Fig 1: Expression and localization of TP63 isoforms and other selected factors associated with CE homeostasis and stratification in absence of Myc. (A, left panel) CE immunostaining of Krt12 (red) and DAPI (blue) on cross-sections of the paraffin-embedded cornea. The Abs utilized were directed to either the N- or the C-terminus of the Krt12 protein. Krt12 accumulation at the CE surface observed in the mutant using either Ab was severely reduced in the CE of the CKO. Bar graphs represent the percentage of Krt12 apical staining along the CE length of cornea cross-sections (mean ± SD, n = 3). (A, right panel) Line-intensity profile plot of Krt12 fluorescence signal across a CE cross-section. The y-axis represents the average intensity of 15 line-intensity scans (31 µm wide) from the basal membrane (BM) to the apical side of the CE taken every 100 µm along the length of each cornea section. The x-axis represents the length of the line scan from the BM to the apical side of the CE (n = 3). (B) CE immunostaining with Ab directed to the epithelial-specific ΔNp63 (n = 4/group). (C) CE immunostaining with the pan (TP) p63 Abs (n = 2 control and 4 CKO). The bar graphs in B and C describe the mean ± SD values of p63 intensities (green fluorescence) over the whole epithelium, normalized by the corresponding nuclear stain (blue fluorescence). (D, left panel) GAPDH-normalized CKO/control chemiluminescence signal ratios (mean ± SD). Number of independent replicates for each protein are indicated on top of the bars. Myc ablation led to a fourfold increase of ΔNp63 and about twofold increases in ETS1 and Notch 1. Krt14 and Krt 12 displayed no substantial differences in two duplicates and were not quantitated. (E) Level of expression of p63 isoforms relative to Gapdh in the wild-type mouse (mean ± SD, n = 3/group). (F) Expression of p63 isoforms and genes involved in the CE differentiation. Data are represented as the fold change in the CKO relative to the control (n = 3/group). ns, not significant (P ≥ 0.05, Student's t-test). *P < 0.05, **P < 0.001. Scale bars: 25 µm.