Fig. 6: RGLG2 promotes the association of BAK1 and BIK1. | Nature Communications

Fig. 6: RGLG2 promotes the association of BAK1 and BIK1.

From: BIK1 protein homeostasis is maintained by the interplay of different ubiquitin ligases in immune signaling

Fig. 6

a RGLG2 associates with BAK1. RGLG2-FLAG was co-expressed with BAK1-HA in Arabidopsis protoplasts, protoplasts were treated with or without 2.5 μM flg22 for 30 min, and IPs were performed using anti-FLAG antibodies. The associated BAK1-HA proteins were detected by immunoblotting with anti-HA antibodies. b RGLG2-BIK1 association partially requires BAK1. RGLG2-FLAG and BIK1-HA were co-expressed in Arabidopsis protoplasts isolated from bak1 or Col-0 plants. IPs were performed using anti-FLAG antibodies. a, b Images shown are representative of two independent experiments. c RGLG2 promotes the association of BIK1 and BAK1. BIK1-HA and BAK1-FLAG were expressed together with or without RGLG2-GFP in Arabidopsis protoplasts. BAK1-FLAG proteins were immunoprecipitated with anti-FLAG antibodies. Images shown are representative of three independent experiments. d RGLG2 promotes the interaction between BIK1 and BAK1 in vitro. The recombinant GST-BIK1 proteins immobilized on GSH beads were incubated with purified His-BAK1CD proteins in the presence or absence of MBP-RGLG2-HA. GST pull-down was performed and the pulled-down His-BAK1CD was detected by immunoblotting with anti-His antibodies. Input proteins were separated on SDS-PAGE and stained with CBB. e BAK1, BIK1, and RGLG1/2 are in the same complex. BAK1-FLAG was co-expressed with BIK1-HA and RGLG2-HA in protoplasts. BAK1-FLAG proteins were immunoprecipitated with anti-FLAG antibodies. f The association of BAK1 and BIK1 is weaker in rglg1 rglg2 than in Col-0. BAK1-FLAG was co-expressed with BIK1-HA in rglg1 rglg2 or Col-0 protoplasts. BAK1-FLAG proteins were immunoprecipitated with anti-FLAG antibodies. d–f Images shown are representative of at least two independent experiments.

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