The encoding and interpretation of FGF signals in mammalian cell fate choice

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During the development of an embryo, cells become different from each other and do so in a manner that is highly regulated in time and in space. This process of diversification is regulated by the coordinated activity of transcription factors inside cells, which determine what the cells become, and by signals emitted and received by cells that allow them to communicate their states with each other. The workings of transcription factors as the masters of cell identity are increasingly well understood: they bind to the DNA and can activate or repress the expression of cohorts of genes that will endow cells with specific functions and identities. Transcription factors act in a combinatorial manner, i.e. some gene expression programs are the result of overlaying the activities of several individual transcription factors. Signals are received by specific receptors and passed on to their targets via a signal transduction pathway in the cell and are used to share information about their state. How signals interact with the activity of transcription factors, remains open to discussion. It could be that an effector of a signalling pathway acts like a transcription factor i.e. that it contributes to some sort of code that defines a unique identity for every cell. Alternatively the effector might have a more permissive role e.g. by opening up a molecular landscape (chromatin structure) for the action of the transcription factors, or as a factor that enhances or weakens the activity of those transcription factors at the level of single cells. Our proposal aims to distinguish amidst these possibilities by focusing on the FGF/MAPK signalling pathway, in two experimental systems that lend themselves to quantitative single cell analysis: the preimplantation mouse embryo and its correlate, the ES cells.FGF is an extracellular signalling molecule which triggers a signalling pathway that activates a protein kinase, MAPK. The activity of MAPK is known to influence the behaviour of cells, in particular their differentiation into more specialized cell types. FGF/MAPK activity is widespread in developing embryos, and although many different cells receive this activity, the response is different in different cells and at different times. Our project is aimed at understanding the basis for this observation. Signalling through MAPK is very common in normal and pathological conditions and we are therefore certain that our findings will have an impact beyond our experimental system. To gain a better understanding of the modes of action of FGF/MAPK signalling, we will focus on the earliest cell fate decisions in a mammalian embryo. Before the embryo implants in the uterus, a group of naïve cells, the Inner Cell Mass (ICM), becomes subdivided into two different populations: the Primitive Endoderm (PrE), that will give rise to extraembryonic tissue, and the naïve pluripotent Epiblast (Epi) that will give rise to the embryo. The decision to become PrE depends on a small number of transcription factors and FGF/MAPK signalling. We can reproduce these events in culture using engineered Embryonic Stem (ES) cells in which we can induce the fate decision event by activating a transcription factor and modulate the response by manipulating FGF/MAPK signalling. By focusing on single cells we can learn details of the decision which cannot be gauged from coarser approaches.An essential part of the problem is to develop reporters for MAPK signalling in single cells and link signalling activity to the transcriptional inputs and outputs of the process. This combination will allow us to obtain quantitative, dynamic data of the process, and inform novel mathematical models of cell decision making.