Mechanism of restriction of normal and cystic fibrosis transmembrane conductance regulator-deficient human tracheal gland cells to adenovirus infection and ad-mediated gene transfer.

F. Gaden (1) , L. Franqueville (1) , Ss Hong (1) , V. Legrand (2) , C. Figarella (3) , P. Boulanger (1)

1 VPV - Virologie et pathogenèse virale
2 Transgene SA [Illkirch]
3 Groupe de Recherche sur les Glandes Endocrines

CF-KM4 (cystic fibrosis transmebrane conductance regulator-deficient) and MM-39 (healthy) cells, two serous cell lines from submucosal tracheal glands, were found to be poorly susceptible to adenovirus (Ad)5 infection and Ad5-mediated gene transduction. The major limiting steps apparently resided in the primary events of Ad5 interaction, i.e., cell attachment and entry. Both CF-KM4 and MM-39 cells failed to express the Coxsackie-Ad receptor (CAR), and experimental data suggested that alpha[2-->6]-linked sialic acid residues of sialoglycoproteins (SAGP) in CF-KM4 cells, and heparan sulfate glycosaminoglycans (HS-GAG) in MM-39, were used as receptors by Ad5 virions. Ad5 attached to SAGP and HS-GAG receptors via its fiber knob domain, but entered the cells via a penton base- and Arg-Gly-Asp (RGD)-integrin-independent pathway. The block to Ad5-mediated gene transfer in MM-39 and KM4 cells could be overcome by conferring to the vector a novel cell-binding specificity. Thus, Ad5 vectors carrying a stretch of 7-lysine residues genetically inserted at the C-terminus of the fiber knob were found to transduce MM-39 cells with a 10- to 20-fold higher efficiency than the original vectors, but the transduction of CF-KM4 was not significantly improved. Retargeting Ad5 to integrin receptors via RGD peptide ligands, inserted at the extremity of the fiber shaft, resulted in a transducing efficiency of 20- and 50-fold higher in MM-39 and KM4 cells, respectively, compared with Ad5 vectors carrying fibers terminated by their natural knob domain.

Mots clés: Adenoviridae/*genetics/metabolism; Adenoviridae Infections/genetics; Bronchi/cytology/virology; Carbohydrate Conformation; Cell Adhesion/genetics; Cells; Cultured; Cystic Fibrosis/genetics/pathology; Cystic Fibrosis Transmembrane Conductance Regulator/*genetics; Endocytosis; Gene Transfer Techniques; Heparan Sulfate Proteoglycan/metabolism; Humans; Microscopy; Electron; Oligopeptides/metabolism; Receptors; Cell Surface/metabolism; Virus/genetics/metabolism; Research Support; Non-U.S. Gov't; Sialoglycoproteins/chemistry/metabolism; Trachea/*pathology/physiopathology/virology; Virion/metabolism

Domaines: Virologie

F. Gaden, L. Franqueville, Ss Hong, V. Legrand, C. Figarella, et al.. Mechanism of restriction of normal and cystic fibrosis transmembrane conductance regulator-deficient human tracheal gland cells to adenovirus infection and ad-mediated gene transfer.. 2006. ⟨hal-00117220⟩