ELECTRIC-FIELD MEDIATED LOADING OF MACROMOLECULES IN INTACT YEAST-CELLS IS CRITICALLY CONTROLLED AT THE WALL LEVEL

The mechanism of electric field mediated macromolecule transfer inside an intact yeast cell was investigated by observing, under a microscop e, the fluorescence associated to cells after pulsation in a buffer co ntaining two different hydrophilic fluorescent dyes. In the case of a small probe such as propidium iodide, a long lived permeabilized state was induced by the field as classically observed on wall free systems . Penetration of a 70 kDa FITC dextran was obtained only by using dras tic conditions and only a very limited number of yeast cells which too k up macromolecules remained viable. Most dextrans were trapped in the wall. A dramatic improvement in transfer of dextrans was observed whe n the cells were treated by dithiothreitol before pulsation. A cytopla smic protein leakage was detected after the electric treatment suggest ing that an irreversible damage took place in the walls of many pulsed cells. Electroloading of macromolecules in intact yeast cells appears to be controlled by a field induced short lived alteration of the env elope organization.