Transcriptional Control of Vertebrate Limb Development

Lead Research Organisation: MRC National Inst for Medical Research

Abstract

Abnormalities in normal heart formation are the most common birth defect, occurring in 1 in 120 live births. If uncorrected, these defects can have serious debilitating effects on quality of life and in severe cases lead to premature death. Other less life-threatening birth defects also dramatically decrease quality of life in affected individuals. Among these, limb defects are the most common. We are interested in studying the causes of two human syndromes that lead to serious abnormalities in normal heart and limb formation. Patients with Holt-Oram Syndrome (HOS) have severely malformed hearts and limbs and those with Ulnar-Mammary Syndrome (UMS) are afflicted with abnormal limb and mammary gland development.|In each syndrome the corruption of the genetic information in a single gene is sufficient to cause all of the associated abnormalities. Armed with this knowledge of the exact genetic cause of each syndrome we propose to investigate the underlying biology of these diseases. Using the mouse and chicken, we are generating syndrome models that provide a powerful method to understand how loss of normal gene function leads to the disruption of heart and limb formation. In both animal models, the development of the heart and limbs occurs in a similar manner to our own and each animal model has experimental advantages we can exploit. Understanding the genesis of these syndromes will provide information vital in order to devise more effective diagnostic and corrective treatments in human patients.

Technical Summary

Limb defects are the second most common congenital abnormality present in human live births and diseases affecting the musculoskeletal system are a significant clinical problem, particularly in the older population. The goal of our work is to understand how the limbs form normally during embryogenesis and the genesis of limb abnormalities and diseases that affect the musculoskeletal system in humans. ||Limbs are formed from specific regions of the flank of the embryo. At early stages of embryonic development, the forelimb and hindlimb buds are morphologically uniform and indistinguishable from one another. During subsequent steps of development however, dramatic changes take place. Each bud develops to form a network of interconnected limb elements e.g. bones, muscles, and tendons-with morphologies characteristic of either the forelimb or hindlimb. We are using vertebrate animal models to understand the mechanisms that control the initiation of limb bud formation and the subsequent construction of the individual limb elements during embryonic development. A longer-term aim is to apply these findings to develop to potential therapeutic approaches that could contribute to maintenance and possible regeneration of these structures in later life.||We use the developing limb as a model system using predominantly mouse and chick model organisms but also zebrafish and frog (and in future, as part of collaboration, we also plan to include Bat). In general our strategy is to interrogate gene function using a combination of coordinated gene misexpression and gene deletion experiments, often in more than one model system. In the mouse, we take advantage of gene conditional alleles and a repertoire of Cre deleter transgenics that we have developed ourselves or have collected. We often include transgene reporters to aid in phenotyping. In the chick, we are using genetic techniques to misexpress genes (either retroviral vectors or electroporation or direct application of recombinant proteins or substances) in combination with more classical embryological approaches. A new and important aspect of current and future work is to develop the use of in vitro techniques as an alternative, complementary approach to our in vivo models. Currently this involves the generation and characterisation of cell lines derived from transgenic reporter mouse lines we have produced and collected.|

Publications

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Bickley SR (2014) Regulatory modulation of the T-box gene Tbx5 links development, evolution, and adaptation of the sternum. in Proceedings of the National Academy of Sciences of the United States of America

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Carkett MD (2011) 1, 2, 3: counting the fingers on a chicken wing. in Genome biology

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Duboc V (2011) Regulation of limb bud initiation and limb-type morphology. in Developmental dynamics : an official publication of the American Association of Anatomists

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Duboc V (2009) Building limb morphology through integration of signalling modules. in Current opinion in genetics & development

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Francis JC (2005) Notch1 signals through Jagged2 to regulate apoptosis in the apical ectodermal ridge of the developing limb bud. in Developmental dynamics : an official publication of the American Association of Anatomists

 
Title Collaboration with Tamsin van Essen 
Description Tamsin is a ceramicist-http://www.vanessendesign.com/- who is developing works based on interaction with the lab over the next 6 months 
Type Of Art Artwork 
Year Produced 2014 
Impact work in progress 
URL http://www.vanessendesign.com/
 
Title Collaboration with artist Ju Gosling 
Description Abnormal Exhibition. Abnormal developed from Ju Gosling aka ju90's artist's residency at the National Institute of Medical Research (NIMR), which took place between 2006 - 2007. Funded by the Wellcome Trust, the residency explored ideas about normality, and asked whether there is a 'Scientific Model of Disability' that is distinct from the 'Medical Model of Disability'? The exhibition toured nationally from 2008-2012 
Type Of Art Artistic/Creative Exhibition 
Year Produced 2010 
Impact national touring exhibition, media coverage, permanent website for exhibition, book produced associated with exhibition 
URL http://www.scientificmodelofdisability.co.uk/
 
Description Contribution to WHO meeting on causes of Thalidomide Embryopathy
Geographic Reach Multiple continents/international 
Policy Influence Type Implementation circular/rapid advice/letter to e.g. Ministry of Health
URL http://www.thalidomidetrust.org/thalidomide-embryopathy-world-health-organisation-report/
 
Description Member of Education Policy Advisory Panel (EPAG) for Gen Soc as part of Society of Biology
Geographic Reach National 
Policy Influence Type Participation in a advisory committee
URL https://www.societyofbiology.org/education
 
Description Vice president corporate affairs, Genetics Society
Geographic Reach National 
Policy Influence Type Membership of a guideline committee
 
Description Commonwealth Fellowship
Amount £2,000 (GBP)
Organisation Government of the UK 
Department Commonwealth Scholarship Commission
Sector Public
Country United Kingdom
Start 03/2011 
End 11/2012
 
Description EMBO Fellowship-2006
Amount £692 (GBP)
Organisation European Molecular Biology Organisation 
Sector Charity/Non Profit
Country Germany
Start 05/2006 
End 04/2008
 
Description EMBO Research Grant-2009
Amount £25,690 (GBP)
Organisation European Molecular Biology Organisation 
Sector Charity/Non Profit
Country Germany
Start 03/2009 
End 02/2011
 
Description EU-Intr-European Fellowship-IEF-(Limb-type ID)
Amount £155,632 (GBP)
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 12/2010 
End 11/2012
 
Description MRC Research Grant
Amount £488,429 (GBP)
Funding ID MR/S000038/1 
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 07/2019 
End 06/2022
 
Title 3D Mouse Limb anatomy database 
Description An interactive 3D anatomy atlas of the embryonic mouse forelimb and hindlimb 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2008 
Provided To Others? Yes  
Impact Improved Anatomy atlas that is more user-friendly and more stable across multiple platforms I get enquiries about the database The database is cited in publications 
URL https://www.emouseatlas.org/emap/ema/modelsummary/modelsummary.html
 
Title Antibody 
Description Tbx5 antibody polyclonal 
Type Of Material Antibody 
Provided To Others? No  
Impact ongoing work functional antibody 
 
Title Mouse limb anatomy database 
Description The Interactive 3D Mouse Limb Anatomy Atlas is a free, downloadable resource of mouse limb anatomy at embryonic stage (E) 14.5 days gestation. Users build, rotate, magnify, and virtually dissect models of the limb using 3D datasets of real forelimbs and hindlimbs using the JAtlas Viewer program. Individual muscles, tendons, and bones are colour-coded and annotated, and users can manipulate 3D models and generate virtual sections through the specimen. 
Type Of Material Biological samples 
Year Produced 2008 
Provided To Others? Yes  
Impact none 
URL http://www.nimr.mrc.ac.uk/3dlimb/
 
Title Pitx1 antibody 
Description Pitx1 antibody polyclonal 
Type Of Material Antibody 
Provided To Others? No  
Impact ongoing work 
 
Title Provider of transgenic mouse lines 
Description transgenic mouse lines eg Prx1Cre, Prx1CreERt2 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2006 
Provided To Others? Yes  
Impact numerous publications 
 
Title Prx1Cre and Prx1CreERt2 transgenic mouse lines 
Description Prx1Cre and Prx1CreERt2 transgenic mouse lines 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2010 
Provided To Others? Yes  
Impact Continually requested research tools that are finding new uses for a broader range of studies including cancer studies Continually cited in published work 
 
Title Tbx4 Antibody 
Description Tbx5 antibody polyclonal 
Type Of Material Antibody 
Provided To Others? No  
Impact ongoing work 
 
Title provider of transgenic mouse lines 
Description transgenic mouse lines 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2010 
Provided To Others? Yes  
Impact numerous publications 
 
Description Collaboration with plastics and reconstructive surgeons 
Organisation Great Ormond Street Hospital Children's Charity (GOSHCC)
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution Hosting and collaborating with ploastic surgeons with specialities in reconstructive surgery of hand Mr Branivan Sivakumar Miss Gill Smith Dr George Murphy Dr Log Murugesan
Collaborator Contribution Developing ongoing collaborative work to explore ways to improved surgical approaches and surgical outcomes in limb reconstructive surgery
Impact Mres project completed and succesful production of preliminary data. plans for further work
Start Year 2013
 
Description Collaboration with plastics and reconstructive surgeons 
Organisation Royal Free London NHS Foundation Trust
Country United Kingdom 
Sector Public 
PI Contribution Hosting and collaborating with ploastic surgeons with specialities in reconstructive surgery of hand Mr Branivan Sivakumar Miss Gill Smith Dr George Murphy Dr Log Murugesan
Collaborator Contribution Developing ongoing collaborative work to explore ways to improved surgical approaches and surgical outcomes in limb reconstructive surgery
Impact Mres project completed and succesful production of preliminary data. plans for further work
Start Year 2013
 
Description DECIPHER/DDD Radial Ray Defects^60 Holder-Espinasse 
Organisation Guy's Hospital
Country United Kingdom 
Sector Hospitals 
PI Contribution I have actively search for and established links with clinicians interested in limb defects. We have developed a strategy to identify patients/families for further study and the key goals for the short and medium term to best deliver impacst on improved patient care and to develop knowledge of the causes and treatments of limb defects
Collaborator Contribution Provide the clinical lead in partnership and have direct access/contact with patients
Impact only established in 2014
Start Year 2014
 
Description Quantifying the effects of treatments to reduce immunogenicity of tendon xeno/allografts 
Organisation Guy's and St Thomas' NHS Foundation Trust
Country United Kingdom 
Sector Public 
PI Contribution Initiating a pilot study with a collaborator, Mr Adil Adjued, Consultant orthopaedic surgeon, GSST to quantify the effects of treatments to reduce immune response/rejevtion of tendon xeno or allografts for surgical replacement following tendon injury/failure.
Collaborator Contribution Provide clinical link for study and samples for analysis
Impact work ongoing. Follwoing results of pilot will (i) publish study in clinical journal (ii) explore possibiklity to extend collaboration and seek funding for further work
Start Year 2016
 
Description Role of Tbx5 in limb soft tissue morphogenesis 
Organisation Medical Research Council (MRC)
Department MRC National Institute for Medical Research (NIMR)
Country United Kingdom 
Sector Academic/University 
PI Contribution Generation nof £D imaging data of limb development
Collaborator Contribution Significantly refined our analysis
Impact Peleg Hasson, April DeLaurier, Michael Bennett, Elena Grigorieva, L.A. Naiche, Virginia E. Papaioannou, Timothy J. Mohun, Malcolm P.O. Logan (2010) Tbx4 and Tbx5 acting in connective tissue are required for limb muscle and tendon patterning. Developmental Cell 18 (1) 148-156 PMID: 20152185
Start Year 2007
 
Description Role of nephronectin in forelimb development 
Organisation Medical Research Council (MRC)
Department MRC National Institute for Medical Research (NIMR)
Country United Kingdom 
Sector Academic/University 
PI Contribution Novel experimental approach adn observations
Collaborator Contribution novel experimental approach and observations
Impact Abu-Daya, A., Nishimoto, S., Fairclough, L., Mohun, TJ., Logan, MP., Zimmerman, LB. (2010) The secreted integrin ligand nephronectin is necessary for forelimb formation in Xenopus tropicalis Dev Biol Oct 23 PMID: 20977901
Start Year 2008
 
Description Using a Prx1 transgenic to study the periosteal stem cell niche 
Organisation UCB Pharma
Country United Kingdom 
Sector Private 
PI Contribution My lab is contributing a unique reagent that we produced that enables a cell-type of interest in bone to be studied and isolated. My lab contributes expertise in embryogensis, genetic tools and genetic approaches and imaging analysis
Collaborator Contribution Experise in bone biology, techniaues in bone biology and novel reagents for studying bone biology (particularly immuno therapy reagents and osteogenic bone matrices). A biopharma outlook on a biological and clinical probelm.
Impact BBSRC-CASE studnetship awarded to a PhD student who I am hositng in my lab but who will also work directly with teams at UCB
Start Year 2016
 
Description Wellcome consortium Phenotyping Mouse cKO 
Organisation The Wellcome Trust Sanger Institute
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution Wellcome Trust Strategic Award 2012-2017 £5,000,000 "Deciphering the Mechanisms of Developmental disease"
Collaborator Contribution Grant has been awarded but not started yet
Impact Grant awarded but not yet started
Start Year 2012
 
Description Advice to author David Nicholls 
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 provided advice to author preparing a novel with main protagonist a scientist

giving advice to author with very large audience to help produce a pro-science book that humanises the practice of science and demsytifies science and scientists to the general public
Year(s) Of Engagement Activity 2014
URL http://www.davidnichollswriter.com/
 
Description Contribution to TV programme 
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 Contributed to Channel 4 programme "Easter Eggs Live" providing facility and expertise to produce images of chicken and Emu embryos developing in ovo for a programme focused on explaining embryonic development (in ovo) to the general public

Good interaction with programme producers. Tangible output and wide audience reached
Year(s) Of Engagement Activity 2013
URL http://www.channel4.com/programmes/easter-eggs-live/4od
 
Description Contribution to WHO meeting on Thalidomide embryopathy 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Patients, carers and/or patient groups
Results and Impact WHO report published
http://www.thalidomidetrust.org/thalidomide-embryopathy-world-health-organisation-report/

After my contribution I estyablihsed new links with workers/policy makers on Thalidomide embryopathy
I have since given a presentation for the Thalidomide trust, been invited to write a report for Thalidomide trust and have established a collaboration with a Japanese group working on Thalidomide embryopathy
Year(s) Of Engagement Activity 2014
 
Description Gave talk for "Pint of Science" festival 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Gave a talk (alongside a clinical collaborator, Bran Sivakumar) to general public describing the work we do to study the rogins of limb defects and improving clinical treatments
Year(s) Of Engagement Activity 2016
URL https://pintofscience.co.uk/event/building-body-parts
 
Description Genetics Society executive committee member 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact genetics Society committee meetings
Genetics Society Executive meetings
Interview panels for genetics Society
Working group for Centenary celebrations
Dealing with Genetics Society journals (Heredity and Genes and Development) Liason with publihsers Springer-Nature and CSHP


genetics Society committee meeting
Year(s) Of Engagement Activity 2013,2015,2016,2017
URL http://www.genetics.org.uk/
 
Description NIMR Open day 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact 100 or so guests visited the NIMR to hear about the work that goes on at the MRC funded research institute

http://www.nimr.mrc.ac.uk/news/nimr-open-day-impresses-visitors/
Year(s) Of Engagement Activity 2013
URL http://www.nimr.mrc.ac.uk/news/nimr-open-day-impresses-visitors/
 
Description Presentation for Thalidomide Trust UK 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact open new avenues for interactions
Talk stimulated debate and future activities

After the talk I was asked to provide advice to former chairman of Trust who is writing a book on origins of Thalidomide
Also met potential collaborators for future work on Thalidomide causes and treatment
Year(s) Of Engagement Activity 2014
URL http://www.thalidomidetrust.org/public/national-advisory-council-nac/
 
Description SChoolls day NIMR 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? Yes
Type Of Presentation Workshop Facilitator
Geographic Reach Local
Primary Audience Schools
Results and Impact Set-up and run a demonstration of chick embryogenesis to enable school children to observe and get hand-on experience of animal models used in research

Always get good feedback and lots of questions from the students
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,2009,2010,2011,2012
 
Description School essay prize winner trip 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Local
Primary Audience Schools
Results and Impact Give presentation and demonstration to school essay prize winners and explain what we do here at NIMR

get positive feedback from students
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,2009,2010,2011,2012
 
Description University of Third Age 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact 180 attendees for regional U3A meeting at NIMR

Good feedback from u3A. will continue collaboration with NIMR for annual meetings
Year(s) Of Engagement Activity 2012
 
Description challenges in 3D bioprinting 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact group of 50-100 discussiing challenges and potential for use of 3d bioprinting in development of translational/regenerative medicine. representation from various interest groups from biologists to chemist to engenineers, patent lawyers industrialists and funding agencies

good networking and catalyst for future meeting to inspire interdisciplinary projects.
Year(s) Of Engagement Activity 2013