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Examination of the Effects of ICP27 on Lytic Replication and the Establishment, Maintenance, and Reactivation of Herpes Simplex Virus 1 in Latency

Presenters Name: 
Kenneth Darcy
Co Presenters Name: 
Primary Research Mentor: 
Anna Cliffe
Secondary Research Mentor: 
9:30 - 10:15
Time of Presentation: 
2019 - 9:30am to 10:15am
Newcomb Hall Ballroom
Presentation Type: 
Presentations Academic Category: 
Grant Program Recipient: 
Not a Recipient

Herpes simplex virus (HSV) establishes a latent infection in neurons where it persists for life. During latency, viral lytic genes are silenced. Periodically HSV reactivates from latency resulting in the production of infectious virus. Reactivation is dependent on the activation of host cell signaling pathways to trigger transcription of the lytic genes. However, during lytic replication, the viral protein ICP27 is required for efficient processing and export of viral mRNAs. Interestingly, we have found that expression of ICP27 occurs earlier than the expression of other lytic transcripts during reactivation of HSV from latency. This indicates that ICP27 may play an important role in the reactivation process, specifically in the earliest phase of lytic protein synthesis. To examine the role of ICP27 during reactivation, I have created shRNA oligos that contain a short hairpin to target the ICP27 mRNA. These oligos were cloned into the pLKO.1-TRC cloning vector and inserts were sequenced to verify the presence of the shRNA construct. Lentiviral stocks were generated for each oligo and their titer was determined via colony formation assay in 6 well plates. The shRNAs will be validated in lytic replication following infection in ARPE cells by western blotting. The role of ICP27 in the maintenance of latency and reactivation will be investigated in an in vitro model of latent HSV infection of primary murine sympathetic neurons and analyzed via RT-PCR and immunofluorescence. These data will inform on whether ICP27 is required prior to expression of other lytic proteins in HSV reactivation from latency.