Understanding how enhancer chromatin transduces extracellular signalling during developmental transitions in human pluripotent cells

A cell's identity is defined by its genes. A cell located in the heart, for example, will express heart genes, but not genes important for liver or brain function. Conversely, liver cells express liver genes but not heart or lung genes. During embryonic development animals are formed by different groups of stem cells, which are able to give rise to the myriad of different cell types found in an adult organism. Cells "know" which cell types they should become because of signals they receive, usually sent by neighbouring cells so that cells in the right place at the right time take on the appropriate identity. Responding appropriately when the levels of these signals reach certain thresholds is crucial not only during development but also in maintaining a healthy state. When these responses fail the consequences include developmental disorders and cancer. When stem cells undergo the process of differentiating into a new cell type, they need to turn on genes appropriate for the new cell type, but also turn off the genes that define a 'stem cell' identity and also keep off any genes associated with other cell types. This process is tightly regulated by a group of proteins called chromatin remodellers. DNA is not "naked" within cells but exists in the form of chromatin, i.e. it is wrapped around proteins in a way that keeps it compact and stable. Chromatin remodellers are able to change how tightly packed specific regulatory parts of the chromatin are. In this way chromatin remodellers are important for making some genes available to respond if the right signal is received by a cell, while other genes are kept silent. We are interested in how the signals which enter a cell are interpreted to produce a change in gene expression. We recently discovered a previously undetected step in an otherwise well studied signalling pathway in mouse pluripotent cells. Signal activation in these cells results in a very fast event in which the chromatin of key regulatory sequences gets reset from a 'waiting' configuration to an 'active' configuration. We further showed that different chromatin remodelling proteins are required for different steps in this resetting event. We now wish to determine how human pluripotent cells respond to activation of these same signals. Our current data indicates that a resetting event occurs in human cells, but we've already detected species-specific differences. In the current study we will move beyond the scope of the mouse work to define how cells respond to changes levels of different signalling pathways, and to determine why some cells are responsive to signals while others are not. We will use what we learn to devise methods for controlling the responsiveness of cells to signals, something that would be of enormous benefit in regenerative medicine and in treatment of diseases where cells exert aberrant signalling responses, such as cancer. We are using cutting-edge methodologies analysing chromatin behaviour at the genome-wide level and at the single molecule level. Overall, this proposal represents a comprehensive investigation of how early embryonic human cells are directed to form the myriad of cell types present in an adult organism, through which we will devise ways in which to control this process.