Differences in the calcium-mediated regulation of gap junctional intercellular communication between a cell line consisting of initiated cells and a carcinoma-derived cell line.

Abstract

Differences in calcium-mediated regulation of gap junctional intercellular communication (GJIC) between a cell line consisting of mouse epidermal initiated cells (3PC) and a mouse epidermal carcinoma-derived cell line (CA3/7) were studied. Under low extracellular calcium ((Ca2+)e) conditions (0.05 mM) CA3/7 cells showed a low level of GJIC compared with 3PC cells. High (Ca2+)e (1.20 mM) raised GJIC between CA3/7 cells to the GJIC level of 3PC cells, which in turn remained unchanged under these conditions. Raising the free intracellular calcium concentration ((Ca2+)i), using a calcium ionophore (ionomycin) or the Ca2+-ATPase inhibitor thapsigargin under low (Ca2+)e conditions, did not affect the GJIC level between 3PC cells, and increased GJIC between CA3/7 cells. Intracellular calcium chelation in 3PC cells under low (Ca2+)e conditions by ethylene glycol-bis(beta-amino-ethyl ether) N,N,N',N'-tetra-acetic acid acetoxy-methyl ester (EGTA-AM) decreased GJIC in this cell line. High (Ca2+)e conditions protected both cell lines from a decreased GJIC by EGTA-AM exposure. Inhibition of calmodulin (CaM) by calmidazolium (CDZ) or N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide (W-7) under low (Ca2+)e conditions, inhibited GJIC in 3PC cells and increased GJIC in CA3/7 cells. Inhibition of Ca2+/CaM-dependent protein kinase (Ca2+/CaM-PK) by 1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-7) decreased GJIC in both cell lines. Western analysis showed that Cx43 was more phosphorylated in both cell lines in concurrence with different effects on the GJIC level. Under conditions in which GJIC was inhibited, a decreased immunostaining of Cx43 on the plasma membrane was found. The level of immunostaining of the cell adhesion molecule E-cadherin on the plasma membranes of both cell types remained unchanged under conditions in which GJIC was changed by modulaters of (Ca2+)i, CaM activity, or the Ca2+/CaM-PK activity. These results indicate that differences exist between 3PC cells and CA3/7 cells in the GJIC regulation by intracellular calcium and calmodulin.