Effect of cytochalasins on F-actin and morphology of Ehrlich ascites tumorcells

Cytochalasins have been used extensively to probe the role of F-actin in di fferent aspects of cellular function. Most of the data obtained are interpr eted on the basis of the well-established depolymerizing effects of cytocha lasins on F-actin preparations in vitro. However, some evidence indicates t hat, in intact cells, different cytochalasins can have varying effects on c ell morphology and F-actin content and organization. To examine this proble m in more detail, we analyzed the effects of cytochalasins on the cell morp hology of and F-actin content and organization in Ehrlich ascites tumor (EA T) cells. After a 3-min exposure to 0.5 muM cytochalasin D, B, or E, F-acti n content was equally reduced in all cases and this correlated with a reduc tion in the amount of cortical F-actin associated with the EAT cell membran e. However, only with CE was cell morphology markedly altered, with the app earance of numerous blebs. At 10 muM, blebbing was present in all condition s and the organization of cortical F-actin was disrupted. F-actin content, however, was not further reduced by this higher concentration and in CD it was identical to control levels. Exposure of EAT cells to similar concentra tions of cheatoglobosin C, an analog of the cytochalasins that has little t o no affinity for F-actin, resulted in a loss of F-actin content, a reducti on in F-actin fluorescence, but no change in cell morphology, including a c omplete lack of bleb formation. Myosin II immunoreactivity, concentrated in the cortical cytoplasm colocalized with F-actin and in an area associated with the Golgi, was reduced by the high-dose cytochalasin. These results de monstrate that caution must be exercised in the use of cytochalasins to pro be the role of F-actin in cellular function and that several parameters mus t be analyzed to obtain an accurate assessment of the effect of cytochalasi n on the actin filament system. (C) 2000 Academic Press.