The epithelial junction protein MarvelD3 in cell proliferation and migration

Lead Research Organisation: University College London
Department Name: Institute of Ophthalmology

Abstract

Epithelia are continuous layers of cells that delineate our tissues and organs. Individual epithelial cells interact with each other via molecular complexes that mediate adhesion but also function as sensors that transmit information about the environment, such as the presence or absence of neighbouring cells, to the cell interior. Integrity of epithelia is important for our organs to develop and function normally, and to protect us from our environment. For example, breaches in epithelial layers such as the skin or in the lining of the intestine can lead to serious infections and can occur due to chronic inflammations or acute infections by viruses and bacteria. Similarly, a characteristic of cancer cells is that they have lost the capability to sense the presence of neighbouring cells and hence continue to proliferate and migrate on top of their neighbours, or leave their tissue of origin by migrating to and invading other tissues and organs and, thereby form metastasis. On the other hand, in many adult tissues cells do no longer multiply and this can lead to reduced cell numbers and loss of normal organ function due to age or tissue damage. It is thus of fundamental importance to understand how cells in a tissue adhere to and recognise each other, how this influences their proliferative and migratory properties, and how we can exploit such mechanisms to manipulate their behaviour to address medical problems.

Here we propose experiments to investigate a new mechanism that, based on our unpublished results, links adhesion between cells to the regulation of cell migration and proliferation. Our first aim is to determine how this mechanism works during epithelial repair processes and, our second aim, how it contributes to normal development of epithelial tissues. Our third aim is to use this information and address a medical problem that is caused by a lack of cell proliferation. The cornea is a tissue at the front of the eye that is required for normal vision. It is formed by an epithelium on the outside and a layer of cells on the inside, called corneal endothelium. As cells in the corneal endothelium do normally not proliferate and regenerate, their numbers decline with age or when the cornea gets damaged. If their numbers are too low, the cornea loses its transparency resulting in a loss of vision. This seriously affects the availability and quality of human corneas for transplantation that are donated to treat patients with damage to the surface of the eye. Hence, we propose experiments to test whether the here-identified mechanism can be exploited to enhance the quality of donated human corneas and thereby enhance the number of corneas that are adequate for transplantation.

Knowledge of how cells sense their neighbours and transmit such information to the cell interior, and how we can manipulate such processes has a wide range of potential applications apart from the one that we will test here. The expected results will help us to think of new ways to aid wound repair after surgery and to treat devastating diseases such as chronic inflammations, certain infections and cancer.

Technical Summary

Epithelial cells adhere to each other via junctional complexes, enabling them to form cellular barriers that separate different tissues and body compartments. These adhesive complexes transmit signals to the cell interior to guide gene expression and cellular processes such as proliferation and migration. Tight junctions are one type of intercellular junctions and restrict paracellular permeability. They are also thought to signal to the cell interior and several signal transduction pathways have been identified that are regulated by tight junctions. However, we know little about how plasma membrane components of tight junctions contribute to the regulation of the junction-associated signalling mechanisms and about the importance of such mechanisms for developmental processes. This proposal focuses on MarvelD3, a recently discovered transmembrane protein of tight junctions. Our preliminary data indicate that MarvelD3 signalling regulates cell proliferation and migration. At least in part, this involves regulation of specific signal transduction mechanisms. Our aims are to identify molecular mechanisms by which MarvelD3 signals, to determine the role of MarvelD3 in a developmental model, and to test whether MarvelD3 signalling can be targeted to improve the quality of corneas donated for transplantation. The expected results will be important for the understanding of how cell-cell adhesion guides epithelial behaviour and to develop possible therapeutic strategies.

Planned Impact

Who will benefit from this research?

The immediate beneficiaries will be scientists working in allied fields at Universities as well as in industry. This includes scientists working in areas such as tissue engineering, infections and wound repair, as well as chronic inflammation and cancer biology. The development of new techniques to enhance the numbers of corneas of sufficient qualities for transplantation has many beneficiaries both nationally and internationally. Corneas for sufficient quality for transplantation are limited and will become even more limited due to our increasing age and medical treatments that affect the quality of donated corneas (e.g., laser treatments). Hence, beneficiaries will also be medical scientists, surgeons and patients, as well as the NHS and thereby the general public.


How will they benefit from this research?

The research will benefit allied scientists by providing them with the molecular details of a new mechanism that links cell-cell adhesion to the regulation of cell proliferation and migration that they can then test in their respective model systems. Translational scientists will benefit in a similar way. For example, scientists interested in carcinomas and tumour spreading will be able to build on our research to exploit the identified molecular pathway to suppress metastasis and tumour proliferation. In our own research environment, these results will benefit researchers and clinicians who work on corneal transplantation as they will be able to adapt techniques and tools developed in the proposal for clinical purposes. The research will also help to target proliferative conditions of the retinal pigment epithelium. A better accessibility of transplants with more endothelial cells will improve patients lives as waiting times will be shorter and transplant lifetime longer. This will have a positive impact on NHS services, reduce costs, and, thereby, benefit the taxpayer. Treating patients with better corneas will in the long term have a positive impact on their wellbeing and associated social costs, and will therefore also benefit society as a whole. The project will also involve training of a postdoctoral fellow and a technician in laboratory techniques that can benefit the private sector as well as public services through the NHS (e.g., preparation and evaluation of corneal transplants).

The expected results are likely to start to benefit other scientists within the lifetime of this grant, and we hope to be in the position to plan strategies how techniques developed for corneal transplants can be developed for clinical applications by the end of this grant.

Publications

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Matter K (2014) SnapShot: Epithelial tight junctions. in Cell

 
Description The tight junction protein MarvelD3 is shown to regulated cell proiferation and migration by tuning the MEKK1/JNK signalling pathway. This tuning mechanism is vital duirng the osmotic shock response during which it is required for junction maintenance and cell survival.

In Xenopus early development, MarvelD3 acts as a neural crest determinant due to its activity of JNK signalling and subsequently regulates the differentiation of neural crest derivatives such as melanocytes and cranial cartilage, we well as ocular development and morphogenesis.

The research also led to the development of new methodology to characterize corneal transplant quality by focusing on the corneal endothelium, another cell type derived from the neural crest. The method was then applied to determine optimal storage and preparation methods for corneal transplants prior to surgery.
Exploitation Route MarvelD3 could be targeted for cancer therapy and used to enhance efficacy of therapeutic drugs. MarvelD3 has also been genetically linked to Malaria resistance and it is possible that this mechanism could be exploited to design new anti-Malaria therapies.
Sectors Pharmaceuticals and Medical Biotechnology

 
Description The research included the development of new methods for the characterization of corneal transplants and the comparative analysis of different storage methods of corneas before they are transplanted. This research has thus informed the clinical community on how best to prepare corneal transplants for storage and surgery, and we expect that this starts to influence clinical practice. The research thus benefits the NHS as well as patients.
First Year Of Impact 2016
Sector Healthcare,Pharmaceuticals and Medical Biotechnology
 
Description PhD fellowship
Amount £100,000 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2016 
End 09/2019
 
Description PhD fellowship
Amount £100,000 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2012 
End 09/2015
 
Description Research Support Grant
Amount £12,395 (GBP)
Funding ID R170006A 
Organisation Moorfields Eye Charity 
Sector Charity/Non Profit
Country United Kingdom
Start 07/2016 
End 11/2016
 
Title Anti-xMarvelD3 
Description Rabbit polyclonal antibodies to detect Xenopus MarvelD3 by immunoblotting and immunofluoresecence 
Type Of Material Antibody 
Year Produced 2018 
Provided To Others? Yes  
Impact Support of other researchers 
 
Title MD3 reagents 
Description Reagents for the biochemical and functional analysis of MD3, a new tight junction-associated membrane protein that regulates subcellular signalling mechanisms (cDNA and siRNAs, adenoviral vectors, antibodies) 
Type Of Material Technology assay or reagent 
Year Produced 2010 
Provided To Others? Yes  
Impact A first paper describing a basic analysis of this paper is in press 
 
Title MDCK cell lines 
Description MDCK cells liens expressing inflammatory regulators to study effect interplay between inflammation and epithelial barrier properites 
Type Of Material Cell line 
Provided To Others? No  
Impact Used at talks and is being incorporated into a publication 
 
Title Xenopus MarvelD3 reagents 
Description Expression constructs for the analysis of Xenopus MarvelD3 
Type Of Material Technology assay or reagent 
Year Produced 2016 
Provided To Others? Yes  
Impact Reagents to express MarvelD3 in Xenopus for rescue of loss of function phenotype and functional analysis 
 
Description Analysis of MarvelD3 in development and tissue function in mice 
Organisation de Duve Institute
Country Belgium 
Sector Academic/University 
PI Contribution We have provided molecular and cellular tools for the analysis of MarvelD3
Collaborator Contribution Our partners provide expertise in genetically manipulated mice and in vivo analysis
Impact A paper published in 2014 (PMID:24567356) The genetically modified mice for conditional depletion of MarvelD3 have now been generated and we started to use them in our laboratory to analyse the function of MarvelD3 in specific tissues.
Start Year 2012
 
Description Improvement of corneal transplants 
Organisation Moorfields Eye Hospital NHS Foundation Trust
Country United Kingdom 
Sector Public 
PI Contribution Expertise in tissue culture and manipulation of cell signalling and cell-cell adhesion
Collaborator Contribution Handling and analysis of corneal transplants
Impact The research programme has recently been supported by a MRC PhD followship for a clinical fellow who works in our laboratory on the corneal transplants Two manuscripts have been published in the Br. J. Ophthalmology that are openly accessible (Bhogal et al., 2016)
Start Year 2012
 
Description Role of tight junctions in infectious disease 
Organisation University of Zurich
Country Switzerland 
Sector Academic/University 
PI Contribution Analysis of junctional signalling mechanisms in relation of infectious agents
Collaborator Contribution Provision of reagents and information
Impact no outputs yet
Start Year 2015
 
Description 4th China-UK Cancer (CUKC) Conference. Cardiff, Wales 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Discussions about junctions and cancer

I was asked whether I would be interested to participate in a follow-up meeting in China
Year(s) Of Engagement Activity 2015
 
Description Annual Student Lecture, IBMC, University of Porto, Portugal 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Postgraduate students
Results and Impact Interest of students for future laboratory visits

Discussions about collaborations
Year(s) Of Engagement Activity 2015
 
Description Cardiff-Peking Universities Cancer Institute, invited lecture 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact lively discussions

exchange of information
Year(s) Of Engagement Activity 2013
 
Description Cell Press Tumour Microenvironment LabLinks Symposium 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact diverse audience reached and interesting discussions

new collaboratoin
Year(s) Of Engagement Activity 2012
 
Description Cell polarity in cell and tissue function 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact General presentation of functional relevance of cell polarity and cell-cell adhesion in tissue function for an audience including graduate and postgraduate students as well as researchers from a wide spectrum of cell and developmental biology
Year(s) Of Engagement Activity 2017
 
Description Distinguished Lecture UCL Medicine 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Professional Practitioners
Results and Impact Discussion of future work
Year(s) Of Engagement Activity 2017
 
Description Engagement with parliament 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Policymakers/politicians
Results and Impact Event with discussion with members of parliament and their staff organised by the Royal Society of Biology
Year(s) Of Engagement Activity 2016
 
Description Epithelial Morphogenesis Symposium, Sapporo 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact interdisciplinary meeting

echange of reagents and protocols
Year(s) Of Engagement Activity 2012
 
Description Eye Research - an equal partner 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Vision Bridge is an organisation dedicated to informing the general public about contemporary eye research and to provide a platform to enable exchange between researchers, the general public and patients.
Year(s) Of Engagement Activity 2018,2019
URL http://visionbridge.org.uk/
 
Description Gordon Research Conference on Signalling by Adhesion Receptors 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Great discussions with peers

established collaborations
Year(s) Of Engagement Activity 2012
 
Description International Conference on the Molecular Structure and Function of Tight Junctions 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact interesting discussions with peers

exchange of research reagents
Year(s) Of Engagement Activity 2012
 
Description Participation in Charity Fund Raising Event 
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 Public/other audiences
Results and Impact Several hundred participants

NA
Year(s) Of Engagement Activity 2011
 
Description PhD students Berlin 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Lecture for students of a PhD programme in Germany and discussions about their own research projects
Year(s) Of Engagement Activity 2017
 
Description Regulation of Cdc42 in epithelial differentiation, Konstanz, Germany 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Invited lecture sparked interesting discussions

discussion of future collaborations
Year(s) Of Engagement Activity 2015
 
Description Regulation of Cdc42 in epithelial differentiation, Zurich, Switzerland 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Supporters
Results and Impact discussions about junctions and infectious disease

Start of a collaboration on tight junctions and infectious disease
Year(s) Of Engagement Activity 2015
 
Description Signal Transduction, Mexican Biochemical Society, Oaxaca, Mexico 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Talked led to discussions about future collaborations with Mexican laboratories

Possible interactions were discussed
Year(s) Of Engagement Activity 2015
 
Description Signalling at tight junctions 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Invited lecture

no actual impacts realised to date
Year(s) Of Engagement Activity 2013
 
Description TV programme 
Form Of Engagement Activity A broadcast e.g. TV/radio/film/podcast (other than news/press)
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Recording short movie to explain cornear research needs
Year(s) Of Engagement Activity 2015
URL http://www.bbc.co.uk/programmes/p01xkzjb
 
Description Visit to Charity Fundraising committee 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact First meeting; 12 people attended, including staff and volunteers of Fight for Sight Charity, who were presented with a short talk about the principle of our research followed by a interactive discussion
Second meeting: large group of participants in combination of information and fund raising event

Have since then regular contacts with charity staff
Year(s) Of Engagement Activity 2009,2012