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. 2013 Jul 1;110(29):12114–12119. doi: 10.1073/pnas.1302154110

Fig. 3.

Fig. 3.

BIK1 associates with BRI1. (A) BIK1 associates with BRI1 in protoplasts. BIK1-FLAG was coexpressed with BRI1-HA in Arabidopsis protoplasts. Co-IP was carried out with an α-FLAG antibody (IP: α-FLAG), and the proteins were analyzed by using Western blot with α-HA antibody. Top shows that BIK1-FLAG coimmunoprecipitated with BRI1-HA (IP: α-FLAG; WB: α-HA). Middle and Bottom show the expression of BRI1-HA and BIK1-FLAG proteins (WB: α-HA or α-FLAG for input control). Protoplasts were treated with 2 μM BL for 2 h. (B) BIK1 associates with BRI1 in transgenic plants. The membrane proteins from 4-wk-old pBIK1::BIK1-HA/pBRI1::BRI1-GFP (#6 and #9) or pBRI1::BRI1-GFP plants were immunoprecipitated with α-HA antibody and analyzed with Western blot using α-GFP antibody (Top). The expression of BRI1-GFP and BIK1-HA in transgenic plants are shown (Middle and Bottom). (C) BIK1 interacts with BRI1 cytosolic domain in vitro. GST-BIK1 proteins were incubated with MBP or MBP-BRI1CD beads (PD:MBP), and the beads were collected and washed for Western blot of immunoprecipitated proteins with α-GST antibody. Asterisk indicates nonspecific bands. The above experiments were repeated three times with similar results.