Figure 5. BHP interacts with BLUS1 both in vitro and in vivo.

(a) In vitro pull-down assay of the interaction of BHP with BLUS1. Both proteins were expressed in E. coli. Extract from the E. coli cells expressing GST or GST-BHP was mixed with that expressing FLAG-BLUS1 and reacted with glutathione-Sepharose 4B beads. The proteins on the beads were subjected to SDS-PAGE and then immunoblotted using anti-GST and anti-FLAG antibodies. (b) In vivo interaction of BHP with BLUS1 determined by a bimolecular fluorescence complementation (BiFC) assay. BLUS1 constructs were co-transformed with BHP into Nicotiana benthamiana leaves. The reconstituted fluorescent signal was observed using a confocal laser microscope. YFP^N and YFP^C represent the N- and C-terminal halves of the YFP protein, respectively. Scale bar represents 100 µm.