Date on Master's Thesis/Doctoral Dissertation

1-2021

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Microbiology and Immunology

Degree Program

Microbiology and Immunology, PhD

Committee Chair

Chung, Donghoon

Committee Co-Chair (if applicable)

Sokoloski, Kevin

Committee Member

Sokoloski, Kevin

Committee Member

Casella

Committee Member

Casella, Carolyn

Committee Member

Graham, James

Committee Member

Clark, Barbara

Committee Member

Lee, Donghan

Author's Keywords

alphavirus; RNA; antiviral

Abstract

Alphaviruses are positive sense, single strand, RNA viruses. These viruses occur on every populated continent. Alphaviruses are divided into two clades, the New-World and Old-World viruses. The New-World viruses include Eastern (EEEV), Western (WEEV), and Venezuelan equine encephalitis viruses (VEEV), and cause neuroinvasive disease. The Old-World viruses include Chikungunya (CHIKV) and Ross River viruses (RRV), and typically cause multijoint arthralgia. There are currently no approved antiviral therapeutics or vaccines for any alphavirus, making them a high priority for antiviral drug design and discovery. A benzamidine inhibitor (ML336) of VEEV was characterized, and determined to inhibit replication of VEEV RNA during infection of BHK-21 cells, a fibroblast model. This activity was due a loss of synthesis of new viral RNA. This compound had no effect on RNA synthesis in uninfected cells, making it a promising target for therapeutic development. The inhibitory activity of ML336 was highly specific for VEEV, having no effect on RNA synthesis of CHIKV. A potential interaction between ML336 and the VEEV nsPs was examined, but these results were inconclusive. ML336 and related compounds were used to generate resistant mutant VEEV. These isolates were sequenced and it was revealed that mutations were concentrated in a region of nsP2 of unknown function. Analysis of these mutant viruses revealed delayed growth, RNA synthesis, and translation of viral proteins in BHK cells. There was also a growth delay seen in SH-SY5Y cells, a model of neuronal infection. These findings indicate that this region of nsP2 is likely involved in RNA synthesis of VEEV, and shows promise as a target of antiviral drug development.

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Virology Commons

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