Directing Integrin-linked Endocytosis of Recombinant AAV Enhances Productive FAK-dependent Transduction

• Published in: Molecular therapy Vol. 20; no. 5; pp. 972 - 983
• Main Authors: Kaminsky, Paul M, Keiser, Nicholas W, Yan, Ziying, Lei-Butters, Diana CM, Engelhardt, John F
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.05.2012; Elsevier Limited; Nature Publishing Group
• Subjects: Adeno-associated virus; Adhesion; Animals; Binding sites; Cells, Cultured; Dependovirus - genetics; Efficiency; Endocytosis - drug effects; Fibroblasts - cytology; Fibroblasts - drug effects; Fibroblasts - metabolism; Focal Adhesion Protein-Tyrosine Kinases - genetics; Focal Adhesion Protein-Tyrosine Kinases - metabolism; Gene therapy; Gene Transfer Techniques; Genes, Reporter; Genetic Vectors; Heparan sulfate; Humans; Infections; Kinases; Ligands; Luciferases; Manganese - pharmacology; Medicine; Mice; Original; Peptides; Peptides - genetics; Peptides - metabolism; Proteins; Receptors, Virus - genetics; Receptors, Virus - metabolism; Signal Transduction - genetics; Vinculin - genetics; Vinculin - metabolism; United States > US; Iowa; Iowa City Iowa
• ISSN: 1525-0016; 1525-0024
• DOI: 10.1038/mt.2011.295

Recombinant adeno-associated virus (rAAV) is a widely used gene therapy vector. Although a wide range of rAAV serotypes can effectively enter most cell types, their transduction efficiencies (i.e., transgene expression) can vary widely depending on the target cell type. Integrins play important roles as coreceptors for rAAV infection, however, it remains unclear how integrin-dependent and -independent mechanisms of rAAV endocytosis influence the efficiency of intracellular virus processing and ultimately transgene expression. In this study, we examined the contribution of integrin-mediated endocytosis to transduction of fibroblasts by rAAV2. Mn++-induced integrin activation significantly enhanced (∼17-fold) the efficiency of rAAV2 transduction, without altering viral binding or endocytosis. rAAV2 subcellular localization studies demonstrated that Mn++ promotes increased clustering of rAAV2 on integrins and recruitment of intracellular vinculin (an integrin effector) to sites of rAAV2 binding at the cell surface. Focal adhesion kinase (FAK), a downstream effector of integrin signals, was essential for rAAV2/integrin complex internalization and transduction. These findings support a model whereby integrin activation at the cell surface can redirect rAAV2 toward a FAK-dependent entry pathway that is more productive for cellular transduction. This pathway appears to be conserved for other rAAV serotypes that contain a capsid integrin-binding domain (AAV1 and AAV6).