Effects of low temperature and calcium on microfilament structure in flagellates of Physarum polycephalum

Microfilament structures of the ridge and the backbone in Physarum flagellates disintegrated selectively within a few minutes upon cooling by ice-water. The elongated cells concurrently rounded up to spherical or irregular and amoeboid shape. When such rounded cells were warmed to 25 °C, the microfilament structures were reconstructed within 1 min and cells subsequently returned to an elongated shape. Disruption of microfilaments by cytochalasin A also caused the rounding up of cells, indicating that the rounding up resulted from disintegration of microfilament structures. This transformation induced by the cold treatment was retarded by preincubation of the cells with EGTA for 15 min, but addition of EGTA immediately before the onset of the cold treatment was less effective. The effect of EGTA was cancelled by simultaneous addition of excess Ca²⁺. Addition of procaine also inhibited the transformation induced by the cold treatment, while caffeine inhibited the recovery of the elongated shape when returned to 25 °C. Furthermore, addition of A23187 at 25 °C in the presence of Ca²⁺ mimicked the effect of the cold treatment. Thus, intracellular release of Ca²⁺ was suggested to be involved in the transformation induced by the cold treatment. Lability of the microfilament structures at a high concentration of Ca²⁺ was directly proved using Triton-permeabilized cells. Therefore we concluded that low temperature disrupts microfilament structures that are necessary for the maintenance of the elongated cell shape by inducing intracellular Ca²⁺ release. However, microfilament structures in Physarum amoeba cells were affected neither by the cold treatment nor by high Ca²⁺ concentration.