Bachelor of Science, Walsh University, 2018, Honors

Herpes simplex virus type-1 (HSV-1) is a double-stranded DNA virus that infects human sensory neurons, causes cold sores, and enters into periods of latency. Heat shock proteins (HSPs) are universally conserved molecular chaperones that are present within all known species, including all prokaryotic and eukaryotic cells. These proteins function by facilitating protein synthesis, assembly, degradation, localization, and regulation of other proteins within the host cell. HSPs are stress-induced and therefore increase in abundance in response to various stressors exerted on the host cell, such as heat shock and viral infection. Of the seven families of HSPs, heat shock protein 70 (HSP70) is of particular interest to researchers due to its role in facilitating protein synthesis and degradation, as well as inhibiting apoptosis from occurring. Previous studies using HSV-1 strain GC and western immunoblotting indicate that HSP70 increases in level of abundance and that transcription of the hsp70 gene is enhanced during a 12-hour time-course infection. This project aimed to determine the location of HSP70 within HSV-1 strain GC infected HeLa cells to discern whether the protein moved from the cytoplasm to the nucleus in conjunction with the translation and translocation of HSV-1 proteins. Samples of uninfected and infected cells were harvested every two hours during an eight-hour time-course infection and the cells were processed to yield nuclear and cytoplasmic extracts. A western immunoblot was then used to detect HSP70 in the extracts, which was subsequently analyzed using densitometry. The results of this study illustrate that the majority of HSP70 remained within the cytoplasm, but did increase within both the nuclear and cytoplasmic fractions as infection proceeded. Nevertheless, the results do not support the theory of HSP70 functioning as a chaperone for the viral proteins, but suggest that HSP70 may be playing another role during HSV-1's reproductive and infectious (open full item for complete abstract)

Committee: Darlene Walro Ph. D. (Advisor) Subjects: Biology

Doctor of Philosophy, The Ohio State University, 2013, Food Science and Technology

Human norovirus (NoV) is the leading cause of acute non-bacterial gastroenteritis worldwide. Despite the significant health, emotional, and economic burden caused by human NoV, there are no vaccines or therapeutic interventions for this virus. This is due in major part to the lack of a cell culture system and an animal model for human NoV infection. Thus, a vector-based vaccine may be ideal for controlling this disease. The major capsid gene (VP1) of a human NoV was inserted into the VSV genome at the glycoprotein (G) and large (L) polymerase gene junction. Recombinant VSV expressing VP1 protein (rVSV-VP1) was recovered from an infectious cDNA clone of VSV. Expression of the capsid protein by VSV resulted in the formation of human NoV virus-like particles (VLPs) that are morphologically and antigenically identical to the native virions. Recombinant rVSV-VP1 was attenuated in cultured mammalian cells as well as in mice. Mice inoculated with a single dose of rVSV-VP1 stimulated a significantly stronger humoral and cellular immune response compared to baculovirus-expressed VLP vaccination. These results demonstrated that that the VSV-based human NoV vaccine induced strong humoral, cellular, and mucosal immunity in a mouse model. To further improve the safety and efficacy of the VSV-based human NoV vaccine, the gene for the 72kDa heat shock protein (HSP70) was inserted into rVSV and rVSV-VP1 vectors as an adjuvant, which resulted in construction of recombinant VSV expressing HSP70 (rVSV-HSP70) and VSV co-expressing human NoV VP1 protein and HSP70 (rVSV-HPS70-VP1), respectively. At the same inoculation dose, both rVSV-HSP70-VP1 and rVSV-VP1 triggered similar levels of specific immunity, even though VP1 expression by rVSV-HSP70-VP1 was approximately five-fold less than that of rVSV-VP1. To compensate for the reduced VP1 expression levels, the inoculation dose of rVSV-HSP70-VP1 was increased five-fold or same dosage of rVSV-VP1 and rVSV-HSP70 was combined vaccinated. Mice (open full item for complete abstract)

Committee: Jianrong Li (Advisor); Hua Wang (Committee Chair); Mark Peeples (Committee Member); Steven Krakowka (Committee Member) Subjects: Food Science; Virology