The Calcium/Calcineurin Pathway Promotes Hemidesmosome Stability through Inhibition of β4 Integrin Phosphorylation

Cell migration depends on cells being able to create and disassemble adhesive contacts. Hemidesmosomes are multiprotein structures that attach epithelia to basal lamina and disassemble during migration and carcinoma invasion. Phosphorylation of the beta 4 integrin, a hemidesmosome component, induces disassembly. Although kinases involved in beta 4 phosphorylation have been identified, little is known about phosphatases countering kinase action. Here we report that calcineurin, a serine-threonine protein phosphatase, regulates beta 4 phosphorylation. Calcineurin inhibitor cyclosporin A (CsA) and calcineurin-siRNA increase beta 4 phosphorylation, induce hemidesmosome disassembly, and increase migration in HaCat keratinocytes, suggesting that calcineurin negatively regulates beta 4 phosphorylation. We found no direct dephosphorylation of beta 4 by calcineurin or association between beta 4 and calcineurin, suggesting indirect regulation of beta 4 phosphorylation. We therefore assessed calcineurin influence on MAPK and PKC, known to phosphorylate beta 4. CsA increased MAPK activity, whereas MAPK inhibitors reduced CsA-induced beta 4 phosphorylation, suggesting that calcineurin restricts beta 4 phosphorylation by MAPK. Calcineurin is activated by calcium. Increased [Ca2+](i) reduces beta 4 phosphorylation and stabilizes hemidesmosomes, effects that are reversed by CsA, indicating that calcineurin mediates calcium effects on beta 4. However, MAPK activation is increased when [Ca2+](i) is increased, suggesting that calcineurin activates an additional mechanism that counteracts MAPK-induced beta 4 phosphorylation. Interestingly, in some squamous cell carcinoma cells, which have reduced hemidesmosomes and increased beta 4 phosphorylation, an increase in [Ca2+](i) using thapsigargin, bradykinin, or acetylcholine can increase hemidesmosomes and reduce beta 4 phosphorylation in a calcineurin-dependent manner. These findings have implications in calcineurin-inhibitor induced carcinoma, a complication of immunosuppressive therapy.