Fig. 7: RDS interacts with RDL and promotes its condensation.

a Bimolecular fluorescence complementation (BiFC) assays showing that RDS interacts with RDL in Arabidopsis protoplasts. YNE, N-terminal half of yellow fluorescent protein (YFP); YCE, C-terminal half of YFP. Scale bars, 10 μm. b MBP pull-down assay showing that RDS interacts with RDL in vitro. MBP fused to RDL (MBP-RDL) was tested for binding to GST fused to RDS (GST-RDS). MBP alone was used as a negative control in the pull-down assays. M, molecular weight of the protein marker. c In vivo co-immunoprecipitation (Co-IP) assay validating the association between RDS-FLAG and RDM16-GFP. RDS-FLAG or FLAG was co-transfected into protoplasts prepared from RDM16:RDM16-GFP plants. M, molecular weights of the protein markers. d RDS promotes condensation of GFP-RDL in vitro. MBP was used as a negative control. Scale bars, 5 μm. e, f Quantification of GFP-RDL droplet size (e) and fluorescence intensity (f) in the samples with the addition of the control protein (MBP) (n = 810) or MBP-RDS (n = 438). g–i Overexpression of RDS promotes of RDL-GFP condensation in Arabidopsis root cells. Representative micrographs of root cells from Arabidopsis seedlings (g) and quantitative analysis of the RDL-GFP fluorescent spots (h) and intensity (i) are shown. In (g), scale bars, 10 μm. n = 154 (h, i). j RDS overexpression does not influence RDL protein abundance. In (e, f, h and i), significant differences were determined using two-sided non-parametric Mann–Whitney U test (*** P < 0.001). All experiments were performed at least three times with similar results.