Do lamin A/C and emerin mutations in satellite cells contribute to Emery-Driefuss muscular dystrophy?

Lead Research Organisation: King's College London
Department Name: Randall Div of Cell and Molecular Biophy

Abstract

Muscle is made of thousands of muscle fibres, which are repaired and regenerated by muscle stem cells called satellite cells. Muscular dystrophies are a group of inherited disorders that are characterised by progressive muscle weakness and degeneration. Since muscle is responsible for movement, its gradual loss in muscular dystrophies can severely affect the quality of life of a patient, and in some disorders, drastically shorten their lifespan. Compromised satellite cell function is thought to contribute to loss of muscle in some of these conditions, including Emery-Dreifuss muscular dystrophy (EDMD). It has been shown that EDMD is caused by problems in genes that make proteins involved in maintaining a healthy nucleus. We intend to examine satellite cells from patients suffering from EDMD to understand how these mutated proteins affected their function and so muscle repair. We will also use mouse models of the disease to study satellite cell biology. Findings on mouse satellite cells should be broadly applicable to man. Theoretically, manipulation of satellite cells could both augment and prolong muscle function in people with muscular dystrophy, which also has the advantage of maintaining a muscle environment still capable of responding to other forms of therapy.

Technical Summary

Autosomal Emery-Dreifuss muscular dystrophy (A-EDMD) is caused by mutations in the LMNA gene, that encodes four intermediate filament-like proteins, the major being lamin A and C. Together with B-type lamins, they form part of a proteinaceous network called the nuclear lamina, which underlies the nuclear membrane. EDMD can also be X-linked, in which mutations of the EMD (aka STA) gene are responsible. EMD encodes emerin, an inner nuclear membrane protein, which forms a complex with lamin A/C, cross-linking chromatin, through the nuclear lamina to the inner nuclear membrane. Mutations in LMNA and EMD are thought to result in satellite cell dysfunction. Since satellite cells are the resident stem cells responsible for repair of skeletal muscle, this dysfunction may directly contribute to EDMD.

Our long-term objective is to understand the role that lamin A/C and emerin plays in satellite cells and how their mutation causes EDMD. Satellite cells from muscles throughout the body will be examined as their heterogeneity could form the basis of different responses of muscle groups to disorders such as EDMD. Expression profiles of these nuclear proteins will be determined in wild-type quiescent satellite cells and during their activation, proliferation, self-renewal and differentiation using immunostaining. Muscle and satellite cells from lamin A/C and emerin null mice will be examined to determine how lack of these proteins affects satellite cell behaviour. Their ability to make muscle and self-renew will also be assayed in vivo using transplantation. Since ~85% of X-linked EDMD result from null mutations in EMD, these studies have direct relevance to the human disorder.

Mutant proteins in most A-EDMD and ~15% of X-EDMD patients cause a gain or change of function. How these same mutated proteins affect mouse satellite cell behaviour will be examined following viral-mediated delivery. Since human and mouse myoblasts fuse to form mosaic multi-nucleated myotubes, many signalling molecules must interact. Findings on mouse satellite cells therefore, should be broadly applicable to human, but muscle cells obtained from EDMD patients will also be examined for cell cycle and differentiation defects. Theoretically, manipulation of satellite cells could both augment and prolong muscle function in all muscular dystrophies, with the advantage of also maintaining an environment capable of responding to other therapeutic interventions. General principles of satellite cell regulation are also relevant to muscle loss in diseases such as cancer and during aging. This project falls within the musculoskeletal research priority area for PSCSB.

Publications

10 25 50
 
Description Consultant for Pharmaceutical company
Geographic Reach Multiple continents/international 
Policy Influence Type Participation in advisory committee
 
Description Muscular Dystrophy Campaign Strategy
Geographic Reach Multiple continents/international 
Policy Influence Type Membership of a guidance committee
 
Description BBSRC London Interdisciplinary Biosciences PhD Consortium (LIDo)
Amount £1,000 (GBP)
Funding ID Student rotating in the lab, who then may decide to undertake the PhD. 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2017 
End 06/2017
 
Description BBSRC London Interdisciplinary Biosciences PhD Consortium (LIDo)
Amount £77,314 (GBP)
Funding ID BB/J014567/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 07/2014 
End 09/2017
 
Description BBSRC London Interdisciplinary Biosciences PhD Consortium (LIDo)
Amount £73,659 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2016 
End 09/2020
 
Description Editing the Nuclear Envelope: Using Human Induced Pluripotent Stem Cells and CRISPR-Cas Technology to Develop Novel Therapies for Skeletal Muscle Laminopathies
Amount £114,556 (GBP)
Funding ID 19GRO-PS48-0188 
Organisation Muscular Dystrophy UK 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2019 
End 09/2023
 
Description London Interdisciplinary Doctoral Programme
Amount £16,015,170 (GBP)
Funding ID BB/M009513/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2015 
End 09/2023
 
Title Collated novel pathogenic EMD mutations underlying X-EDMD 
Description Collated novel pathogenic EMD mutations from a large American cohort. 
Type Of Material Biological samples 
Year Produced 2011 
Provided To Others? Yes  
Impact Increases known mutations underlying X-EDMD significantly Increases the number of mutations in the literature - should help developing better genotype/phenotype correlations. Mutations available at http://www.umd.be/ 21697856 Brown Charlotte A (Aug, 2011) Novel and recurrent EMD mutations in patients with Emery-Dreifuss muscular dystrophy, identify exon 2 as a mutation hot spot., Journal of human genetics 56, 8, 589-94 
URL http://www.umd.be/
 
Title LMNA exon skipping 
Description Lmna constructs lacking exon 5 in retroviral expression vector. 
Type Of Material Technology assay or reagent 
Provided To Others? No  
Impact Allowed us to determine that LMNA lacking amino acids encoded by exon 5 were redundant to LMNA function and so could provide a potential therapy for certain diseases involving missense mutations. Scharner, J., Figeac, N., Ellis, J.A. and Zammit, P.S. (2015). Ameliorating pathogenesis by removing an exon containing a missense mutation: a potential exon-skipping therapy for laminopathies. Gene Therapy 22, 503-515. 
 
Title Muscle stem cell dysfunction and disease progression 
Description Found that muscle stem dysfunction could contribute to Emery-Dreifuss Muscular Dystrophy by examining muscle stem cells from mice that are null for the Lmna or Emd genes. Also created new mouse models by crossing these mice with others that have reporter genes expressed in muscle stem cells or myonuclei. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2012 
Provided To Others? Yes  
Impact Provides a methodology that opens a new avenue in understanding of the disease mechanism of EDMD. 
 
Title New human disease causing LMNA mutations for EDMD published 
Description Collated new pathological mutations that cause Emery-Dreifuss MD, tested some on cell function in culture. Published in 20848652 Scharner Juergen (Feb, 2011) Novel LMNA mutations in patients with Emery-Dreifuss muscular dystrophy and functional characterization of four LMNA mutations., Human mutation 32, 2, 152-67 
Type Of Material Biological samples 
Year Produced 2011 
Provided To Others? Yes  
Impact Increases the number of mutations in the literature - should help developing better genotype/phenotype correlations. Mutations available at http://www.umd.be/ 
URL http://www.umd.be/
 
Title Retroviruses containing pathological LMNA species 
Description Cloning of pathogenic LMNA mutant species from human patients allows them to be efficiently expressed in cells to determine how they affect cell function. 
Type Of Material Technology assay or reagent 
Year Produced 2014 
Provided To Others? Yes  
Impact Better understanding of how different mutations in the LMNA gene affect cell function to lead to disease. Included in 20848652 Scharner Juergen (Feb, 2011) Novel LMNA mutations in patients with Emery-Dreifuss muscular dystrophy and functional characterization of four LMNA mutations., Human mutation 32, 2, 152-67 
 
Description Bioinformatics 
Organisation King's College London
Country United Kingdom 
Sector Academic/University 
PI Contribution Identifying and locating suitable mutations in LMNA and analysing effects on protein structure, charge and interactions.
Collaborator Contribution Provided bioinformatics expertise and software
Impact •Scharner, J., Lu, H.-C., Fraternali, F., Ellis, J. A. and Zammit , P. S. (2014). Mapping disease-related missense mutations in the Ig-like fold domain of lamin A/C reveals novel genotype-phenotype associations for laminopathies. Proteins: Structure, Function, and Bioinformatics 82(6):904-15. doi: 10.1002/prot.24465
Start Year 2012
 
Description Collation and testing of novel LMNA mutations 
Organisation Becton, Dickinson and Company
Country United States 
Sector Private 
PI Contribution Tested novel LMNA mutations for their effects on myogenic cell nuclear morphology
Collaborator Contribution Provided genetic diagnosis data from patients with suspected EDMD
Impact 20848652 Scharner Juergen (Feb, 2011) Novel LMNA mutations in patients with Emery-Dreifuss muscular dystrophy and functional characterization of four LMNA mutations., Human mutation 32, 2, 152-67
Start Year 2009
 
Description Collation and testing of novel LMNA mutations II 
Organisation Ohio State University Wexner Medical Center
Department Department of Neurology
Country United States 
Sector Academic/University 
PI Contribution Testing the effects of novel LMNA mutations underlying EDMD on myogenic cell nuclei
Collaborator Contribution Providing novel LMNA mutations for testing
Impact 20848652 Scharner Juergen (Feb, 2011) Novel LMNA mutations in patients with Emery-Dreifuss muscular dystrophy and functional characterization of four LMNA mutations., Human mutation 32, 2, 152-67
Start Year 2009
 
Description Exon-skipping on the LMNA gene 
Organisation University of Oxford
Department Department of Physiology, Anatomy and Genetics
Country United Kingdom 
Sector Academic/University 
PI Contribution Personnel and animal models to perform the experiments
Collaborator Contribution Expertise in exon-skipping and provided the necessary antisense -oligonucleotides for experiments
Impact Zammit published a paper containing the results of LMNA exon skipping. • Scharner, J., Figeac, N., Ellis, J.A. and Zammit, P.S. (2015). Ameliorating pathogenesis by removing an exon containing a missense mutation: a potential exon-skipping therapy for laminopathies. Gene Therapy 22, 503-515.
Start Year 2010
 
Description Making IPS cells 
Organisation University College London
Department Division of Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution Contributing EDMD patient cells and helping make IPS
Collaborator Contribution Expertise in making IPS cells
Impact Will make IPS cells
Start Year 2012
 
Description Measurement of muscle contraction 
Organisation King's College London
Department School of Biomedical Sciences KCL
Country United Kingdom 
Sector Academic/University 
PI Contribution Supplied EDMD mouse models for measurement of force generation
Collaborator Contribution Allowing force generation in muscles of EDMD mouse models to be measured
Impact 21364987 Gnocchi VF, Scharner J, Huang Z, Brady K, Lee JS, White RB, Morgan JE, Sun YB, Ellis JA, Zammit PS (, 2011) Uncoordinated transcription and compromised muscle function in the lmna-null mouse model of Emery- Emery-Dreyfuss muscular dystrophy., PLoS ONE 6, 2, Multi-disciplinary - combining cell biology and muscle physiology
Start Year 2009
 
Title Exon skipping sequences 
Description Paper accepted that will then potentially allow patent protection for the sequences involved in exon skipping. • Scharner, J., Figeac, N., Ellis, J.A. and Zammit, P.S. (2015). Ameliorating pathogenesis by removing an exon containing a missense mutation: a potential exon-skipping therapy for laminopathies. Gene Therapy 22, 503-515. 
Type Therapeutic Intervention - Drug
Current Stage Of Development Initial development
Year Development Stage Completed 2013
Development Status Actively seeking support
Impact Provides a potential therapy for some laminopathies. Identifies the potential to treat certain diseases that arise from missense mutations in proteins with structural repeats by exon skipping. 
 
Title Identification of new EMD mutations for diagnosis of EDMD 
Description Novel EMD mutations published on the http://www.umd.be/ database, allowing rapid confirmation after genetic testing for a diagnosis of X-EDMD. 
Type Management of Diseases and Conditions
Current Stage Of Development Refinement. Clinical
Year Development Stage Completed 2011
Development Status Under active development/distribution
Impact Novel EMD mutations published on the http://www.umd.be/ database, allowing rapid confirmation after genetic testing for a diagnosis of X-EDMD. 
URL http://www.umd.be/
 
Title Identification of new LMNA mutations for diagnosis of EDMD 
Description Novel LMNA mutations published on the http://www.umd.be/ database, allowing rapid confirmation after genetic testing for a diagnosis of EDMD. 
Type Management of Diseases and Conditions
Current Stage Of Development Refinement. Clinical
Year Development Stage Completed 2010
Development Status Under active development/distribution
Impact Novel LMNA mutations published on the http://www.umd.be/ database, allowing rapid confirmation after genetic testing of a diagnosis of EDMD. 
URL http://www.umd.be/
 
Description House of Lords - MDUK Vice-President's lunch 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Patients, carers and/or patient groups
Results and Impact MDUK Vice-President's lunch at the House of Lords that is attended by patients suffering from muscular conditions/carers/sponsors/donors/scientists/clinicians. Important occasion to explain the causes of muscular conditions and the research that is being conducted to understand and treat such disorders.
Year(s) Of Engagement Activity 2017,2018
 
Description Muscular Dystrophy Campaign conference 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Paper Presentation
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Members of the group gave a talk on muscular dystrophy to the Muscular Dystrophy Campaign's National Conference, to an audience of patients, carers, fund-raisers and supporters.

UK Neuromuscular Translational Research Conference 2016
Patient Day
The Kings Centre, Oxford, OX2 0ES
Monday 22 March 2016


After the talk there was a questions and answers session. Many suffers of muscular dystrophy later commented on how the talk had helped them to understand their condition.
Year(s) Of Engagement Activity 2009,2012,2013,2016
 
Description Muscular Dystrophy Campaign event 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Participants in your research and patient groups
Results and Impact Muscular Dystrophy Campaign hosted event where researchers spoke of their work and impacts to a lay audience composed of patients, carers, fund-raisers and supporters of the charity. Followed by an active discussion session with the above.

Charity fund-raisers enthused about ongoing research funded by the charity which will hopefully result in increased support for research.
Year(s) Of Engagement Activity 2010
 
Description Muscular dystrophy Campaign 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Type Of Presentation Workshop Facilitator
Geographic Reach National
Primary Audience Participants in your research and patient groups
Results and Impact Strategy research meeting to discuss funding priorities.

Prioritization of funding allocation.
Year(s) Of Engagement Activity 2012
 
Description School outreach 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Schools
Results and Impact ~25 6th formers used microscopes to examine muscle tissue and were engaged with the topic generating an active discussion.
100 or so year 7 pupils undertake some histological staining and examine sections under the microscope.

Hopefully an increased level of interest in university and science in general.

Successful, as will be repeated next year.
Year(s) Of Engagement Activity 2012,2016
 
Description Target MDC/MDC website, FSHSoc 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Regularly edit/contribute articles/quotes to Muscular dystrophy campaign publication (Target MD) and website e.g.
http://www.muscular-dystrophy.org/research/news/2700_new_insight_into_how_muscle_stem_cells_work

Also FSHSoc

url: http://www.fshsociety.org/assets/pdf/FSHSociety_FSHWatch_SummerFall2014.pdf
https://www.fshsociety.org/wp-content/uploads/2014/03/FSH_Society_WATCH_Fall-2015-FINAL-REV1-LR.pdf

Raises awareness of our work (MRC and from other funders) with patients, carers, fund-raisers and stakeholders.
Year(s) Of Engagement Activity 2009,2010,2011,2012,2013,2014,2015
URL https://www.fshsociety.org/wp-content/uploads/2014/03/FSH_Society_WATCH_Fall-2015-FINAL-REV1-LR.pdf