Varicella zoster virus (VZV) is the causative agent of chicken pox and shingles. The overall objective of this proposal is to understand the nature of physical and functional interactions between the complex VZV major transactivator, IE62, and specific components of the cellular transcription apparatus. Such information is critical to our understanding of the complex pathogenesis of VZV infection. It is particularly important in terms of the live attenuated VZV vaccine. Vaccination is now recommended for all children and recently the FDA approved the vaccine for use in the elderly for the prevention of zoster based on an extensive clinical trial. Currently little is known concerning attenuation of the vaccine virus. Since the IE62 gene accumulates the majority of mutations in the attenuated vaccine strains, understanding its interaction with host functions could lead to the development of second generation vaccine whose molecular mechanism of attenuation is well understood. The work proposed involves three Specific Aims.
Specific Aim 1 : Functional Interaction of IE62 with Cellular Factors. We will examine the functional interaction between the IE62 protein and two essential cellular transcription factors the Mediator complex and HCF-1. This will be done by mapping and mutation of interacting domains and expression of these mutations in the context of the viral genome.
Specific Aim 2 : Role of DNA Binding in the Mechanism of IE62 Activation. We will determine the specificity and affinity of the IE62- DNA Interaction and the effect of mutations in the DNA-binding domain on IE62 DNA-binding and transactivation and viral gene expression.
Specific Aim 3 : Interrelationship Between IE62 Structure and Function. We will mutate the IE62 SEAC domains and assess there function in in vitro transcription assays. The growth properties of virus carrying mutations in these domains will be determined. We will determine if the two transcriptional activation domains present in the IE62 homodimer are both required for IE62 function and what functions they perform. These studies will help us to understand the ways in which this important viral protein influences the functions of cells following infection. Since the live attenuated VZV vaccine will be used to prevent chicken pox and shingles throughout the U.S. population it is important to understand how the virus interacts with its human host. This work should also result in information that could be used in strategies to develop better anti-viral drugs and vaccines.
