Investigating the Function of Vaccinia Virus Protein A33 during Morphogenesis, Egress, and Infectivity

Thesis (Ph.D.)--University of Rochester. School of Medicine & Dentistry. Dept. of Microbiology and Immunology, 2011.
Vaccinia virus, the prototypical member of the orthopoxviruses, was used as a live attenuated vaccine for the eradication of smallpox. It is an enveloped DNA virus with a large genome of about 200 kbp that is predicated to encode approximately 200 functional open reading frames. During vaccinia virus infection, two morphologically and antigenically distinct forms of infectious virions are produced: intracellular mature virions (IMV) and extracellular virions (EV). The first form of infectious virions produced within the viral factory is IMV. Due to the intracellular envelopment that occurs at the trans-Golgi network or early endosome, a small subset of IMV obtain an additional double membrane envelope and become intracellular enveloped virions (IEV). IEV are transported to the cell surface along microtubules and released from the cytoplasm by fusion of their outermost membrane with the plasma membrane. Virions that are retained on the cell surface are called cell-associated enveloped virion (CEV) and those released from the cell surface are called extracellular enveloped virion (EEV). CEV and EEV are collectively termed EV, which are responsible for severe systemic infection. Although eight proteins encoded by the virus, A33, A34, A36, A56, B5, F12, F13, and K2, are known to be specific for IEV/EV, understanding of the molecular mechanism of intracellular envelopment is far from complete. The major goal of this thesis study is to elucidate the function of A33 during viral morphogenesis, egress, and/or infectivity. A recombinant virus that has A33R deleted (v∆A33R) has been shown to produce small plaques due to the inability of the virus to induce actin tails. Thus, A33 is known to be required for efficient cell-to-cell spread. However, its exact role during infection remains to be determined. To study poxvirus morphogenesis in the absence of A33, a recombinant virus that has A33R deleted and expresses B5R-GFP in place of the normal B5R (vB5R-GFP/∆A33R) was generated