Transgenes used are as indicated. (A) Agroinfiltration-transfected tobacco leaf epidermal cells. Figure 9—figure supplement 1A shows FER-GFP and GFP-LLG1 localization when expressed alone. (B–E) Transformed Arabidopsis protoplasts. (B) Protoplasts with cell membrane-located reconstituted Venus signal (upper panel) and pronounced intracellular signal (lower panel) were observed on comparable levels (n>3 replicate assays, with ∼50 BiFC positive cells each). Figure 9—figure supplement 1 shows GFP-LLG1 localization when expressed alone, single vector controls, and reconstituted Venus signal in cells expressing the full-length FER as one of the split Venus pair. (C) The majority of protoplasts (>90%, n=triplicate samplings, ∼50 BIFC positive cells each) co-transformed by ARF1(Q71L) and the split Venus pair showed strong intracellular Venus signals that resembled the ER. Arrow indicates perinuclear; arrowheads indicate dynamic reticulate structure (see Videos 1, 2). (D, E) Colocalization of the split Venus signal with the ER marker, RFP-ER. Boxed areas in (D) were contrast-enhanced (equally) to highlight the signals. *, chlorophyll fluorescence. (A, C) Wide-field, (B, D, E) confocal images. Scale bars: 25 μm (A), 10 μm (B–E).