Promoter-associated histone modifications and establishment of the developmental gene expression program during early embryogenesis

Animal bodies including that of humans develop from a single cell, the fertilised egg. The development of embryos from fertilised eggs happens through proliferation of embryonic cells and a coordinated sequence of changes of these cells, which leads to formation of a variety of tissues. This process is governed by the coordinated activation of thousands of genes at the right time and in the right cells and is called genetic regulation of development. Each of the cells carry the same set of genes yet the activity of genes varies greatly in various cell types. When and where genes get switched on is a key determinant of normal development and is encoded in the DNA sequence present in every cell.

However, there is an additional mechanism of gene control, which operates by selective packaging of DNA (the genes and their switches) in the cell nucleus. The packaging of DNA (called chromatin) varies between cells and is important for marking the genes for activation. The marking and order of DNA packaging is inherited between cells during cell proliferation and may also be inherited from the oocyte and the sperm after fertilisation. This process is called epigenetic inheritance and has important implications in how inherited information is conveyed from parent to the embryo.

In this project we will study how one particular type of epigenetic information may contribute to the regulation of genes during embryo development. We will use a small animal model zebrafish, which is popular for genetic analysis of development due to the similarities in the way fish and human embryos regulate their genes during development and because the small transparent fish embryos can be easily cultured and monitored in laboratory conditions. We will study how proteins important in DNA packaging (histones) are marked in gametes and embryos to ask whether they are important for conveying epigenetic information from the parents to the developing embryo. We will explore the DNA codes of epigenetic regulation using biochemical assays that will tell us about histone marks on gene switches, and by analysing the DNA sequences which carry the codes for DNA packaging at the sites of the switches of genes active during embryo development. Our findings will be important for understanding the regulation of genes in normal development and will be useful for those studying the reasons for abnormal gene regulation, which causes a variety of disease both in development and in adult life.

BACKGROUND - Understanding the mechanisms of epigenetic regulation and their phenotypic consequences in development is a key prerequisite to unravelling the disease-causing mechanisms involving epigenetic change. The mechanisms of epigenetic signals (DNA methylation, histone posttranslational modifications (PTM) and RNAs) in the maintenance and reprogramming of cell fates remains elusive. Premarking of histones on promoters is present at early ontogenic stages and in sperm, suggesting predictive roles in establishment of a developmental gene expression.

OBJECTIVES - We address histone modifications in transcription regulation during earliest stages of embryo development. We shall exploit the transcription-free stages of zebrafish development to study the temporal regulation of chromatin modifications from gametes to transcriptionally active embryo and ask: 1. Does the pattern of PTMs in gametes predict pattern of chromatin modifications. 2. Can parent-specific histone PTMs be traced in the developing vertebrate embryo and do parental PTM patterns impact developmental gene expression? 3. What sequence information (CpG content) is required for the deposition of H3K4me3 and transcription start site choice?

METHODS - We will use small cell number ChIP and RNA sequencing in embryos and gametes, and apply our recent advances in deciphering core promoter codes to decipher the signals associated with deposition of the transcription initiation machinery, histone PTMs and nucleosome positioning signals by genome-scale analysis.

EXPECTED OUTCOME - This work will shed light on the relationship between chromatin regulation and transcription initiation at zygotic genome activation of the vertebrate embryo. It will elucidate the contribution of histone PTMs to developmental program of gene expression for normal ontogeny of a complex organism and provide critical evidence and a resource to study transgenerational epigenetic inheritance in a tractable vertebrate model.

The projects will have a large academic impact, described in detail in "Academic beneficiaries". It is perfectly aligned with the BBSRC strategic priority "Data Driven Biology" - it includes analysis and interrogation of next-generation sequencing datasets, analysis of the impact of genome variation, the extraction of quantitative information, and the development of new visualisation approaches for answering biological questions. Even though it addresses a fundamental scientific problem, the results and the methodologies developed within the project have the potential for a broader impact:

IMPACT ON HEALTH - The research focuses on the understanding of fundamental processes during early development, including the mechanism of activation of developmental programmes and the genes that control it, often associated with genetic and multifactorial disease. Understanding their early activation will have impact on understanding their function and the generation of hypotheses about the mechanism of disease. The events during the transition from gametes to dividing embryo and its activation will characterise key epigenetic events following fertilisation and provide a wealth of data that can be exploited for studies of stem cell biology, and transgenerational epigenetic inheritance and their applications.

IMPACT OF NEW METHODOLOGIES IN DEVELOPMENTAL GENOMICS - The experimental protocols we plan to develop for enabling genome-wide studies on limited amount of material will have impact on future efforts with cell type specific epigenomics. Applications will include include human (patient) samples, other animal model organism, and commercial species.

IMPACT OF THE GENERATED DATA AND SOFTWARE TOOLS - To maximise the impact of genome-wide datasets produced in the course of the project, it is necessary to provide them in an easily accessible and documented way. The combination of the data and tools to manipulate them is expected to have additional impact beyond the areas directly addressed by the project itself.

IMPACT ON THE EXPERTISE IN COMPUTATIONAL AND DEVELOPMENTAL BIOLOGY - The expertise in computational genomics and epigenomics in particular, is currently in high demand that vastly exceeds the supply of qualified, talented people. Training postdoctoral researchers in an interdisciplinary setting by collaborating groups with strong track record in computational genomics and developmental genetics bridges the training gap between computational and wet lab and has a potential to produce future research leaders. The computational methods developed on the project will benefit the community by increasing productivity through reusing and extending the softwares created here.

GENERAL IMPACT ON THE UK SCIENCE - University of Birmingham and Imperial College London are currently expanding their activities to become world leaders in the areas of genomics, integrative and systems biology. The competitive research program that we propose here will represent a large step towards that goal. Our training activities will provide exposure to our research and skills to talented students in developmental and computational biology across Europe, and serve as a platform for their recruitment to the UK.

IMPACT ON GENERAL PUBLIC AND APPRECIATION OF SCIENCE - Our outreach activities will span the audiences from general public to students. Within general public, we will organise activities that cover age groups from primary school pupils to adults. In addition to conveying the importance of our research, we shall address topics such as why zebrafish is a good organism to study human biology and disease, how whole genomes are studied, how biologists struggle with vast amounts of data etc. We expect this to benefit both the general public and the scientific community by increasing public appreciation of science and scientists, and by early exposure to current research.

The details of implementation are provided in "Pathways to Impact".