Protein Kinase-C Regulates Keratin K15 Expression In Differentiating Keratinocytes

Objectives: K15 is a keratin which is specifically expressed in the basal keratinocytes of most stratified epithelia including oral and other internal epithelia. Studies have shown K15 to be a bulge stem cell marker in human hair follicles. In basal cell carcinoma (BCC) the expression of this protein is reportedly induced. About 80% of BCCs are reported to occur in the head and neck region. However, no study has yet characterised the mechanism of K15 expression in normal keratinocytes. Therefore, the aim of this study was to define a molecular mechanism that regulates K15 expression in stratified epithelia. Methods: N/Tert-1 keratinocytes were induced to differentiate by suspension culture, phorbol ester and culture density. The basal and differentiation-specific gene expressions were quantified using real time absolute qPCR and the protein levels were analysed by western blotting. Role of PKC pathway was established by using known PKC inhibitor GF109203X in suspension and monolayer cultures. Isoform specific PKC siRNAs were used to investigate the role of candidate PKC isoforms involved in the regulation of K15 expression. Results: Here we report for the first time that K15 expression is activated along with other differentiation-specific markers during keratinocyte differentiation induced by suspension culture, phorbol ester or high culture density. These observations are consistent with suprabasal expression of K15 in foetal and adult esophagus as reported in literature. Inhibition of PKC pathway by GF109203X showed suppression of K15 expression in suspended keratinocytes but had no effect on density induced K15 expression. Knockdown of PKC eta showed significant reduction of K15 expression in suspension culture. Conclusion: The data therefore suggests that there are two different mechanisms of K15 induction during differentiation; one is by suspension culture which is regulated by PKC pathway while the other is cell-cell junction mediated.