Fig. 4

ROP recruits the BDR-WAL complex to the plasma membrane. a, b BiFC (a) and FRET (b) assays between BDR and ROP11, ROP11G17V, or ROP11T22N in tobacco leaf epidermis. Values are mean ± s.d. (n > 9), **p < 0.01; ***p < 0.001 (ANOVA with Scheffe test). c BDR1-GFP and tagRFP-ROP11 in Arabidopsis-cultured cells. N indicates the nucleus. Graphs show intensity profiles along the white lines. BDR1-GFP is localized to the cytoplasm in the absence of ROP11 or in the presence of ROP11T22N, but is localized to the plasma membrane in the presence of ROP11 or ROP11G17V. Green and red arrowheads indicate peaks of BDR1-GFP and tagRFP-ROP11 intensity, respectively. Note that the BDR1-GFP peak overlaps with those of tagRFP-ROP11 and tagRFP-ROP11G17V but not with that of tagRFP-ROP11T22N. d Reconstitution of active ROP11 domains in leaf epidermal cells. ROP11, tagRFP-ROPGEF4PRONE, and ROPGAP3 were co-expressed. Either or both BDR1-ECFP and EYFP-WALΔN were co-expressed alongside. Intensity profiles along the white lines are shown at the right panels. Note that EYFP-WALΔN was present around ROPGEF4 clusters in the presence of BDR1-ECFP but not in the absence of BDR1-ECFP. e Schematic illustration of ROP signaling pathways in the pits. Activated ROP recruits BDR to the boundary of the pit membrane (red dotted circle), which in turn recruits WAL. WAL promotes assembly of actin microfilaments. The actin bundle directs secondary cell wall ingrowth to overarch the pit membrane (ROP-BDR-WAL-actin pathway). Besides the BDR pathway, activated ROP recruits the MIDD1-Kinesin-13A (KIN13A) complex to the entire area of pit membrane (yellow dotted circle) to promote disassembly of cortical microtubules (MT), thereby inhibiting deposition of secondary cell walls (ROP-MIDD1-Kinesin13A pathway). Bars = 50 µm (a) and 10 µm (c and d). Source data are provided as a Source Data file