Gene regulatory networks in early development

Lead Research Organisation: MRC National Inst for Medical Research

Abstract

We study the very early stages of embryonic development when errors in our inherited genetic makeup can have serious consequences.|Understanding the genes that control our growth from fertilized egg to adult human is the key to understanding the impact of our individual genetic heritage on our health and life expectancy. We all have slightly different gene sequences from each other, and some of these differences, or polymorphisms, can have a profound impact on how we develop. |In the future we hope to be able to use gene therapy to ameliorate the adverse impact of these differences, but first we have to understand what each of these genes is doing in our bodies; when they are switched on (or off) during development, exactly where they are acting, and what other genes they may be affecting.|By looking at the first few hours of development in a non-human model organism (the frog Xenopus), we hope to improve our understanding of the critical initial steps of the developing embryo as it begins to use its own DNA to replace the proteins and other genetic material it received from its mother. Although fundamental, this process is still poorly understood, with only a handful of the genes involved being determined so far. |Recent technological advances now make it possible for us to analyse the complete set of active genes in a tissue sample or embryo through the generation of millions of tiny sequence fragments, and this will be the key to unpicking this crucial phase of development.|

Technical Summary

Elucidation of the gene regulatory networks managing the transcriptional programme of early development in vertebrates in an unbiased, genomic-scale manner.|Sequence errors in the genes involved during the early stages of vertebrate development can have very serious consequences for the viability and correct progression of the rapidly growing embryo. Our ability to best manage the consequences of genetic variability that have an impact in this period will depend on a detailed and thorough understanding of the mechanisms involved. We propose to investigate this using the Xenopus tropicalis model system, with its combination of highly accessible early embryos, diploid genome, and relative physiological similarity to humans. |A key step in early development is the point at which the embryo switches from synch-ronous cell division, under control of maternally derived gene products, to asynchronous division and the establishment of the zygotic transcriptional programme. This midblastula transition will be the primary focus of our research. The main objectives will be to try to understand better the processes that trigger it, and to lay out the initial programme of gene activation from the zygotic genome, including timings and functional relationships. |To investigate the transition in an unbiased manner requires a systematic, genomic-scale approach. The first step will be to use next-generation sequencing technology to create high-resolution time series of the expression levels of active molecular components in the early embryo. From these expression profiles we will identify the activated transcription factors, and this knowledge will feed into further rounds of experimental work, to identify and characterise the functional relationships between genes. |Experimental techniques will primarily be RNA-Seq on the Illumina platform for the transcriptional profiling, ChIP-Seq on the same platform for analysis of transcription factor targets and gene locus activation, and the production of in situ images for gene expression localization. We may also look at the use of other techniques for investigating gene networks, such as synthetic lethal screens. |Computational analysis will be applied early on to predict candidate relationships between genes from the relative timing and shape of their expression profiles, and the knowledge of which gene products are probably transcription factors. Later on in the project, comput-ational techniques, which will likely need to be developed, will be used to amalgamate all the experimental data and refine our understanding of the functional relationships between genes. The integrated sum of these relationships will define the gene regulatory network for this phase of embryonic development.|

Publications

10 25 50
 
Description Computational skills for wet lab biologists
Geographic Reach Multiple continents/international 
Policy Influence Type Influenced training of practitioners or researchers
 
Description National Institutes of Health/University of Virginia
Amount £64,639 (GBP)
Organisation National Institutes of Health (NIH) 
Sector Public
Country United States
Start 08/2011 
End 05/2016
 
Description National Institutes of Health/Yale University
Amount £300,366 (GBP)
Organisation National Institutes of Health (NIH) 
Sector Public
Country United States
Start 08/2011 
End 04/2015
 
Title sectional in situ analysis 
Description Extension of in situ annotation method to allow computational representation of gene expression patterns in tissue section images, and subsequent querying by axpression pattern to find (e.g.) candidate neural stem cell markers. 
Type Of Material Technology assay or reagent 
Year Produced 2011 
Provided To Others? Yes  
Impact publication in Developmental Neurobiology 
 
Title online database of gene expression profiles 
Description online database of gene expresion profiles in Xenopus tropicalis early development 
Type Of Material Database/Collection of data 
Year Produced 2014 
Provided To Others? Yes  
Impact described in: High-resolution analysis of gene activity during the Xenopus mid-blastula transition. Collart C, Owens ND, Bhaw-Rosun L, Cooper B, De Domenico E, Patrushev I, Sesay AK, Smith JN, Smith JC, Gilchrist MJ. Development. 2014 May;141(9):1927-39. doi: 10.1242/dev.102012. PMID: 24757007 
URL http://genomics.nimr.mrc.ac.uk/apps/profiles/
 
Description Xenopus miRNA in situ screen 
Organisation University of East Anglia
Department School of Biological Sciences UEA
Country United Kingdom 
Sector Academic/University 
PI Contribution Providing input into experimental design and data processing. Perform QC on the data, and upload into our existing datbase/iammge annotation system, XenMARK. Support the annotation process
Collaborator Contribution Originated the experiment. Created the in situs and organised teh data for upload, and teh web hosting iof images. Will be organising the annotation team.
Impact 693 in situ images of Xenopus MiRNAs awaiting annotation.
Start Year 2010
 
Description Xenopus oocyte transcriptome 
Organisation University of Cambridge
Department Gurdon Institute
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided computational expertise and practical analysis of high throughput sequencing data to characterise the transcriptome of Xenopus oocytes
Collaborator Contribution Developed our understanding of the pre-egg phases of Xenopus development, in particluar the mRNA burden of the developing oocyte.
Impact Publication in 2011. Combined computational expertise with wet lab expertise.
Start Year 2009
 
Description immune response in flies 
Organisation King's College London
Department Centre for Molecular & Cellular Biology of Inflammation
Country United Kingdom 
Sector Academic/University 
PI Contribution Ideas and design expertise for an pilot experiment to assess immune response in flies. Agreement to analyses data when received.
Collaborator Contribution Original research results upon which this experiment is based. Agreement to pay for the cost of the experiment, and to generate the mRNA samples required.
Impact None yet
Start Year 2012
 
Description sectional in situ analysis 
Organisation University Paris Sud
Department Institute of Neurobiology
Country France 
Sector Academic/University 
PI Contribution Provide the computational expertise and the database and coding framework to instantiate our ideas on in situ image annotation of sectional images.
Collaborator Contribution Providing the motivation and laboratory work to extend our ideas on in situ image annotation to sectional images.
Impact One publication and an extension to our publicly availabe database of in situ images. Combines wet lab research with computational ideas on data analysis and visualisation.
Start Year 2010
 
Description spinal cord regeneration 
Organisation Pontifical Catholic University of Chile
Country Chile 
Sector Academic/University 
PI Contribution experimental design sequence library construction data analysis
Collaborator Contribution primary experimental rationale experimental protocols experimental work data analysis
Impact multi-disciplinary Larrain - experimental expertise and resources Gilchrist - sequencing and compuational analysis
Start Year 2014
 
Description Mill Hill Essay 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? Yes
Primary Audience Public/other audiences
Results and Impact My essay in the impact of next generation sequencing technologies on ourselves was included in the 2010 Mill Hill Essays booklet.

(unknown)
Year(s) Of Engagement Activity 2010
 
Description Teachers professional development day 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Primary Audience Schools
Results and Impact 44 science teachers attending an evening of four talks and Q&A given by PIs at NIMR. Discussion was lively and informed.

In addition to what they got from the talks, the teachers found value in the connections they made between themselves at he event.
Year(s) Of Engagement Activity 2010