An immunofluorescence study of the calcium-induced coordinated reorganization of microfilaments, keratin intermediate filaments, and microtubules in cultured human epidermal keratinocytes

Indirect immunofluorescence microscopy has been used to investigate the coordinated reorganization of microtubules, microfilaments, and keratin intermediate filaments in cultured human epidermal keratinocytes following a switch from low-Ca⁺⁺ (0.15 mM) medium to high-Ca⁺⁺ (1.05 mM) medium. A dramatic reorganization occurs concurrently in the three major cytoskeletal components shortly after the calcium switch. The most prominent features are the alignment of keratin filaments at the plasma membranes of apposed cells, the induction of microfilament rings, the restriction of microtubules to the area within the boundaries of the microfilament rings, and the alignment of actin bundles at cell borders. Additional changes are observed in terminally differentiated cells. This is the first report that describes simultaneous changes in the organization of the three major cytoskeletal components of epidermal keratinocytes. Cytochalasin D and demecolcine (colcemid) studies were performed to determine whether the organization of microtubules, microfilaments, and keratin filaments, as well as the calcium-induced reorganization of these cytoskeletal elements, may be dependent on the existence of structural relationships between them. These studies demonstrate that the disruption of microfilaments results in the formation of a latticelike keratin network, with a close association of actin and keratin being maintained. The formation of keratin filament alignments occurs even in the absence of intact microfilaments. In addition, it was found that the Ca⁺⁺-induced reorganization of microfilaments and keratin filaments is not dependent on an intact microtubule network. Furthermore, the reorganization of actin into concentric rings can be dissociated from changes in the organization of keratin filaments.