Progressive Multifocal Leukoencephalopathy (PML) is a subacute demyelinating disease of the human central nervous system (CNS). PML primarily afflicts individuals with defects in cell-mediated immunity and there is strong evidence suggesting that PML results from infection of glial-origin cells, i.e., oligodendrocytes and astrocytes with the human neurotropic virus, JCV. Evidently, these cells are destroyed or transformed during the course of this disease, resulting in demyelinating lesions in the brain. Previous observations by us and others have indicated that the neurotropism of JCV is determined primarily at the level of viral early and late gene transcription. Analysis of viral RNAs during the course of infection indicate differential activation of major viral early and late promoters during the course of infection. Since the early and late promoters are divergent and overlapping, it is of particular interest to determine the mechanism(s) responsible for the preferential use of the early and late promoters before and after DNA replication, respectively. since the transition from early to late occurs during/after DNA replication, when the viral early gene product, T-antigen, is present in the cells, we hypothesize that T-antigen, in concert with cellular factor(s), orchestrates differential expression of viral promoters. The central goal of this research proposal is to identify the mechanism(s) that mediates viral gene expression during the course of infection, and determines the regulatory components (cis- and trans- elements) that are participating in this process. The experimental design utilizes a variety of biochemistry, molecular virology and genetics techniques to identify the key regulatory component(s) that plays a critical role in the stimulation of viral RNA synthesis and viral DNA replication. The information obtained from these experiments should increase our current understanding of cell-type specific gene transcription in the CNS and should provide insight into the replication of viral and cellular DNA in the brain.
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