Src kinases and Rac1 function in basal protrusion formation. (A,B) Control-treated kidney new nephrons (A; PP3) show invasive basal protrusions (arrowhead) that are missing in Src family inhibitor-treated kidneys (B; PP2; arrowhead indicates basal surface). (C) Quantification of surface curvature revealed PP2 caused a significant loss of basal protrusions (DT, distal tubule). Data are mean±s.d. (PP3: 0.47±0.17, n=19; PP2: 0.23±0.09, n=13). (D) Basal protrusions in control new nephrons (arrowhead) are formed with a central actin core (phalloidin; magenta) and maintain a narrow invasive morphology. (D′) Enlarged magnification of a slice from the same image stack as in D highlights the actin core structure (phalloidin; magenta) of protrusions. (E) Rac1 inhibition with EHT1864 caused mislocalization of actin to basal protrusion cell membranes, distension of protrusions and a ‘blebbing’ morphology (arrowhead). (E′) Enlarged magnification of a slice from the same image stack as in E. (F) EHT1864-induced distension of basal protrusions quantified by protrusion cross-sectional area. Data are mean±s.d. (control: 2.21±1.13, n=31; Rac1 inhib: 5.262±3.13, n=34). Blue fluorescence is Hoechst-stained nuclei. Scale bars: 5 µm in A,B,D,E; 2 µm in D′,E′. ****P<0.0001 (unpaired t-test).
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