NUCLEAR ION-CHANNEL ACTIVITY IS REGULATED BY ACTIN-FILAMENTS

Actin filaments are novel second messengers involved in ion channel re gulation. Because cytoskeletal components interact with the nuclear en velope, the actin cytoskeleton may also control nuclear membrane funct ion. In this report, the patch-clamp technique was applied to isolated nuclei from amphibian A6 epithelial cells to assess the role of actin filaments on nuclear ion channel activity under nucleus-attached or - excised conditions. The most prevalent spontaneous nuclear ion channel species, 76% (n = 46), was cation selective and had a maximal single- channel conductance of similar to 420 pS. Nuclear ion channels also di splayed multiple subconductance states, including channel activity of 26 pS that was frequently observed. Nuclear ion channel activity on ot herwise quiescent patches was induced by either addition of the actin cytoskeleton disrupter cytochalasin D (CD; 5 mu g/ml, 60%, 3 of 5 patc hes) or actin (10-1,000 mu g/ml) to the bathing solution of nucleus-at tached patches (59%, 13 of 22 patches). Actin also induced ion channel activity in quiescent excised inside-out patches from the nuclear env elope (80%, 4 of 5 patches). In contrast, addition of bovine serum alb umin (10-1,000 mu g/ml) to the bathing solution of nucleus-attached pa tches was without effect on nuclear ion channel activity (5 of 5 patch es). The monoclonal antibody MAb414, specific for nuclear pore complex proteins, completely prevented either spontaneous or cytosolic actin- induced nuclear ion channels under nucleus-attached conditions (4 of 4 patches) but not intranuclear actin-induced nuclear ion channels unde r excised inside-out conditions (3 of 3 patches). In nucleus-attached patches, channel activity was readily activated by addition of the G-a ctin-binding protein deoxyribonuclease I to nucleus-attached patches ( 56%, 5 of 9 patches) or further addition of the actin-cross-linker fil amin in the presence of actin (57%, 4 of 7 patches). The data indicate that dynamic changes in actin filament organization may represent a n ovel mechanism to control nuclear function.