Building epithelial cytoarchitecture:regulation of actin dynamics by cell-cell junctions
Lead Research Organisation:
Imperial College London
Department Name: Div of Biomedical Sciences
Abstract
Cell-cell adhesion and migration are key events in the life of many different cell types, and are essential for the organization of cells into higher ordered structures such as blood vessels, lung, skin, muscle, etc. Epithelial cells differentiate and form sheets of tightly packed cells: these sheets form protective layers wrapping up the body (skin) and the inside of internal cavities (i.e. stomach, intestines, etc). The choice of whether to migrate or stick to their neighbours has implications for epithelial function in health and disease. Thus, it is important to understand how a cell is kept inside an epithelial sheet or detaches and become shapeless, homeless and aimless. Epithelial cells can not function properly in the absence of tight cell-cell adhesion and a cuboidal morphology. We are interested in understanding how cells glue to their neighbours in order to regulate their shape and differentiation. We aim at identifying the molecules that participate in these processes and investigate their role in epithelia. Insights into how epithelial cell shape is established and maintained will provide clues on how to ameliorate diseases that have compromised epithelial function.
Technical Summary
The acquisition of a cuboidal morphology (polarization) is a hallmark of epithelia and occurs after cell division, wound closure, and during morphogenesis. Extensive reorganization of epithelial actin cytoskeleton is triggered following assembly of cadherin-dependent cell-cell adhesion. This cytoskeleton remodelling is essential for the functionality of epithelial sheets, stability of adhesive receptors at junctions and for the appropriate localization of signalling complexes. Yet, in spite of the importance of epithelia in development, homeostasis and diseases, the mechanisms via which a cuboidal morphology is acquired are poorly understood. Understanding how actin structures are assembled following cell-cell adhesion will provide major insights into the relevant signalling pathways for polarization. We have found that when cell-cell contacts are induced in keratinocytes, an increase in actin polymerization and myosin function (contractility) is observed very quickly. These processes are coordinated spatially and temporally into two distinct actin populations: junctional-actin and thin bundles. Here we aim to identify specific cytoskeletal proteins that are modulated by cell-cell adhesion and participate in the formation and/or maintenance of junctional-actin and thin bundles. As strategy, we will deplete the protein levels of specific proteins using siRNA during induction of cell-cell contacts and perform a screening using a siRNA library of actin binding proteins. Once relevant proteins are identified, we will perform functional assays to determine in which step of epithelial polarization these actin binding proteins operate: stabilization of cell-cell contacts or cell height increase (which reflects the functions of junctional-actin and thin bundles). We will use a combination of dynamic studies (live imaging), biochemical, biophysical and cell biology experiments together with rescue analysis of the specific phenotypes induced by siRNA depletion.
Publications
Kelly D
(2015)
Automated multiwell fluorescence lifetime imaging for Förster resonance energy transfer assays and high content analysis
in Analytical Methods
Schultz F
(2019)
Cardiomyocyte-myofibroblast contact dynamism is modulated by connexin-43.
in FASEB journal : official publication of the Federation of American Societies for Experimental Biology
V. M.M. Braga
(2017)
Cell-cell junctions
Erasmus JC
(2015)
Cooperation of distinct Rac-dependent pathways to stabilise E-cadherin adhesion.
in Cellular signalling
Erasmus JC
(2017)
Corrigendum: Defining functional interactions during biogenesis of epithelial junctions.
in Nature communications
McCormack J
(2013)
Cycling around cell-cell adhesion with Rho GTPase regulators.
in Journal of cell science
Erasmus JC
(2016)
Defining functional interactions during biogenesis of epithelial junctions.
in Nature communications
Braga VM
(2018)
Editorial Overview: Integration of dynamic processes in cell behaviour and tissue architecture.
in Current opinion in cell biology
Sri-Ranjan K
(2022)
Intrinsic cell rheology drives junction maturation.
in Nature communications
Brezovjakova H
(2019)
Junction Mapper is a novel computer vision tool to decipher cell-cell contact phenotypes.
in eLife
Tzircotis G
(2011)
RhoG is required for both Fc?R- and CR3-mediated phagocytosis.
in Journal of cell science
Kalaji R
(2012)
ROCK1 and ROCK2 regulate epithelial polarisation and geometric cell shape.
in Biology of the cell
Machesky L
(2017)
So far, yet so close: a-Catenin dimers help migrating cells get together.
in The Journal of cell biology
Braga V
(2016)
Spatial integration of E-cadherin adhesion, signalling and the epithelial cytoskeleton.
in Current opinion in cell biology
Keraudren, K.
(2011)
Two-step watershed segmentation of epithelial cells
Fischer A
(2018)
Vascular Permeability: Flow-Mediated, Non-canonical Notch Signalling Promotes Barrier Integrity
in Current Biology
Description | We designed and optimized a methodology to identify regulators of cell-cell contacts and thus understand how cells stick to each other and how they regulate how strong their bond should be. |
Exploitation Route | The methodology developed in this grant may be of interest for screening of compounds that inhibit junction dissociation. It would be interesting for basic researchers, drug development and pharmaceutical companies. In addition, we developed an automated software to quantify distinct junction phenotypes. |
Sectors | Healthcare Pharmaceuticals and Medical Biotechnology |
Description | BHF Centre for Research Excellence |
Amount | £40,000 (GBP) |
Organisation | British Heart Foundation (BHF) |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 11/2009 |
End | 10/2010 |
Description | Cross-talk between Ajuba and Rac signalling in the stabilization of cadherin adhesion |
Amount | £435,943 (GBP) |
Funding ID | MR/J007668/1 |
Organisation | Medical Research Council (MRC) |
Sector | Public |
Country | United Kingdom |
Start | 03/2012 |
End | 12/2015 |
Description | Multi-user equipment |
Amount | £784,699 (GBP) |
Funding ID | WT066143 |
Organisation | Wellcome Trust |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 01/2015 |
End | 12/2020 |
Description | Newton Fund (MRC-CONFAP) |
Amount | £84,730 (GBP) |
Funding ID | MR/M026310/1 |
Organisation | Medical Research Council (MRC) |
Sector | Public |
Country | United Kingdom |
Start | 01/2016 |
End | 12/2016 |
Description | Science without Borders |
Amount | £203,427 (GBP) |
Funding ID | 4803318142983356 |
Organisation | National Council for Scientific and Technological Development (CNPq) |
Sector | Public |
Country | Brazil |
Start | 02/2014 |
End | 01/2016 |
Title | Development of Machine Learning methods to quantify junction phenotypes driven by different stimuli (2017) |
Description | A major road block in the analysis of junction phenotypes is the ability to recognize cell borders in a monolayer of epithelial cells, which is painstakinly done manually and with enorumous time invested in each image. Such detailed analysis is not feasible for high-throughput analysis in screens. Training datasets teach computer how to identify cell-cell contacts, which can then tbe deployed in large datasets with minimal input from user. Development of the program was done using a large dataset of images obtained under different conditions obtained via the distinct grants funding my research. |
Type Of Material | Technology assay or reagent |
Year Produced | 2017 |
Provided To Others? | No |
Impact | Methodology is currenly being validated. It will be made available after thorough optimization, validation and adding a user-friendly interface. |
Title | high-throughput image analysis of junctions |
Description | A pipeline for automated quantification of phenotypes on cell-cell contacts as published in our paper: Erasmus, J, Bruche, S; Pizarro, L; Maimari, N; Pogglioli, T; Wheeler, A; Lees, J; Tomlinson, C; Zalivina, I; Alberts, A; Russo, A & Braga, VMM. Defining functional interactions during biogenesis of epithelial junctions. Nature Commun. 7, 13542, 2016 |
Type Of Material | Technology assay or reagent |
Year Produced | 2016 |
Provided To Others? | Yes |
Impact | N/A methodology is available online. |
URL | http://www.imperial.ac.uk/bioinformatics-data-science-group/resources/software/rnaiscreener/ |
Description | Defining the role of MTSS1 in epithelial cell-cell contacts |
Organisation | Beatson Institute for Cancer Research |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | This study identified the mechanism via which MTSS1 stabilizes cell-cell adhesion via modulation of actin filaments. |
Start Year | 2010 |
Description | Sean Sun - Modelling of biophysical parameters that generate a mature cell-cell contact |
Organisation | Johns Hopkins University |
Country | United States |
Sector | Academic/University |
PI Contribution | We provided the biological question and image datasets. |
Collaborator Contribution | Sean Sun - Hopkins Universitymodeller |
Impact | Manuscript submitted: Ranjan, K.; Alonso-Mardones, JS; Swiatlowska, P; Liu, S.;Rothery, S.; M. Stevens; Sun,S.; Gorelik, J & Braga, VMM. Cell confinement impacts on cortical elasticity, configuration and dynamics of junctions |
Start Year | 2015 |
Description | 51. 3rd World Congress on Cancer Biology and Immunology-2019", Italy KEYNOTE SPEAKER |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | Seminar at conference |
Year(s) Of Engagement Activity | 2019 |
Description | Invited Speaker Valbonne France |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Type Of Presentation | Keynote/Invited Speaker |
Geographic Reach | International |
Primary Audience | Other academic audiences (collaborators, peers etc.) |
Results and Impact | Invited to give a seminar at the Institut de Pharmacologie Moléculaire et Cellulaire, Valbonne France networking with academic staff from department |
Year(s) Of Engagement Activity | 2012 |
Description | Keynote speaker Symposium Epithelial intercellular junctions as dynamic hubs to integrate forces, signals and cell behaviour". Aachen University |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Undergraduate students |
Results and Impact | conference Keynote speaker |
Year(s) Of Engagement Activity | 2016,2017 |
Description | Lecture "Pioneers in Cell Dynamics and Imaging", Cells in Motion Clusters of Excellence, University of Muenster, Germany. 2018 |
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 | networking |
Year(s) Of Engagement Activity | 2018 |
Description | Mechanical Forces in Biology, EMBL Symposia, Heidelberg - Germany. 2017 |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | networking, data dissemination |
Year(s) Of Engagement Activity | 2017 |
Description | Physics and Biology confront cell-cell adhesion" Aussois, France October 14-19th |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | Seminar presented at conference - networking and recognition |
Year(s) Of Engagement Activity | 2019 |
Description | Summer Global School at Imperial College |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | Yes |
Geographic Reach | International |
Primary Audience | Schools |
Results and Impact | Together with Dr Anita Hall (Faculty of Natural Sciences) we were the leaders for the FoNS and FoM Stream Module. This stream aimed to provide students with insights and hands-on experience of cell biology experiments and their impact in medical research and drug development, cancer research in particular. All members of the Braga laboratory were involved in preparing cell samples to understand tumour metastasis and imaging techniques. The activities can be seen in the video https://spectrecom.wistia.com/medias/oipf0gc1je Consolidate and develop current knowledge of science, engineering, technology and maths Improve study skills whilst working in a range of stimulating environments Develop practical skills as well as theoretical knowledge Work in teams to develop interpersonal and organisational abilities Find out about what university life is like |
Year(s) Of Engagement Activity | 2015 |
URL | https://www.imperial.ac.uk/be-inspired/global-summer-school/ |
Description | Talk at Brazilian Embassy Networking event |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Other audiences |
Results and Impact | Workshop "Innovation in Healthcare" - discussion on engagement and collaborations - how is it best fomented? |
Year(s) Of Engagement Activity | 2017 |
Description | Workshop Women in Science |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | Women Scientists discussing their career progression in Academia, what worked well, what could have been different |
Year(s) Of Engagement Activity | 2009 |
Description | invited speaker Liverpool |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Type Of Presentation | Keynote/Invited Speaker |
Geographic Reach | National |
Primary Audience | Other academic audiences (collaborators, peers etc.) |
Results and Impact | invited to give a seminar at the Dept Cellular and Molecular Physiology, Liverpool, UK networking with colleagues at the Dept. |
Year(s) Of Engagement Activity | 2012 |
Description | talk conference Mechanobiology Institute Singapore, 10th Anniversary |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | networking, Science discussions, collaborations |
Year(s) Of Engagement Activity | 2018 |