Mechanisms of Cellular Communication: The Role of Merlin in Pancreatic Development

PROJECT SUMMARY More than 30 million Americans have pre- or existing diabetes. An innovative approach to treat diabetes is to generate functional insulin-producing -cells for transplantation in vitro. This approach is currently in clinical trials, but a recognized problem is suboptimal progenitor generation. -cells originate from a multipotent progenitor population in the early pancreatic epithelium. Elucidating mechanisms that regulate progenitor specification and maturation in vivo using mouse models will instruct in vitro differentiation efforts. The Hippo pathway is emerging as a key mechanotransducive pathway involved in regulating organ development, however, mechanisms that regulate Hippo signaling in the context of pancreatic development remain unclear. This proposal will examine Merlin, a key regulator of Hippo signaling, to determine if it relays biomechanical signals during tissue morphogenesis. We have found that deleting Merlin leads to severely aberrant pancreatic morphogenesis and cell differentiation. Currently, we aim to elucidate both how Merlin is regulated and what its molecular function is in pancreatic development. The central hypothesis of this proposal is that Merlin enables pancreatic lumenogenesis by interpreting cues from cytoskeletal contractility to organize the apical membrane. In Aim 1, I will examine if Merlin directs pancreas formation non- cell autonomously in vivo by using mouse genetics and in vitro by using a 3-D spheroid model. Using a mosaic deletion system, I will assess if Merlin is required for propagation of tension-based cues. In Aim 2, I will determine the cellular function of Merlin; specifically, I will determine if Merlin facilities vesicular trafficking events required for lumenogenesis by using a novel live imaging system that our lab developed. In Aim 3, I will examine if mechanical cues regulate Merlin phosphorylation status and function in a 2D cell culture, through direct application of tension using cell stretching system. I will further determine if Merlin phosphorylation is required for YAP1/TAZ inhibition. Together, this data will provide us with insight into how mechanical cues guide pancreatic morphogenesis and cell fate. More broadly, our results will shed light on how progenitor specification and maturation occur and will guide regenerative medicine efforts.