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Journal of Virology, September 2005, p. 11142-11150, Vol. 79, No. 17
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.17.11142-11150.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Genetic and Molecular In Vivo Analysis of Herpes Simplex Virus Assembly in Murine Visual System Neurons

Departments of Anatomy and Ophthalmology, University of California San Francisco, San Francisco, California 94143-0452,¹ Program in Virology and Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115²

Received 7 April 2005/ Accepted 15 June 2005

Herpes simplex virus (HSV) infects both epithelial cells and neuronal cells of the human host. Although HSV assembly has been studied extensively for cultured epithelial and neuronal cells, cultured neurons are biochemically, physiologically, and anatomically significantly different than mature neurons in vivo. Therefore, it is imperative that viral maturation and assembly be studied in vivo. To study viral assembly in vivo, we inoculated wild-type and replication-defective viruses into the posterior chamber of mouse eyes and followed infection in retinal ganglion cell bodies and axons. We used PCR techniques to detect viral DNA and RNA and electron microscopy immunohistochemistry and Western blotting to detect viral proteins in specific portions of the optic tract. This approach has shown that viral DNA replication is necessary for viral DNA movement into axons. Movement of viral DNA along ganglion cell axons occurs within capsid-like structures at the speed of fast axonal transport. These studies show that the combined use of intravitreal injections of replication-defective viruses and molecular probes allows the genetic analysis of essential viral replication and maturation processes in neurons in vivo. The studies also provide novel direct evidence for the axonal transport of viral DNA and support for the subassembly hypothesis of viral maturation in situ.

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