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Journal of Virology, October 2000, p. 9184-9196, Vol. 74, No. 19
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Endocytosis and Nuclear Trafficking of Adeno-Associated Virus Type 2 Are Controlled by Rac1 and Phosphatidylinositol-3 Kinase Activation

Salih Sanlioglu,1,2 Peter K. Benson,1,3 Jusan Yang,1 E. Morrey Atkinson,4 Thomas Reynolds,4 and John F. Engelhardt1,2,*

Department of Anatomy and Cell Biology and Center for Gene Therapy,1 Department of Internal Medicine,2 and Department of Otolaryngology-Head and Neck Surgery Institute,3 University of Iowa College of Medicine, Iowa City, Iowa 52242, and Targeted Genetics Corporation, Seattle, Washington 981014

Received 12 May 2000/Accepted 14 July 2000

Adeno-associated virus (AAV) is a single-stranded DNA parvovirus that causes no currently known pathology in humans. Despite the fact that this virus is of increasing interest to molecular medicine as a vector for gene delivery, relatively little is known about the cellular mechanisms controlling infection. In this study, we have examined endocytic and intracellular trafficking of AAV-2 using fluorescent (Cy3)-conjugated viral particles and molecular techniques. Our results demonstrate that internalization of heparan sulfate proteoglycan-bound AAV-2 requires alpha Vbeta 5 integrin and activation of the small GTP-binding protein Rac1. Following endocytosis, activation of a phosphatidylinositol-3 (PI3) kinase pathway was necessary to initiate intracellular movement of AAV-2 to the nucleus via both microfilaments and 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 Vbeta 5 integrin-mediated endocytosis of AAV-2 occurs through a Rac1 and PI3 kinase activation cascade, which directs viral movement along the cytoskeletal network to the nucleus.

* Corresponding author. Mailing address: Department of Anatomy and Cell Biology, University of Iowa, College of Medicine, 51 Newton Rd., Room 1-111 BSB, Iowa City, IA 52242. Phone: (319) 335-7753. Fax: (319) 335-7198. E-mail: john-engelhardt{at}uiowa.edu.

Journal of Virology, October 2000, p. 9184-9196, Vol. 74, No. 19
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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