Calcium ions as efficient cofactor of polycation-mediated gene transfer

• Published in: Biochimica et biophysica acta Vol. 1445; no. 1; pp. 21 - 30
• Main Authors: Haberland, Annekathrin, Knaus, Thomas, Zaitsev, Sergei V., Stahn, Renate, Mistry, Ajay R., Coutelle, Charles, Haller, Hermann, Böttger, Michael
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 14-04-1999
• Subjects: Calcimycin - pharmacology; Calcium - pharmacology; Calcium ion; Calcium Phosphates - pharmacology; Calcium precipitate; Cations, Divalent - pharmacology; Cell Line; Commercial transfectant; DNA - chemistry; H1 histone; Histones; Humans; Nifedipine - pharmacology; Nuclear protein; Polyamines; Time Factors; Transfection; Transfection - methods; Calcium precipitate; Transfection; Calcium ion; H1 histone; Commercial transfectant; Nuclear protein
• ISSN: 0167-4781; 0006-3002; 1879-2634
• DOI: 10.1016/S0167-4781(99)00017-2

We investigated the effect of calcium on the transfection of non-viral DNA transfer systems. Cationic proteins such as the nuclear protein H1, the polycation polylysine and a number of commercial transfection agents exhibited high transfection rates in the presence of Ca
2+. Without Ca
2+ H1 and HMG1 were inactive in transfection of the human permanent endothelial cell line ECV 304 while cationic liposomes such as Lipofectin and Lipofectamine did not show any Ca
2+ dependence. More detailed experiments showed that Ca
2+ was replaceable by the lysosomotropic agent chloroquine. Furthermore, it was possible to separate the transfection-enhancing role of Ca
2+ from the actual transfection process by adding Ca
2+ to the cells after the transfection period and still to obtain a significant transgene expression. This makes it possible to distinguish between cellular uptake of H1 (or mediator)-DNA complexes and endocytotic release. We also replaced soluble Ca
2+ by Ca-phosphate precipitates not containing DNA and obtained similar transfection results. This allowed us to suggest that the addition of free Ca
2+ to the transfection medium resulted in nascent Ca-phosphate microprecipitates. The known fusogenic and membranolytic activity of such microprecipitates could facilitate the transport through and the release of the transfecting complexes from the endosomal/lysosomal compartment.