Herpes simplex virus (HSV) causes lifelong latent infections in humans. It is responsible for significant disease, ranging from cold sores and genital infections to blindness and fatal encephalitis. There is no effective vaccine and no cure. The long-term objective of this project is a detailed understanding of the intracellular pathway taken by HSV entering during infection. The endocytosis pathway utilized by HSV during entry into human epithelial cells was identified 20 years ago. However, the specific vesicular compartments traversed by HSV during its journey to the nucleus are not known. We demonstrated that HSV does not travel the conventional lysosome-terminal endocytosis pathway during viral entry. Based on our previous results and new preliminary studies, we hypothesize that incoming HSV utilizes a retrograde transport pathway to the Golgi apparatus for successful entry into physiologically relevant cells. Completion of this pilot study will demonstrate that HSV transits the Golgi complex prior to arrival at the nucleus during successful entry. Our experimental design employs techniques of cell biology, biochemistry, microscopy, and molecular virology. Completing this study will establish critical features of the entry pathway taken by HSV in epithelial cells, the in vivo target of primary and recurrent infection. These studies will support the innovative concept that the Golgi complex and Golgi-related functions mediate HSV entry and will lay the foundation for identification of the intracellular site of HSV membrane fusion. Furthermore, the results will aid in the development of new preventions and therapeutics for HSV.