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Intracellular Movements and Reorganization of Electrically Fused Sunflower Protoplasts

Early events during the fusion process of sunflower protoplasts have been investigated using videoenhanced microscopy and fine structure analysis. Electrical pulses applied for membrane breakdown induced an immediate cessation of cytoplasmic streaming, which resumed 0.5-5 min after pulsing. This observation may provide a marker for fusion product viability and an indicator to optimize fusion parameters. Correlated with the cessation of cytoplasmic streaming, internal vesicles formed probably from the ER. Two types of fusion events were observed: (1) protoplasts interconnected by cytoplasmic bridges showing no rounding up and subsequent individual cell divisions and (2) protoplasts connected by a broad fusion plane rounding up in ca. 1 h. This process was accompanied by accumulation of cytoplasm in the fusion plane, formation of cytoplasmic strands connecting the fused protoplasts, and dislocation of the nuclei. The creation of viable fusion products required not only membrane fusion but also complete reorganization of the cytoplasmic material.