Elucidating novel pluripotency signalling networks

I will investigate the proteins that control stem cell behaviour.

Stem cells are a very important and interesting cell type, as they can either divide into a larger population of stem cells ("self-renew"), or develop into one of the many types of tissues and organs in the human body ("differentiate"). The balance between these alternative cell fates is fundamental to human growth and development, and this process often go awry in human disease. In addition, it is possible for researchers to instruct stem cells to develop into adult tissues and organs in the laboratory. Therefore, by fully understanding stem cell behaviour, we may be able to both treat diseases and manipulate stem cells to generate replacement tissues and organs for patients; for example, by using stem cells to produce heart muscle to repair a damaged heart following a heart attack.

Whether stem cells self-renew or differentiate is controlled by signals from their immediate environment. However, the ways in which stem cells interpret these signals is poorly understood. We therefore need to identify the lines of communication used by stem cells to interpret environmental signals leading to a change in stem cell behaviour. Knowledge of the communication lines that tell a stem cell how to behave will potentially allow us to re-wire these communication systems so that we can either keep stem cells growing indefinitely, or instruct them to develop into adult tissues and organs.

One of the most important components of cell communication systems are proteins known as kinases, which play an important role in determining stem cell behaviour. Kinases can be readily blocked using chemicals, which disrupts the lines of communication. This is a relatively simple way to rewire stem cell communication systems so that the stem cells behave in a particular way. Therefore, this research project aims to uncover the kinases that control stem cell behaviour, knowledge which potentially will be exploited to form cells of a particular tissue or organ using chemicals.

In my recent research we have found a kinase known as ERK5, which is used by stem cells to stop self-renewing and start development towards adult tissues and organs. I aim to investigate how ERK5 is connected to ESC communication systems, and whether we can use chemicals to alter ERK5 communication patterns to modify stem cell behaviour. I also aim to identify other kinases that are involved in other important ESC communication systems, and again use chemicals to shut down these systems and artificially change stem cell behaviour in the laboratory.

Pluripotent Embryonic Stem Cells (ESCs) have the capacity to differentiate into all cell types in the adult body, and their application for the purposes of regenerative medicine and disease modelling has shown significant promise in recent years. However, it remains a challenge to harness the developmental capacity of ESCs, thereby capturing distinct pluripotent and differentiated states for use in regenerative therapeutics. A highly promising novel approach is to identify components of the phosphorylation machinery that control pluripotency and differentiation. Phosphorylation networks ultimately modulate the function of key regulatory targets such as transcriptional circuits to direct developmental potential. However, the kinases involved in these regulatory systems have not been extensively explored.

Here, I propose to uncover fundamental molecular bases of pluripotency regulation and identify strategies to manipulate key signalling components using small molecules. The primary aim of this project is to identify novel kinases and mechanisms by which phosphorylation controls gene expression to modulate the developmental potential of ESCs. Preliminary data from my lab suggests that the ERK5 kinase signalling pathway is a key regulator of pluripotency, and I will explore the molecular mechanisms underpinning this regulation. I will also exploit an inter-disciplinary combination of cutting-edge chemical biology, gene editing technology and phenotype-based analysis to identify and specifically perturb novel kinases that modulate ESC pluripotency. My extensive preliminary data, including identification of the ERK5 pathway and validation of a high-throughput screen for kinase inhibitors that modulate pluripotent ESC states, demonstrates the feasibility of the proposal. This research will generate new opportunities for chemical manipulation of ESC phosphorylation networks, allowing distinct developmental fates to be synthetically programmed in the laboratory.

This project aims to identify novel kinases and signalling pathways that control ESC pluripotency and differentiation. Therefore, this research will benefit:

1. The academic researchers directly involved in the project, by developing their research skills and experience through this varied, ambitious and multi-disciplinary research. The researchers involved will also benefit by acquiring further professional and transferable skills, in particular the PDRA, who will be assisted and encouraged to develop public engagement and communication skills in a variety of ways.

2. The worldwide academic community, including researchers interested in signal transduction, protein kinases, ESC biology including human ESCs, developmental pathways and cellular programming for regenerative medicine/disease modelling. More broadly, my research will also benefit scientists and clinicians who work in application and translation of Stem Cell technologies. These communities will immediately gain new ideas and scientific knowledge from our data, which in future will lead to development of novel methodologies and protocols to improve our basic understanding and medical application of Pluripotent Stem Cells. In addition, my laboratory will generate and make available many tools and reagents to study kinases identified in this research (including antibodies, cDNA constructs and mutants, kinase inhibitors and knock-out ESC lines) to facilitate research projects in other laboratories.

3. Commercial private companies aiming to develop effective platforms for reprogramming, culturing and differentiating Pluripotent Stem Cells for regenerative applications and disease modelling. In particular, we envision that our findings will be primarily exploited by MRC-PPU partner companies and other interested parties to elaborate novel protocols and methods to improve the feasibility and success of Stem Cell approaches for use in commercially-viable disease treatments. Improved Pluripotent Stem Cell culture protocols using kinase inhibitors is a key outcome which may be realised rapidly, whilst long-term applications will include elaboration of novel protocols and procedures, based on manipulation of kinase signalling, to facilitate generation of differentiated tissues and lineages for regenerative therapies and disease modelling. Finally, private companies with access to small molecule programmes and sophisticated chemical biology will potentially identify and develop drugs based on kinase inhibitors identified in our research, which may in future be used as drugs to treat disease.

4. The general public will benefit from this research through increased awareness and understanding of science, particularly in my field of Stem Cell biology. I will be an active participant in public outreach, introducing simple concepts of Stem Cells and their potential uses in treatments of disease, and publicising exciting findings from my laboratory to those beyond the academic and pharmaceutical communities. The MRC-PPU is committed to a number of outreach forums, which I will utilise to deliver effective public outreach. In the longer-term, this research aims to contribute to improvements in public health and welfare by providing a more detailed understanding of pluripotency and differentiation, and mechanisms by which these can be modulated, to facilitate development of effective regenerative therapeutics and disease modelling strategies to improve treatment of a variety of diseases.