Development of a reporter system to study Yes-associated protein-1 activity

Abstract

Yes-associated protein (YAP) is a transcriptional co-activator that promotes regulation of genes active in growth and proliferation. YAP activation upon cancer therapy is widely associated with therapy resistance in various tumors.

YAP activation has recently been shown to mediate therapy resistance in epidermal growth factor receptor (EGFR) mutant lung cancer. While EGFR-targeted therapy eradicates most tumor cells, some cells activate YAP to survive treatment. YAP facilitates the survival of tumor cells by directly repressing the expression of pro-apoptotic Bcl2 modifying factor (BMF). Preventing YAP activation upon EGFR-targeted therapy leads to highly increased BMF expression, resulting in enhanced apoptosis.

While the role of YAP in mediating resistance to EGFR-targeted therapy is well-established, the upstream activators of YAP following targeted therapy remain unknown. This project aimed to develop a cell-based reporter system for monitoring YAP activity in living cells to facilitate future screening of YAP upstream regulators upon EGFR-targeted therapy. The reporter system was built upon the direct relationship between YAP activity and BMF expression in EGFR-mutant lung cancer cells, utilizing the mNG21-10/11 split-fluorescent protein system to monitor the expression of BMF in living cells. While the reporter system was successfully constructed, we failed to detect fluorescent signal from the reporter. This is most likely due to either insufficient BMF expression or non-optimal complementation of the split-fluorescent protein components. In conclusion, further optimization is needed to construct a fully functional reporter system.