Wednesday, February 5, 2025
2/5/2025
PROJECT SUMMARY The ongoing theme of this program is to study the virus-cell interactions that affect HSV gene expression, with the primary focus on the mechanisms that regulate viral and cellular transcription during the course of infection. Progress during the last funding period has provided new insights into how the genes of HSV are transcribed in a regulated cascade during the course of viral infection. The studies also showed how viral processes and molecules can interact with and possibly regulate cellular transcription. We propose to extend on these studies and investigate new virus-cell interactions that affect virus and cell transcription. Ultimately, knowledge of how HSV genes are transcribed, and the virus-cell interactions that contribute to their regulated expression, may allow for strategies to block activated transcription and hence virus multiplication. The HSV genome during productive infection is a much more dynamic structure than classic cellular chromatin. The viral nucleoprotein allows for the rapid progression through the infectious cycle so that by 3hpi, all the events that must occur on the viral genome to produce new virus can be observed. This includes events that contribute to the shutoff of host cell transcription. This renewal focuses on virus-cell interactions occurring on the cellular (aim 1) and viral (aim 2) genomes, as well as the utilization of a cellular complex by the virus that is crucial for all pol II transcription, the mediator (aim 3). We have shown that the major viral regulatory protein, ICP4, binds to promoter/start-site regions of specific cellular genes early in infection as a function of the epigenetic state of the cell. The first aim will investigate the mechanism of interaction between ICP4 and the cellular genome, determine how ICP4 affects its expression, and explore the possibility that it functions differently in different cell types. The second will focus on the structure of the viral genome, and how viral proteins, including ICP4, affect the accessibility of the genome, and hence viral transcription. During the last funding period we showed that ICP4 interacts with and recruits a specific form of the mediator to the viral genome. In the third aim we will further explore the ICP4-mediator interaction, and test a model for the recruitment of mediator by ICP4 and its utilization for the activated and regulated transcription of viral genes. To undertake these studies, we have developed and refined a set of proteomics and genomics tools to specifically probe the dynamic state of viral genomes, and the viral and cellular proteins that associate with them. These tools combined with virus genetics allows us to elucidate the mechanistic underpinnings of processes occurring on the viral genome, including transcription.
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