Browsing by Author "Staab, Peyton"
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Staab, Peyton (2024)Kaposi sarcoma (KS) is a cancer characterized by spindle cells thought to originate from endothelial cells. The most aggressive form is AIDS-related KS, which is one of the most common cancers in Sub-Saharan African regions with high HIV burden. The causative agent of KS, Kaposi Sarcoma Herpesvirus (KSHV), displays a biphasic lifestyle of latent and lytic phases. During the latent program, the viral DNA is tethered to the host chromatin without integration, through the viral protein LANA. This mechanism allows the KSHV genome to exploit the human cellular replication machinery to replicate and maintain viral DNA genomes. Recent, unpublished findings suggest that this mechanism is supported by interactions between LANA and the host transcription factor SOX18. The Ojala lab has shown that this key developmental factor SOX18 is significantly upregulated in KSHV-infected cells. Importantly, SOX18 inhibition by a small molecule inhibitor SM4 decreases the intracellular viral DNA copies and characteristic infected spindle cell phenotype. However, the mechanism behind the upregulation of SOX18 is not understood. Additionally, a previous BioID screen for SOX18 protein interactions discovered that an important structural protein and the ATPase of the host chromatin remodeling cBAF complex, ARID1A and BRG1, respectively, were the highest confidence interaction partners in KSHV-infected cells. Therefore, it was hypothesized that SOX18 and the cBAF complex, and their interactions, co-operate in viral DNA replication and could represent promising therapeutic targets for KS and other SOX18-dependent diseases. This study found that host signaling pathway NF-κB may be responsible for SOX18 upregulation. A BrdU pulldown assay with ectopic SOX18 expression showed enhanced KSHV DNA synthesis, which could be hampered with SM4 inhibition. These results further elucidate the role of SOX18 in viral genome replication. Furthermore, proximity ligation assays and immunofluorescence were performed to investigate interactions between SOX18, KSHV LANA and cBAF in KSHV-infected cells. Cell viability assays, protein expression by Western blotting, and KSHV genome copy quantification upon treatment with cBAF inhibitors were performed. In addition to SM4, inhibition of cBAF led to a regression of the spindle cell phenotype and reduced KSHV genome copies and showed that KSHV-infected cells are sensitized to cBAF inhibition. Infection with a constitutively latent virus supported these results. This thesis sheds light on the mechanism of SOX18 interaction with LANA and host chromatin remodeler in viral latent DNA replication and provides a deeper insight into possible future therapeutic strategies for KS.
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