Endocytosis and nuclear trafficking of adeno-associated virus type 2 are controlled by Rac1 and phosphatidylinositol-3 kinase activation

Adeno-associated virus (AAV) is a single-stranded DNA parvovirus that cause s no currently known pathology in humans. Despite the fact that this virus is of increasing interest to molecular medicine as a vector for gene delive ry, relatively little is known about the cellular mechanisms controlling in fection. In this study, we have examined endocytic and intracellular traffi cking of AAV-2 using fluorescent (Cy3)-conjugated viral particles and molec ular techniques. Our results demonstrate that internalization of heparan su lfate proteoglycan-bound AAV-2 requires alpha V beta 5 integrin and activat ion of the small GTP-binding protein Rac1. Following endocytosis, activatio n of a phosphatidylinositol-3 (PI3) kinase pathway was necessary to initiat e intracellular movement of AAV-2 to the nucleus via both microfilaments an d microtubules. Inhibition of Rac1 using a dominant N17Rac1 mutant led to a decrease in AAV-2-mediated PI3 kinase activation, indicating that Rac1 may act proximal to PI3 kinase during AAV-2 infection. In summary, our results indicate that alpha V beta 5 integrin-mediated endocytosis of AAV-2 occurs through a Rad and PD kinase activation cascade, which directs viral moveme nt along the cytoskeletal network to the nucleus.