Herpesvirus Research

The overall goal of my research is to understand herpes simplex virus (HSV) entry into susceptible cells animal models of HSV-1 and HSV-2 disease in their human host. HSV entry requires binding of gD to either HVEM or nectin-1. This interaction triggers virus-cell fusion involving three other HSV glycoproteins. HVEM is a member of the TNF receptor superfamily. Nectin-1 is an adhesion molecule located at adherens junctions of cells and is a member of the Ig superfamily. A major accomplishment was to solve the three-dimensional structure of glycoprotein D bound to HVEM. The structure of this complex has enabled us to carry out structure-based mutagenesis of both proteins. Three additional gD structures have enabled us to understand how the gD/receptor interaction triggers later steps of the entry process. Our studies have shown that the binding sites for both receptors are hidden in the native protein and that a major conformational changed in gD accompanies receptor binding. Our structural studies are ongoing with very active collaborations to solve the structures of the other entry glycoproteins. We are close to knowing the 3-dimensional structure of gB, a highly conserved protein that is a critical component of the fusion machinery. Studies to understand the structure and funtion of the other component of the fusion complex, called gH/gL is also an active area our research. In more biologic studies, we and others have also found that virus entry can occur by at least three different mechanisms: direct fusion withthe plasma membrane, pH dependent endocytosis and pH independent but receptor dependent endocytosis. The "rules" that govern which pathway is used and in which cell type is an active area of investigation. Our current direction is to understand how the interaction between gD and its receptors triggers the next steps of virus entry and cell fusion. Finally, we are examining entry using real-time imaging with viruses and/or receptors that bear fluorescent tags. We found that gD causes a major down-regulation of its receptors both on the cell that the virus infects and on neighboring uninfected cells. This fascinating property has led us into very interesting aspects of cell biology.

Poxvirus Research

We recently began work on vaccinia virus (VV) glycoproteins. One goal is to develop an antibody cocktail to replace vaccinia immune globulin (VIG) as a therapy for adverse reactions to the live vaccine. A second goal is to develop a subunit vaccine against smallpox using proteins from vaccinia virus or from variola virus (smallpox). We have cloned several VV envelope proteins in a baculovirus expression system, purified the recombinant proteins and developed polyclonal and monoclonal antibodies. The monoclonal antibodies are being evaluated in passive immunization studies. The proteins are being evaluated for efficacy as vaccines in mouse and monkey models. Thus far, we have found that a combination of antibodies to various glycoproteins is effective as a passive immune therapeutic. Importantly, we have also shown that a combination of three envelope proteins of VV protects mice against a lethal VV challenge. Preliminary studies indicate that this combination is also effective in protectin monkeys against a monkeypox challenge. A third goal is to understand the role of each these proteins (and others) in poxvirus entry. A variety of experimental approaches are being taken, modeled on our herpesvirus work.

Post-doctoral fellows, Research Associates and Research Ass't Professors (2005)
Claude Krummenacher, PhD
Richard S.B. Milne, PhD
Tina  Cairns, PhD
Florent Bender, PhD
Lyria Aldaz , PhD

Graduate Students (all in CAMB)
Brian Hannah
Eric Lazear
Chwan Hong Foo
Katie Stiles

Other Lab Personnel
J.Charles Whitbeck, PhD
Huan Lou
Manuel Ponce de Leon, MS
Li Zuo, DMD