 μm CHX and 75 μm MG132. C, Coimmunoprecipitation (Co-IP) assays between MYB30 and RHA2b. MYC-MYB30 and FLAG-RHA2b were transiently expressed in Arabidopsis protoplasts. Soluble extracts from the protoplasts were detected directly (Input) or immunoprecipitated with an anti-MYC antibody (IP: MYC) and detected with anti-MYC and anti-FLAG antibodies (WB: MYC or FLAG). D, Split-YFP complementation imaging assays in Arabidopsis protoplasts. MYB30-cYFP and RHA2b-nYFP or RHA2a-nYFP were coexpressed in Arabidopsis protoplasts. MYB3 was used as a negative control. 4′,6-Diamino-phenylindole (DAPI) was used to stain the nuclei. DIC, Differential interference contrast. Bars = 50 μm.