Merkel cell carcinoma (MCC) is a rare and aggressive form of skin cancer that metastasizes rapidly and resists most current therapies. Approximately 80% of MCC tumors are associated with Merkel cell polyomavirus (MCPyV), which clonally integrates into the host cell genome prior to oncogenic expansion. MCPyV+ MCC oncogenic maintenance relies on the continuous expression of the viral tumor antigens, LTT and sT, such that cancer cell death occurs upon inhibition of tumor antigen expression. Additionally, the ability to express LT/sT in a potential host cell determines the virus’ narrow infectious tropism. We therefore sought to understand the molecular mechanisms that control the expression of the viral oncogenes at the transcriptional level during infection and oncogenesis. Through inhibitor screening experiments, we identified the histone acetyltransferases (HATs) p300 and CBP as key regulators of MCPyV transcription. We discovered that p300/CBP mediate viral gene expression through both direct acetylation of viral genome-associated histones and through coactivation of the NCRR-binding transcription factor NF-κB. We also demonstrated that HAT inhibitors (HATis) specific to p300/CBP were highly effective at reducing LTT expression in MCPyV+ MCC cell lines, leading to tumor cell death. We investigated the cell-type specific factors that regulate MCPyV transcription in order to improve our understanding of the cell type(s) that may support MCPyV infection and tumorigenesis. We identified the B cell development factors EBF1 and PAX5 as regulators of MCPyV transcription in only MCPyV+ MCC and not during infection, demonstrating that viral gene expression is regulated differently in the contexts of infection and cancer. Furthermore, we examined the effects of a hypoxic microenvironment on MCPyV gene expression, and found that hypoxic conditions could induce or repress viral transcription depending on whether the host cell is a site of active inflammatory signaling. In our studies, we have identified multiple factors and mechanisms that regulate MCPyV gene expression during infection and oncogenesis. We have also established a novel MCPyV+ MCC treatment strategy of targeting LTT/sT expression through downregulation of their transcription via small molecule inhibition.