UNDERSTANDING THE INTERPLAY OF ENHANCERS, CHROMATIN PRIMING ELEMENTS AND SIGNALS REGULATING DYNAMIC GENE EXPRESSION IN DEVELOPMENT

The formation of a fully functional organism during embryonic development is primarily coordinated and controlled by extra-cellular signalling modulators and regulators of gene expression. A fascinating aspect of embryonic development is that the timing of how different cells form is always the same. The reason for this synchrony is that all information for our body plan is encoded in our DNA, which contains instructions on when our genes should be expressed and when they should be silent. Moreover, developing cells communicate via signals to synchronize their development and activate genes to produce proteins, called transcription factors which can "read" the genetic code. These factors work together with many other proteins to ensure that genes are expressed in the right cell and at the right time. However, our DNA is packaged into a compact structure called chromatin which on the one hand ensures that it fits into the nucleus, but on the other hand restricts transcription factor access. How chromatin structure, transcription factors and signals work together to properly regulate gene expression is unclear. To answer this question, we will use an in vitro differentiation system that mimics normal development in vitro. We will collect data on all genes, and we will use computational biology methods to answer the fundamental question of how the balance and timing of gene expression are controlled in development.

The developmental control of gene expression involves the cell-type specific expression of transcription factors and their binding to specific chromatin landscapes with defined structural features modulating factor binding to enhancer and promoter elements. We still have a limited understanding of the molecular details ensuring that genes are expressed at the correct time and in the correct cell type. One answer to this question is the finding that not all cis-regulatory regions are bona-fide enhancer elements. It was found that the onset of RNA-synthesis at developmentally regulated genes involves marking them for future expression at defined genomic regions, a process called chromatin priming. How such elements interact with enhancers has a fundamental influence on the dynamics of tissue specific expression and determines where and when a gene is fully expressed. We also know that external signals can drive a specific developmental trajectory and impact on both enhancers and priming elements. We therefore hypothesize that the correct sequence of chromatin priming regulated by external signals is essential for temporal gene expression control. Ideally, we would like to know the dynamic activity of such elements to be able to predict the tissue-specific activity of genes.

This grant is a collaboration between wet lab researchers, bioinformaticians and mathematical modellers. We will use the differentiation of mouse embryonic stem cells into blood to examine at the global level the cis- and trans regulatory requirements for chromatin priming and its role in temporal gene expression control. We will gather systems-level kinetic data from normal and perturbed differentiation to feed into the construction of predictive mathematical models for temporal gene regulation. This work will provide novel mechanistic insights into how gene expression programs are coordinated in development and generate new computational tools to predict their response to perturbations.

Who will benefit from this research?
Our work will have an impact not only on our immediate research field but also far beyond. If successful, our work will benefit a number of diverse research fields. This includes:
1. Developmental biology because the relevance of signalling pathways and transcription factors for the regulation of developmental-stage-specific gene expression is not well understood and shared by many developmental processes in all mammals. Our functional enhancer annotations will be widely used, in particular by researchers who perform single cell chromatin accessibility assays.
2. Researchers working on transcriptional regulation in a chromatin context - these mechanisms are highly conserved in mammals.
2. Stem cell research because the molecular details of early haematopoietic specification are still not fully understood.
3. Mathematical modelling and bioinformatics research because our genome-wide data and those from perturbed systems will provide ample opportunity for mathematical modelling and developing novel methods for data integration. In addition, we have already produced extensive datasets from our LoLa work, which will be complemented by the information of inducible factors.

How will they benefit from this research?
1. We will make our system-wide data sets publicly available.
2. We will generate data that will be highly relevant to scientists studying other developmental/differentiation pathways both in academia and industry.
3. Our work will generate a number of resources such as a collection of tracer cell lines. We will make this resource available to all members from industry and academia who wish to use it.
4. We will develop novel bioinformatics and statistical machine learning pipelines, which we will distribute as open-source software, as we have done with a variety of algorithms developed for the ENCODE Consortium, including the widely used Irreproducible Discovery Rate (IDR).
5. Our work will enhance the skills base in the UK. Future advances in biology and medicine will depend on building a skills base consisting of researchers who will be capable of thinking both in cellular and molecular terms as well as in system-wide terms, and researchers working on this grant will be exposed to the forefront of research in this field.
6. Along the same lines, we will attract PhD and Master's students into this area. Birmingham is part of an MBTIP studentship scheme together with Leicester and Warwick and the CCB and Cazier/Brown run Master's courses in bioinformatics.