Fig. 9: PpREN disrupts the membrane clustering of PpRopGAP1.

a Competitive pull-down analyses of PpRopGAP1 and PpROP4CA in the presence of PpREN or the GAP domain of PpREN (rGAP). Equal amounts of MBP-PpRopGAP1 and MBP-PpREN (or MBP-rGAP) were pulled down by GST-PpROP4CA. b Localization of the free Nter portion of PpRopGAP1 and its fusion with the GAP domain of PpREN. The arrow indicates weak enrichment at the apical membrane. Scale bar: 10 µm. c Quantification of intensity ratio between membrane-associated and cytoplasmic proteins. The number of cells used for quantification is shown below the bars. d Localization of the endogenous PpRopGAP1 in WT and REN overexpression (REN-ox) lines. Scale bar: 10 µm. e Quantification of intensity ratio between membrane-associated and cytoplasmic PpRopGAP1 (left) and distances of PpRopGAP1 signal to the cell tip (right). The number of cells used for quantification is shown below the bars. f Kymographs showing PpRopGAP1 movement. Kymographs were generated at one lateral surface along the tip to the basal region. Note that PpRopGAP1 is enriched at the subapical membrane in WT cells and its signal extends to the basal membrane in REN-ox cells. Scale bars: horizontal, 10 µm; vertical, 10 s. g Quantification of velocities of PpRopGAP1-labeled particles. The numbers of mobile particles from WT (5 cells) and REN-ox (4 cells) cells are shown below the bars. Data in (c, e) are presented as box-and-whisker plots, showing the interquartile range (box), the median (horizontal line), minimum and maximum values (whiskers), and individual data points. Data in (g) are presented as violin plots, showing the quartiles (thin lines) and the median (central line). Statistical analyses were performed using two-tailed student’s t-tests. ns, not significant. *p < 0.05. ****p < 0.0001. The exact p-values are available in Source Data. Similar results in (a) were obtained in three independent experiments. Source data are provided as a Source Data file.