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MBC in Press, published online ahead of print January 18, 2002
Mol. Biol. Cell 10.1091/mbc.01-06-0280

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Submitted on June 4, 2001
Revised on September 24, 2001
Accepted on November 1, 2001

Macromolecular uptake is a spontaneous event during mitosis in cultured fibroblasts: implications for vector-dependent plasmid transfection

Anne Fernandez1*, Pierre Pellegrin1, Ned J.C. Lamb1, and René Bennes1

1 Institut de Génétique Humaine, CNRS UPR 1142, 141 rue de la Cardonille, 34396 Montpellier, France

* Corresponding author. E-mail address: Anne.Fernandez{at}igh.cnrs.fr.

The process through which macromolecules penetrate the plasma membrane of mammalian cells remains poorly defined. We have examined if natural cellular events modulate the capacity of cells to take up agents applied extraneously. Here we report that during mitosis and in a cell type-independent manner, cells exhibit a natural ability to absorb agents present in the extracellular environment up to 150 kDa as assessed using FITC-dextrans. This event is exclusive to the mitotic period and not observed during G0, G1, S or G2 phase. During mitosis, starting in advanced prophase, oligonucleotides, active enzymes and polypeptides are efficiently taken into mitotic cells. This uptake of macromolecules during mitosis still takes place in the presence of cytochalasin D or nocodazole, showing no requirement for intact microtubules or actin filaments in this process. However, cell rounding up, which still takes place in the presence of either of these drugs in mitotic cells, appears to be a key event in this process. Indeed, limited trypsinization of adherent cells mimics both the cell retraction and macromolecule uptake observed as cells enter mitosis. A plasmid DNA encoding GFP (3.3Mda) coated with a 18 aa peptide is efficiently expressed when applied onto synchronized G2/M fibroblasts whereas little or no expression is observed when the coated plasmid is applied onto asynchronous cell cultures. This shows that such coating peptides are only efficient for their encapsulating and protective effect on the plasmid DNA to be "vectorised" rather than acting as true vectors.




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