Fig. 1

CNF1 affects epithelial cell growth. (A) MTT assay of IEC-6 cells after 6 days of culture in the presence of scalar concentrations of CNF1 or CNF1-C866S. One representative experiment out of two is shown. (*p < 0.05; ***p < 0.001). (B) Trypan blue exclusion count of IEC-6 after 3 and 6 days of culture in the presence of the indicated concentrations of CNF1 or CNF1-C866S (N = 3). One representative experiment out of three is shown. (*p < 0.05 vs. CNF1-C866S and PBS). (C) Percentage of Annexin V and/or PI-positive cells after 6 days of culture in the presence of CNF1 or CNF1-C866S (25 ρM) (N = 3). One representative experiment out of three is shown. (***p < 0.001 vs. CNF1-C866S and PBS). (D) Histogram plot of the cell cycle distribution of untreated IEC-6 cells or IEC-6 cells exposed to 25 ρM CNF1 for 24 h. One representative experiment out of three is shown. (E) Western blot analysis of p21, Cyclin D1, p53 and phosphorylated p53 (pp53), in cell lysates from untreated and CNF1-treated IEC-6 cells at different time-points from treatment. One representative experiment out of two is shown. (F) Western blot analysis of the hypophosphorylated (pRB) and hyperphosphorylated (ppRB) forms of RB in cell lysates from untreated and CNF1-treated IEC-6 cells for the indicated times. (G) Representative fluorescence micrographs of F-actin staining in untreated and CNF1-treated IEC-6 24 h after treatment. Scale bar, 10 μm