A Structural and Functional Investigation of the Essential HSV-1 Protein ICP27

Herpes simplex virus 1 ICP27 is an essential multifunctional regulatory protein that assumes different roles during infection. ICP27 disrupts multiple aspects of cellular gene expression, contributing to the shutoff of host protein synthesis. ICP27 also recruits RNA polymerase II to viral replication sites and mediates the export of viral mRNA. Furthermore, ICP27 has also been implicated in nuclear protein quality control, cell cycle control, activation of stress signaling pathways and apoptosis. ICP27 performs its activities by interacting with RNA and a myriad of proteins. However, how ICP27 is able to interact with so many different binding partners and how these interactions are regulated during the viral life cycle is not well understood. ICP27 has been reported to form homo-dimers and to undergo an intramolecular N- to C-terminal interaction. It’s hypothesized that these different configurations provide the flexibility it needs to interact with various binding partners and events or factors that promote one isomer over the other may serve to regulate different ICP27 activities. At present, how these configurations are related, and their specific functions, have not been well characterized. Moreover, the molecular details as to how ICP27 targets host RNA processing and the wider impacts its activities have on the host transcriptome have also not been detailed. In these studies, we (1) use a bimolecular fluorescence complementation and western blot-co-immunoprecipitation approach to perform a structural analysis in order to elucidate the functions of ICP27’s different conformations, (2) utilize a combination of biophysical and molecular techniques in order to detail the molecular mechanism of how ICP27 inactivates SR protein kinase 1, an important regulator of SR proteins, to inhibit cellular splicing, and (3) perform next-generation sequencing to globally Identify ICP27 modulations to host gene expression. The results from our studies show: (1) ICP27’s conformational isomers utilize different protein domains for their assembly and these isomers can interact with different binding partners, (2) the ICP27 RGG box binds to the SRPK1 substrate docking groove in a similar manner and affinity as SRPK1’s cellular substrate SRSF1, and (3) ICP27 promotes accumulation of host antisense transcripts during infection. Together, the information gained in this dissertation expands our understanding of the molecular principles by which ICP27 functions