Lactosylated poly-L-lysine targets a potential lactose receptor in cystic fibrosis and non-cystic fibrosis airway epithelial cells.
Poly-L-lysine with 40% of the epsilon -amino groups substituted with lactosyl residues facilitated the internalization of lactosylated poly-L-lysine/cDNA complexes into cystic fibrosis (CF) and non-CF airway epithelial cells. It was previously shown that lactosylated poly-L-lysine enhanced the transfer of cDNA into the cell nucleus, resulting in transfection. The cell entry of lactosylated poly-L-lysine/cDNA complexes, however, has not been elucidated and we hypothesized that entry of the complex was by receptor-mediated endocytosis. It is shown here that binding of the vector/cDNA complexes to the cell membrane was inhibited by lactose but not N-acetyl glucosamine. Examination by electron microscopy revealed the complexes in clathrin-coated pits. Furthermore, the complexes colocalized with transferrin during cell entry and were shown in early endosomes. These results demonstrated that lactosylated poly-L-lysine/cDNA complexes enter airway epithelial cells via receptor-mediated endocytosis utilizing lactose-binding receptors, which employ the clathrin-coated pit for internalization. Taken together with the fact that nuclear translocation also is enhanced by lactose, these results demonstrate why lactosylated poly-L-lysine is an excellent vector for transfection of airway epithelial cells. Moreover, other carbohydrates covalently linked to poly-L-lysine for targeting other specific cell types, combined with lactosyl residues, can be designed for the development of other molecular conjugates for gene transfer.
Klink
Daniel
D
The Cystic Fibrosis Center and Department of Pediatrics, University of Pennsylvania School of Medicine, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania 19104, USA.
Yu
Qian-Chun
QC
Glick
Mary Catherine
MC
Scanlin
Thomas
T
D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism; D003034:Coated Pits, Cell-Membrane / Q000378:metabolism; D003550:Cystic Fibrosis / Q000378:metabolism*; D018076:DNA, Complementary; D004705:Endocytosis; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D006801:Humans; D007785:Lactose / Q000378:metabolism*; D008854:Microscopy, Electron; D011107:Polylysine / Q000378:metabolism*; D014132:Trachea / Q000166:cytology / Q000378:metabolism / Q000473:pathology* / Q000648:ultrastructure
D002966:Clathrin; D018076:DNA, Complementary; D011107:Polylysine; D007785:Lactose