Cell Surface Nucleolin Serves as Receptor for DNA Nanoparticles Composed of Pegylated Polylysine and DNA

• Published in: Molecular therapy Vol. 16; no. 2; pp. 333 - 342
• Main Authors: Chen, Xuguang, Kube, Dianne M, Cooper, Mark J, Davis, Pamela B
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-02-2008; Elsevier Limited
• Subjects: Antibodies; Cell Membrane - metabolism; Cell Nucleolus - metabolism; Cells; Cells, Cultured; Cytoplasm; Cytoplasm - metabolism; DNA - chemistry; DNA - genetics; Drug dosages; Epithelial Cells - cytology; Epithelial Cells - metabolism; Gene therapy; Gene Transfer Techniques; HeLa Cells; Humans; Hypotheses; Microscopy, Fluorescence; Nanoparticles; Nanoparticles - chemistry; Nucleolin; Pediatrics; Peptides; Phosphoproteins - genetics; Phosphoproteins - metabolism; Polyethylene glycol; Polylysine - chemistry; Protein Binding; Proteins; Rhodamines - chemistry; RNA, Small Interfering - genetics; RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism; Transfection; Viruses; United States > US; Ohio
• ISSN: 1525-0016; 1525-0024
• DOI: 10.1038/sj.mt.6300365

Compacted DNA nanoparticles deliver transgenes efficiently to the lung following intrapulmonary dosing. Here we show that nucleolin, a protein known to shuttle between the nucleus, cytoplasm, and cell surface, is a receptor for DNA nanoparticles at the cell surface. By using surface plasmon resonance (SPR), we demonstrate that nucleolin binds to DNA nanoparticles directly. The presence of nucleolin on the surface of HeLa and 16HBEo- cells was confirmed by surface biotinylation assay and immunofluorescence. Rhodamine-labeled DNA nanoparticles colocalize with nucleolin on the cell surface, as well as in the cytoplasm and nucleus, but not with transferrin or markers of early endosome or lysosome following cellular uptake. Reducing nucleolin on the cell surface by serum-free medium or siRNA against nucleolin treatment leads to significant reduction in luciferase reporter gene activity, while overexpressing nucleolin has the opposite effect. Competition for binding to DNA nanoparticles with exogenous purified nucleolin decreases the transfection efficiency by 60–90% in a dose-dependent manner. Therefore, the data strongly suggest that cell surface nucleolin serves as a receptor for DNA nanoparticles, and that nucleolin is essential for internalization and/or transport of the nanoparticles from cell surface to the nucleus.