Increased connexin-43 and gap junctional communication correlate with altered phenotypic characteristics of cells overexpressing the receptor for hyaluronic acid-mediated motility.

Gap junctional intercellular communication (GJIC) and connexin expression are often altered during cell migration, growth, and transformation, each of which is accompanied by cytoskeletal reorganization. Recently, transfection of fibroblast cells with various isoforms of the receptor for hyaluronic acid-mediated motility (RHAMM) was shown to have profound and differential effects on motility, growth, and cell contact behavior as well as on elements of the actin-containing cytoskeleton. These cells thus provide an ideal system in which to investigate parameters implicated in regulation of GJIC as well as expression of connexin-43 (Cx43) in fibroblasts. We used 10T1/2 fibroblast cell lines transfected with RHAMM isoforms or a dominant negative mutated form of RHAMM that blocks the function of endogenous RHAMM. Increased RHAMM expression in the various cell lines was correlated with increased Cx43 and GJIC. These changes were accompanied by a loss of contact inhibition and decreased focal adhesions in all, and elevated motility of most but not all, cell lines tested. RHAMM-induced transformation also resulted in elevated GJIC and Cx43 levels. Reversion to normal growth, motility, and focal adhesion density following transfection of H-ras-transformed fibroblasts with the mutant form of RHAMM was associated with decreases in both Cx43 expression and GJIC. Transfection of 10T1/2 fibroblasts with RHAMM II (exons 5-14) produced altered contact behavior and increased both Cx43 and GJIC but had no effect on motility. All cells expressing high levels of RHAMM, regardless of the isoform, exhibited a lower density of focal adhesions, which corresponds to a reduced organizational state of the cytoskeleton. These results indicate that regulation of GJIC most strongly correlates with altered focal adhesion and cytoskeleton organization that can lead to various secondary responses, including motility, growth, and transformation, and suggest that RHAMM regulates GJIC and Cx43 expression possibly through its actions on focal adhesions and the associated cytoskeleton.