Receptor-mediated endocytosis and nuclear transport of a transfecting DNA construct

Andrey A. Rosenkranz, Sergey V. Yachmenev, David A. Jans, Natalya V. Serebryakova, Vitaly I. Murav'ev, Reiner Peters, Alexander S. Sobolev

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Abstract

A soluble construct consisting of a plasmid carrying the gene of the SV40 large T-antigen and an insulin-poly-l-lysine conjugate is able to selectively transfect PLC/PRF/5 human hepatoma cells which possess insulin receptors. Transfection can be efficiently competed by excess free insulin. To examine intracellular transport of the construct, it was fluorescently labeled and its accumulation on and in cells visualized by video-enhanced microscopy and quantitative confocal laser scanning microscopy. After 2 h at 37 °C, the labeled construct was found predominantly in intracellular acidic compartments, with a substantial portion of fluorescence localized both near and in the cell nucleus. Binding, endocytosis, and nuclear localization of the labeled conjugate could all be competed by excess free insulin, thus indicating that entry of the conjugate into cells was specifically mediated by the insulin receptor.

Original languageEnglish
Pages (from-to)323-329
Number of pages7
JournalExperimental Cell Research
Volume199
Issue number2
DOIs
Publication statusPublished - 1 Jan 1992
Externally publishedYes

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