Rga3 colocalizes with active Cdc42 and binds the cortex through its C1 domain. (A) Images of Rga3-GFP and Rga4-RFP. (B) Average cortical profiles of Cdc42-mCherry^SW and Rga3-GFP fluorescence intensity normalized to maximum and minimum values. n = 24 tips. (C) Images of Rga3-GFP strains expressing a pREP41-Cdc42-mCherry^SW (WT or Q61L) plasmid (left). Arrowheads indicate zones of Rga3-GFP at the cell sides. Average cortical profiles of Rga3-GFP fluorescence (right). In the strain carrying the Cdc42^Q61L allele, only rod-shaped cells were analyzed. n = 18. (D) Schematic representation of Rga3 fragments whose localization was tested upon GFP tagging and plasmid expression. Cortex-binding ability is summarized on the right. Representative images of plasmid-expressing rga3Δ cells are shown at the bottom. Note that Rga3 fragments are excluded from vacuoles. CC, coiled coil. (E) Image of cells expressing a triple-tandem copy of Rga3 C1 domain from plasmid. (F) Images of Rga3-GFP and Rga3^ΔC1-GFP expressed from the native genomic locus (left). Average cortical profiles of Rga3-GFP and Rga3^ΔC1-GFP fluorescence intensity (middle). Whole-cell intensity of Rga3-GFP and Rga3^ΔC1-GFP (right). n = 11. (G) Calcofluor and GFP fluorescence images of rga4Δrga6Δ cells expressing Rga3-GFP or Rga3^ΔC1-GFP (left). Average cell length and width of the same strains (n > 50; right). All micrographs are medial plane spinning-disk confocal images. Bars, 3 µm. Student’s t test P value is indicated. Error bars show SD.