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

Abstract

Extracellular Ca2+ (Ca2+o) acting 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 the CaR regulates cell-cell adhesion through the Fyn/Src tyrosine kinases. Here 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, 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 Ca2+o level induced interactions among CaR, Rho A, E-cadherin and a scaffolding protein filamin A at the cell membrane. Inactivation of CaR expression by adenoviral expression of a CaR antisense cDNA inhibited Ca2+o-induced activation of endogenous Rho. Ca2+o-activation of Rho required a direct interaction between the 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.

The Calcium-Sensing Receptor as a Regulator of Cellular Fate in Normal and Pathological Conditions

“…The CaSR coordinates these activities through regulation of a myriad of signaling pathways, including EGFR signaling, MAPK cascades and Ecadherin signaling [3,36,37,47,[50][51][52][62][63][64]. However, few studies have assessed the cell-or tissue-specificity of the cascades activated by engagement of the CaSR or determined the particular roles of the signaling cascades, leaving considerable room for discovery in this field.…”

“…In addition, keratinocyte differentiation has been shown to depend on the physical interaction between the CaSR, filamin A and Rho. Moreover, the Rho-filamin axis would drive a diverse array of kinase activities [64,65].…”

“…Finally, CaSR signaling also has been implicated in regulating colonic epithelial cell differentiation and keratinocyte differentiation through the activation of E-cadherin signaling in a Rhodependent manner with filamin A involvement (Fig. 2) [62][63][64]. Chakrabarty and colleagues demonstrated that stimulation of the CaSR through extracellular calcium increased the expression of the tumor suppressor on colon carcinoma cells, and concomitantly reduced the activation of -catenin/T-cell factor, leading to the suppression of their malignant behavior [63].…”

Structural and Functional Diversity of Desmosomes

“…This scenario predicted that adherens junction components or a different factor, perhaps the calcium receptor, CaR, responded to extracellular calcium by initiating intracellular calcium release (Tu et al, 2011;Xie & Bikle, 2007;Tu et al, 2007;Tu et al, 2008). Confi rming the involvement of intracellular calcium, Darier disease patients carrying a mutant form of the intracellular calcium pump, SERCA2, suffer from severe skin erosions attributed to desmosome loss (Sakuntabhai et al, 1999).…”

Focal adhesion complex proteins in epidermis and squamous cell carcinoma

“…93 Filamin A has been shown to be required for calcium-induced keratinocyte differentiation in culture, though the mechanism by which this occurs is unknown. 94 Migfilin, also known as Filamin binding LIM protein 1, also binds to Filamin A and plays a role in the actin cytoskeleton. However, the FBLIM1 (gene that encodes Migfilin) knockout mouse is normal with no noticeable defect in skin or tissue morphology.…”