Composition of a regulatory locus and the impact on phenotype and disease

Lead Research Organisation: University of Edinburgh

Abstract

It was assumed that most mutations/variations that cause human diseases would be due to changes in the coding regions of genes affecting the structure of proteins. This view has changed dramatically over recent years and it is now clear that the majority of mutations related to disease predisposition and even variability within the human population occur outside genes. The assumption is that these differences reside in regions of the DNA (known as regulatory domains or enhancers) that control protein production in the cell. Our research focusses on how changes in these regulatory domains can lead to congenital abnormalities. Although development of the human foetus is highly reliable, in approximately 1 in 100 cases deformities occur. Some of the most frequent abnormalities that occur affect the skeleton of the arms and legs and structure of the head and face. We have identified mutations in regulatory domains responsible for a spectrum of deformities in these features of the newborn, by using mammalian model systems such as the mouse and our goals are to investigate the mechanism by which this class of mutations causes congenital defects.

Technical Summary

Genome wide association studies (GWAS) highlight SNPs (single nucleotide polymorphisms) in noncoding DNA associated with disease traits for complex or common diseases. Undoubtedly, cis-regulatory mutations cause multiple human congenital defects which show a direct association of regulatory mutations with effects on gene expression. Enhancer adoption and recently, ectopic enhancer activity were shown to occur in patients with chromosomal deletions or duplications that remove TAD boundaries. Hence, examination of the mechanisms by which regulatory mutations perturb gene expression is fundamental to understanding the basis of congenital defects.
We showed that disrupted gene regulation of the signalling molecule SHH is responsible for skeletal abnormalities and others have shown involvement in brain and craniofacial defects. Mutations in the Shh limb-specific cis-regulator, the ZRS, cause appendicular skeletal defects which has subsequently, led to the categorization of a broad spectrum of defects into the ‘ZRS associated syndromes’. Our studies range from defining causative mutations to attempting to identify regulatory processes involved in disease, focusing on causative mechanisms and investigating general principles that underlie long range gene activation. We showed that dominant regulatory mutations result from both gain and loss of transcription factor binding capacity. One series of point mutations in human hijacks the normal regulatory machinery involved in setting the Shh expression boundary to generate ectopic expression and skeletal defects.
In addition we are investigating the role that chromatin organization plays in supporting enhancer activity and controlling gene expression. In general, we are investigation the overall architectural components required for cis-regulators to operate over large genomic distances. Known chromatin organizers such as CTCF binding sites are being surveyed to understand their direct role on gene expression and their contribution to the phenotype of the developing mouse. Systematic removal of CTCF sites in the large Shh regulatory domain results in change in 3D chromatin organization and one site underlies the appearance of the holoprosencephaly (craniofacial and brain defects) spectrum in mouse. We are presently developing imaging techniques to map chromatin changes that occur during developmental gene activation and to observe the dynamics in live cells. Our pursuit of long-range regulatory mechanisms are important in understanding developmental gene activation, and the contribution of non-coding mutations to congenital abnormalities.

Publications

10 25 50
 
Description Masters Programme
Geographic Reach National 
Policy Influence Type Influenced training of practitioners or researchers
Impact Developed Masters by research programme for Edinburgh University
 
Title Mice with chromosome 5 engineered 
Description Serial deletions of large region of Chromosome 5 that include the Shh locus 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Provided To Others? No  
Impact Gain insight into developmental regulation of brain, limb and gut 
 
Title Model for studying the human condition holoprosencephaly 
Description A regulatory mutations that lowers the expression of the sonic hedgehog 
Type Of Material Model of mechanisms or symptoms - human 
Year Produced 2017 
Provided To Others? No  
Impact These findings will be published in the upcoming year 
 
Title Skeletal Changes 
Description Modification of skeletal development by creating a series of mutations using the CRISPR/Cas genome engineering system 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Provided To Others? No  
Impact Manuscript being prepared 
 
Title limb cell lines 
Description A cohort of immortalised cell lines from developing limb tissues. New lines now contain a fluorescent marker for Shh expression for genetic screens 
Type Of Material Cell line 
Year Produced 2010 
Provided To Others? Yes  
Impact These lines have provided deeper insights into limb development and Shh signaling. 
 
Title osteoarthritis model 
Description A mouse mutations that results in osteoarthritis that shows a novel pathway responsible for joint inflammation and deformation 
Type Of Material Model of mechanisms or symptoms - human 
Provided To Others? No  
Impact A better understanding of the molecular mechanism responsible for osteoarthritis 
 
Description Acardi-Goutieres Syndrome 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided specialised methodology and intellectual input
Collaborator Contribution Has provided mutation data and phenotypic analyses
Impact Generated a mouse model for Aicardi-Goutieres Syndrome
Start Year 2007
 
Description Congenital disease and genome conformation 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Contribution to a body of work to unravel a disease mechanism
Collaborator Contribution Contribution to work and provided patient samples
Impact Publications
Start Year 2020
 
Description Congenital disease and genome conformation 
Organisation University of California, San Francisco
Department Department of Bioengineering and Therapeutic Sciences
Country United States 
Sector Academic/University 
PI Contribution Contribution to a body of work to unravel a disease mechanism
Collaborator Contribution Contribution to work and provided patient samples
Impact Publications
Start Year 2020
 
Description Corneal development 
Organisation University of Edinburgh
Department Centre for Integrative Physiology
Country United Kingdom 
Sector Academic/University 
PI Contribution Contribute experimental information and advice to students and Postdoc.
Collaborator Contribution Contribute experimental information and data
Impact 19387074 doi.org/10.1016/j.scr.2015.10.016 doi.org/10.1016/j.exer.2016.05.021
 
Description Embryonic growth and patterning 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Meier Gorlin Syndrome has been identified in patients and we made a genetic model to further probe the molecular disease mechanism Mouse model for analyzing cell proliferation and pattern formation in limb development
Collaborator Contribution Provides a genetic means to determine growth rate on organogensis. Also brought in a student to work collaboratively on the project
Impact A mouse model for the Meier-Gorlin (primitive dwarfism) syndrome.
Start Year 2011
 
Description Genomic architecture 
Organisation University of California, San Francisco
Country United States 
Sector Academic/University 
PI Contribution Generated a number of mutations targeted to affect chromatin organization
Collaborator Contribution provided the expertise in analysing the chromatin structure
Impact Data generated for a future publication
Start Year 2017
 
Description Innate immunity 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided specialised methodology and intellectual input
Collaborator Contribution provided basic immunology data to advance our understanding of the innate immune system
Impact Generated a number of mouse mutations which affect the innate immunity system
 
Description Long-Range Regulation 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution expertise, intellectual input,access to data,
Collaborator Contribution expertise, intellectual input
Impact Multidisciplinary. One manuscripts in preparation and work ongoing on nsecond
Start Year 2015
 
Description Matthew wood Syndrome 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided specialised methodology and intellectual input
Collaborator Contribution Provided studentship and patient data
Impact Analysis of genetic models (mouse and zebrafish) to understand retinoic acid involvement in Matthew Wood (PDAC) syndrome
Start Year 2008
 
Description Preaxial Polydactyly Patients 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Sharing patient information
Collaborator Contribution Sharing information on patientsProvided patient data
Impact 18463159
Start Year 2007
 
Description Preaxial Polydactyly Patients 
Organisation University of Oxford
Department Weatherall Institute of Molecular Medicine (WIMM)
Country United Kingdom 
Sector Academic/University 
PI Contribution Sharing patient information
Collaborator Contribution Sharing information on patientsProvided patient data
Impact 18463159
Start Year 2007
 
Description Syndactyly 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Chromosomal analysis and molecular biological data
Collaborator Contribution Provided patient data and phenotype analysis and chromosomal analysis
Impact Generated an animal model for human syndactylism DOI:10.1002/humu.21615
Start Year 2006
 
Description UBR5 
Organisation Cancer Research UK
Department Edinburgh Cancer Research UK Centre
Country United Kingdom 
Sector Academic/University 
PI Contribution Intellectual input, experimental resources and technical assistance
Collaborator Contribution Studentship
Impact Collaborative studentship doi:10.1371/journal.pone.0157079
Start Year 2009
 
Description Wt1 
Organisation Medical Research Council (MRC)
Department MRC Human Genetics Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided specialised methodology and intellectual input
Collaborator Contribution Basic biochemistry analysis
Impact Multidisciplinary (mouse genetics and Biochemistry) doi:10.1038/ng.494
Start Year 2006
 
Description evolution of Shh 
Organisation University of Cape Town
Department Department of Zoology
Country South Africa 
Sector Academic/University 
PI Contribution Functional analysis of Shh in mammalian flight
Collaborator Contribution Initial identification of element
Impact Manuscript in prepaation
Start Year 2012
 
Description Interview for National Radio Program (Radio 4) 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact General discussion of genes and their roles for the layperson
Year(s) Of Engagement Activity 2020
 
Description Interview for a lay science book on evolution and development 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Several interviews by the author of a science book for the general public. Our work was discussed extensively in one chapter
Year(s) Of Engagement Activity 2019