Supplementary Figure 2: Rac1 regulates apical polarity induction in multipotent progenitors and apical domain size in Ngn3 cells. | Nature Cell Biology

Supplementary Figure 2: Rac1 regulates apical polarity induction in multipotent progenitors and apical domain size in Ngn3 cells.

From: EGFR signalling controls cellular fate and pancreatic organogenesis by regulating apicobasal polarity

Supplementary Figure 2

(a) Representative Rac1 western blot in E15.5 Ctrl and Rac1;pdxCre KO (Loading control: α-tubulin). (b) Representative 3D-projections of E14.5 Ctrl and Rac1;pdxCre KO dorsal pancreata stained for Muc1. (c) Polarity quantification in E15.5 Ctrl and Rac1;pdxCre KO. n = 3 pancreata, from two litters, pTT = 0.0324. (d,e) Representative sections of E12.5 Ctrl and Rac1;pdxCreER KO, stained for β-gal, Muc1 and E-cad, and polarity quantification. n = 3 pancreata, from two litters, pTT = 0.0002. (f,g) Apical width quantification of E12.5 and E15.5 Ctrl and WT/KO cells in Rac1;pdxCreER KO. E12.5: n = 3 pancreata (from two litters), Het/KO pTT = 0.356 (n.s.), Het/WT pTT = 0.942 (n.s.). E15.5: n = 3 pancreata (from two litters), pTT = 0.674 (n.s.. h) Representative sections of E15.5 Ctrl and Rac1;pdxCre KO stained for Ngn3 and Ecad. (i–k) Ngn3/Ecad mm2 (i), Sox9/Ecad mm2 (j) and Ngn3 + /Sox9 + quantification in E15.5 Ctrl and Rac1;pdxCre KO. (i) n = 4 pancreata (two litters), pTT = 0.0394; (j) n = 3 pancreata, from two litters, pTT = 0.0122; (k) n = 4 pancreata, from two litters, pTT = 0.7289. (l) Representative sections of E15.5 Ctrl and Rac1;pdxCre KO stained for Ngn3 and Ecad. (m) qPCR analysis of Ngn3 in E15.5 Ctrl and Rac1;pdxCre KO (normalization to β-actin). n = 7 Ctrl, 13 KO pancreata, (three litters), pTT = 0.0022. (n) Ngn3 intensity quantification in E15.5 Ctrl and Rac1;pdxCre KO. n = 3 pancreata from each condition (two litters), pTT = 0.01011. (o,p) Representative flow cytometry plots and quantification of%Ngn3High cells in E15.5 Ctrl and Rac1;pdxCre KO. n = 4 Ctrl, 5 KO pancreata, from two independent experiments, pTT = 0.0093. (q) Apical width quantification from E15.5 Ctrl and Rac1;pdxCre KO. n = 3 pancreata (two litters), pTT = 0.0132. (r) Apical volume quantification from E15.5 Ctrl and Rac1;pdxCre KO. n = 3 pancreata (two litters), pKW = 0.04953. (s,t) Representative sections of E17.5 Ctrl and Rac1;pdxCre KO stained for Insulin (Ins) and Ecad and quantification of β- and α-cell differentiation. n = 4 pancreata (two litters), Insulin(β): pTT = 0.0326, Glucagon(α): pTT = 0.6323. Error bars represent ± SEM. Scale bars, 20 μm (d,l), 50 μm (b,h), 100 μm (r). Statistic source data are found in Supplementary Table 3. Unprocessed blots are shown in Supplementary Fig. 11. Statistical analysis: two-tailed student’s t-test (pTT) or Kruskal-Wallis (pKW).

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