Expression and Functional Significance of the Ca~(2+)-Activated Cl- Channel ANO6 in Dendritic Cells

摘要

Background/Aims: Migration of dendritic cells (DCs), antigen presenting cells that link innate and adaptive immunity, is critical for initiation of immune responses. DC migration is controlled by the activity of different ion channels, which mediate Ca~(2+) flux or set the membrane potential. Moreover, cell migration requires local volume changes at the leading and rear end of travelling cells, which might be mediated by the fluxes of osmotically active solutes, including Cl-. The present study explored the functional expression, regulation and role of Cl- channels in mouse bone marrow-derived DCs. Methods/Results: In whole-cell patch clamp experiments we detected outwardly rectifying Cl- currents which were activated by elevation of cytosolic Ca~(2+), triggered either by ionomycin in the presence of extracellular Ca~(2+) or mobilization of Ca~(2+) by IP3. Most importantly, Ca~(2+)-activated Cl- channels (CaCCs) were activated by CCL21 (75 ng/ml), an agonist of the chemokine receptor CCR7. The currents showed sensitivity to Cl- channel blockers such as tannic acid (10 μM), digallic acid (100 μM) and more specific CaCC blockers niflumic acid (300 μM) and AO1 (20 μM). According to RT-PCR and Western blot data, Anoctamin 6 (ANO6) is expressed in DCs. Knock-down of ANO6 with siRNA led to inhibition of CaCC currents in DCs. Moreover, chemokine-induced migration of both immature and LPS-matured DCs was reduced upon ANO6 knock-down. Conclusion: Our data identify ANO6 as a Ca~(2+)-activated Cl- channel in mouse DCs, show its activation upon chemokine receptor ligation and establish an important role of ANO6 in chemokine-induced DC migration.