Inactivation of the calcium sensing receptor inhibits E-cadherin-mediated cell-cell adhesion and calcium-induced differentiation in human epidermal keratinocytes

Extracellular Ca2+ (Ca-o(2+)) is a critical regulator that promotes differentiation in epidermal keratinocytes. The calcium sensing receptor (CaR) is essential for mediating Ca2+ signaling during Ca-o(2+)-induced differentiation. Inactivation of the endogenous CaR-encoding gene CASR by adenoviral expression of a CaR antisense cDNA inhibited the Ca-o(2+)-induced increase in intracellular free calcium (Ca-i(2+)) and expression of terminal differentiation genes, while promoting apoptosis. Ca-o(2+) also instigates E-cadherin-mediated cell-cell adhesion, which plays a critical role in orchestrating cellular signals mediating cell survival and differentiation. Raising Ca-o(2+) concentration ([ Ca2+](o)) from 0.03 to 2 mM rapidly induced the co-localization of alpha-, beta-, and p120-catenin with E-cadherin in the intercellular adherens junctions (AJs). To assess whether CaR is required for the Ca-o(2+)-induced activation of E-cadherin signaling, we examined the impact of CaR inactivation on AJ formation. Decreased CaR expression suppressed the Ca-o(2+)-induced AJ formation, membrane translocation, and the complex formation of E-cadherin, catenins, and the phosphatidylinositol 3-kinase (PI3K), although the expression of these proteins was not affected. The assembly of the E-cadherin-catenin-PI3K complex was sensitive to the pharmacologic inhibition of Src family tyrosine kinases but was not affected by inhibition of Ca-o(2+)-induced rise in Ca-i(2+). Inhibition of CaR expression blocked the Ca-o(2+)-induced tyrosine phosphorylation of beta-, gamma-, and p120-catenin, PI3K, and the tyrosine kinase Fyn and the association of Fyn with E-cadherin and PI3K. Our results indicate that the CaR regulates cell survival and Ca-o(2+)-induced differentiation in keratinocytes at least in part by activating the E-cadherin/PI3K pathway through a Src family tyrosine kinase-mediated signaling.