The Calcium-Sensing Receptor-Dependent Regulation of Cell-Cell Adhesion and Keratinocyte Differentiation Requires Rho and Filamin A

Extracellular Ca2+ (Ca2+o) functioning through the calcium-sensing receptor (CaR) induces E-cadherin-mediated cell-cell adhesion and cellular signals mediating cell differentiation in epidermal keratinocytes. Previous studies indicate that CaR regulates cell-cell adhesion through Fyn/Src tyrosine kinases. In this study, we investigate whether Rho GTPase is a part of the CaR-mediated signaling cascade regulating cell adhesion and differentiation. Suppressing endogenous Rho A expression by small interfering RNA (siRNA)-mediated gene silencing blocked the Ca2+o-induced association of Fyn with E-cadherin and suppressed the Ca2+o-induced tyrosine phosphorylation of β-, γ-, and p120-catenin and formation of intercellular adherens junctions. Rho A silencing also decreased the Ca2+o-stimulated expression of terminal differentiation markers. Elevating the Ca2+o level induced interactions among CaR, Rho A, E-cadherin, and the scaffolding protein filamin A at the cell membrane. Inactivation of CaR expression by adenoviral expression of a CaR antisense complementary DNA inhibited Ca2+o-induced activation of endogenous Rho. Ca2+o activation of Rho required a direct interaction between CaR and filamin A. Interference of CaR-filamin interaction inhibited Ca2+o-induced Rho activation and the formation of cell-cell junctions. These results indicate that Rho is a downstream mediator of CaR in the regulation of Ca2+o-induced E-cadherin-mediated cell-cell adhesion and keratinocyte differentiation.