Objective
Progenitor, or stem cells, that give rise to insulin-producing islet cells emerge from embryonic epithelium within the early pancreatic bud. This work focuses on the basic molecular mechanisms that shape the cellular microenvironment and 3D epithelial architecture where pancreatic endocrine cells are born. Understanding these basic processes will lead to novel insights into regenerative and replacement therapies for diabetes mellitus.
Background Rationale
In order to properly differentiate, cells read and respond to cues from their surrounding environment. These cues are transduced into changes in gene expression, which lead to changes in overall cell fate. While traditional models focus on chemical cues, recent evidence has suggested that the mechanical forces placed on a cell also guide differentiation. Insulin-producing β--cells arise from a highly dynamic 3D epithelial tissue structure, termed the epithelial plexus. Epithelial plexus formation occurs through a process tube-forming process termed lumenogenesis, in which cells constrict and rearrange themselves to form tubes. Lumenogenesis involves the generation of tension-based forces that places mechanical stress on cells. After plexus formation, select progenitor cells within the plexus undergo further constriction and differentiate into endocrine cells. Thus, both plexus formation and endocrine differentiation involve transitions in the mechanical environment, suggesting that the biomechanical cues may influence both of these processes.
How are biomechanical cues transduced into genetic responses? Work from our lab and others has shown that the Hippo pathway is a key mechanosensory pathway involved in directing development. This proposal will focus on an upstream regulator of the Hippo pathway, known as Merlin. Merlin is located on periphery of the cell, and is poised to interpret cues from both the tension places on cell junctions and from deformations to the cytoskeleton. Merlin can impact gene expression through regulation of transcriptional co-activator, Yap. Thus, we believe that understanding the function of Merlin in the pancreatic epithelium will be instrumental in understanding how pancreatic progenitors properly differentiate.
Description of Project
An innovative approach to treating Type 1 diabetes is the generation of functional beta cells in the lab for transplantation into human patients. While companies such as Vertex have succeeded and are currently in clinical trial, problems remain with suboptimal progenitor differentiation. To better understand how we can increase differentiation of these cells, we choose to learn more about how the cell differentiation in the body using mouse embryos as a model system.
Recent evidence has pointed to the roll of mechanical cues in directing the differentiation of endocrine cells. Our proposal here will work to uncover a mechanosensory molecular circuit that guides differentiation of endocrine cells. Specifically, our work will focus on a protein called Merlin. Merlin is regulator of the Hippo pathway, a key pathway involved in sensing mechanical strain placed on cells and transducing that to changes in gene expression. Our preliminary data indicate loss of Merlin produces dramatic effects on the pancreas; at the earliest stages of pancreas development, Merlin mutants are full of cysts and intriguingly have an excess of endocrine cells. This proposal will uncover how Merlin functions to facilitate proper pancreatic development and endocrine differentiation.
Anticipated Outcome
We have already found that loss of Merlin from the early pancreatic epithelium causes dramatic disruption of plexus formation and cellular differentiation. By E14.5, loss of Merlin causes large cysts throughout the pancreatic epithelium, failure of plexus formation, and dysregulation of Hippo pathway components. Intriguingly, Merlin mutants also display an increase in β-cell mass. Our work suggests that Merlin may act as a negative regulator of endocrine differentiation. In Aim 1 of this grant, we will determine how Merlin regulate formation of the pancreatic epithelial plexus, and whether it is involved in facilitating communication between neighboring cells. In Aim 2 of this grant, we will determine if Merlin represses certain transcription factors to facilitate endocrine differentiation. Finally, in Aim 3, we will investigate if Merlin functions to stabilize actin in cells fated to become ductal cells. Together, this work will increase our understanding of the mechanisms that endocrine progenitors use to respond to mechanical cues, and how those mechanical cues facilitate differentiation.
Relevance to T1D
Type 1 diabetes affects about 1.8 million Americans and is a devastating disease which leads to loss of insulin-producing pancreatic β-cells. There is substantial interest in understanding how cues from the environment facilitate differentiation of these cells. Recent work has shown that cells respond to mechanical cues to properly differentiate, but the underlying molecular circuitry is unclear. Our work here will uncover one such circuit, which can then be used to enhance regenerative medicine efforts.