Fig. 2: Reduction of Cdc42 mobility at the plasma membrane compromises cell polarity and viability.

A Schematic figure of the constructed Cdc42 variants. The prenylation CAAX box is replaced by 1, 2 or 3 tandem copies of ritC (red), which targets Cdc42 peripherally to the plasma membrane (gray). Cdc42 is tagged with mCherry (purple) inserted in a poorly conserved region¹⁷. B D and r_D (diffusion and membrane unbinding rates, respectively) of Cdc42-mCherry^SW variants, derived from FRAP measurements. The measures were made at cell sides, which captures Cdc42-GDP dynamics. Points are best D and r_D fits for individual cells. Dashed contour lines indicate 96% of the maximum R² value of the averaged R² plots for all fits. Gray shaded regions have predicted tip occupation below indicated levels. C Airyscan2 images of Cdc42-mCherry^SW variants and CRIB-3GFP (marking Cdc42-GTP) in diploid cells also contain an untagged copy of WT cdc42. Representative image out of >100 imaged cells. D Average Cdc42 and CRIB cortical profiles from cell poles in diploid cells as in (C). Shaded areas show standard deviation. n \(\ge\) 84 half-tips for each strain. E Averaged airyscan2 images of Cdc42-mCherry^SW variants and CRIB-3GFP (marking Cdc42-GTP) in haploid cells. Representative image out of >100 imaged cells. F Average Cdc42 and CRIB cortical profiles from cell poles in haploid cells as in (E). Shaded areas show standard deviation. n \(\ge\) 24 half-tips for each strain. G Tetrad dissections from indicated diploid strains. Growth on rich non-selective (YE) and cdc42-mCherry^SW-XritC-selective (YE-G418) plates is shown. H Serial 10-fold dilutions of indicated strains on rich medium. Scale bars are 5 µm. For (B, D, F), source data are provided as a Source Data file.