SPATIOTEMPORAL CONTROL OF YAP LOCALISATION AND FUNCTION IN VITRO AND IN VIVO

Abstract

The regulation of organ size is an important aspect during growth and regeneration and is controlled by inputs from biochemical signalling pathways and mechanical stimuli. The Hippo-YAP pathway links between external physical forces and internal transduction of these mechanical cues as biochemical signals. YAP, a transcription co-activator shuttles between the cytoplasm and nucleus depending on upstream signals. The activity of YAP is highly dependent on its subcellular localisation, where nuclear localisation allows it to bind to transcription factors and elicit a genetic response. We synthesised an optogenetic tool that enables us to control the localisation of YAP. We studied the import and export dynamics of optoYAP, verifying the independence of each activation cycle. In cell culture, activated optoYAP promotes target gene expression and cell proliferation. We demonstrate that optoYAP can override a cell’s innate response to substrate stiffness to generate anchorage-independent growth. Similarly, opto-fYap can be used in zebrafish embryos to modulate target genes. We also established a transgenic zebrafish line expressing opto-fYap in cardiomyocytes, demonstrating its viability in the adult zebrafish. Overall, we show that optoYAP is functional in both cell culture and in vivo, providing a powerful tool to manipulate the localisation and therefore function of YAP.