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Fig. 5. HK1.fos/ ${Delta}$ 5Pten KAs exhibit high p53 and novel p21^WAF expression associated with a threshold level of GSK3β inactivation. Western analysis of p53, total (t-) and phosphorylated (p-) GSK3β and AKT were compared with p21^WAF, cyclin D1 and cyclin E2 expression in pathology-matched KAs (similar keratosis/papilloma ratios) and age-matched preneoplastic phenotypes. Hyperkeratotic (HK) K14.cre/ ${Delta}$ 5Pten^flx ear epidermis displayed little detectable p21^WAF or p53, and slightly increased expression of P-AKT, cyclin D1 and cyclin E2, with P-GSK3β being higher in the tagged (T) wound-promoted biopsy. Normal (N) appearing HK1.fos epidermis was negative for p21^WAF and p53 expression, with low P-GSK3β and decreased P-AKT levels, alongside slightly elevated cyclin D1. HK1.fos papillomas (PAP) expressed little p21^WAF and p53, but displayed increased P-GSK3β expression compared with P-AKT, together with elevated cyclins. HK1.fos/ ${Delta}$ 5Pten^flx epidermis (HK) expressed barely detectable p21^WAF, limited p53 and moderate P-GSK3β expression, whereas P-AKT expression was less than K14.cre/ ${Delta}$ 5Pten^flx controls. All HK1.fos/ ${Delta}$ 5Pten^flx KAs expressed high levels of p21^WAF and p53 that mirrored significant increases in P-GSK3β inactivation. However, P-AKT expression remained similar to K14.cre/ ${Delta}$ 5Pten^flx epidermis. All KAs exhibited elevated cyclin D1 and cyclin E2 expression, particularly in ear-tagged samples (KA^T). β-Actin served as a loading control.

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