Description
Chapter 1 Ovarian cancer patients commonly experience relapse following treatment, suggesting the presence of a population of chemoresistant tumor-initiating cells (TICs) capable of repopulating a tumor. NF-κB signaling promotes ovarian cancer tumorigenesis and previous studies demonstrate that classical signaling supports chemosensitive, proliferative cells, while alternative signaling supports chemoresistant TICs. However, NF-κB is an unfit signaling pathway to target for combating relapse due to its critical role in immune function. Here, I investigate a role for WNT10A in OV90 ovarian cancer cells and implement ChIP-seq to identify genes regulated by the classical or alternative NF-κB transcription factors, RelA or RelB, respectively, that support TICs and promote ovarian cancer relapse. Our findings suggest that WNT10A is negatively regulated by RelB in TICs and may be regulating different stemness qualities in OV90 cells. Additional studies are required for defining the role of WNT10A in ovarian cancer and the regulatory interplay between WNT10A and RELB. Chapter 2 Cures for blood malignancies are curtailed by a shortage of hematopoietic stem cell (HSC) donors coupled with the potential for donor tissue rejection. By furthering our understanding of HSC development and maintenance we can make headway for autologous HSC transplants as a therapy for diseases of the blood. Previous studies identified the Wnt9a-Fzd9b ligand-receptor pairing to be specifically required for hematopoietic (HSC) emergence and proliferation, with EGFR acting as a required cofactor in the Wnt9a-Fzd9b signaling cascade. To investigate the intracellular mechanism by which this signaling occurs, a proximity labeling assay was conducted, revealing that components of clathrin-mediated endocytosis (CME) are recruited to Fzd9b upon stimulation with Wnt9a. Here, I investigate the effects of both clathrin knockdown and small molecule endocytic inhibitors on Wnt9a- Fzd9b signaling. I also generated EGFR fusion proteins to identify the EGFR domains required for transducing the Wnt9a-Fzd9b signal. Our findings indicate a complex endocytic network with Wnt receptor internalization playing a potential role in signal amplification. Ongoing studies will need to validate the fusion proteins and employ intracellular protein visualization to identify the route(s) that the signaling complex traffic to and the subsequent effect(s) this has on signaling.
