Vinculin and associated signalling networks in the regulation of cell motility
Lead Research Organisation:
University of Manchester
Department Name: Life Sciences
Abstract
The process whereby cells in the body move from place to place is called cell migration. Cell migration is fundamental for the development of organisms, wound healing, and the immune respose in the body. All these physiological processes are regulated by the interaction of the cell with its environment. The major group of molecules controlling these interactions are cell surface proteins called integrins. Integrins span the cell membrane and are able to attach with their extracellular part to proteins located in the outside surrounding of the cell (extracellular matrix). With their intracellular part, integrins associate with proteins that link them to the cells' highly flexible and dynamic skeleton (actin cytoskeleton). The interaction of integrins with the cytoskeleton is indirect. One of the key molecules, which control the link between integrins and the actin cytoskeleton, is named 'vinculin'. Mice lacking this protein die early during development, and cells deficient in vinculin are highly metastatic. Vinculin is able to interact with 11 other proteins, which have the potential to exert important signals regulating the cell communication with its environment. My aim is to understand how vinculin coordinates the numerous signals and thus modulates the communications between the cell and its outside surroundings. We will explore the function of vinculin by visualizing and monitoring its dynamic behaviour and interactions with other proteins in live cells using advanced imaging techniques. We will genetically modify vinculin and its ability to interact with its potential binding partners and thus test how vinculin controls a network of signals that is important for coordinated cell migration. For coordinated cell movements, the cell needs to exert pulling forces on the extracellular environment. Our recent discoveries suggest that vinculin interactions with the contractile skeleton in the cell are essential for the transmission of forces to the integrins that bind with its extracellular part to the extracellular matrix. Using highly developed nanotechnological devices, we will determine how vinculin signals contribute to such force transmission from the interior to the outside of the cell. Using devices that will mimic forces in the cells outer environment (such as pressure, stretching and shear forces in the body), we will be able to test how vinculin transfers signals from the outside of the cell to the inside cytoskeleton. The outcome of this study will provide us with important information for the better understanding of how cell motility is regulated in health and disease.
Technical Summary
The primary sites for cell adhesion to the substrate are focal adhesions (FA). At FAs, over 100 molecules regulate the link of integrins to the actomyosin force machinery. FAs are therefore molecular entities that regulate cell motility as well as the transmission of forces to the substrate, and vice versa. Here I seek support to define and integrate the contribution of vinculin, a central player for focal adhesion regulation. Vinculin in its active state has the potential to bind 11 other FA components, which in turn have multiple binding partners. With its prominent binding sites for talin and actin, vinculin is located at the interface between adhesion receptors (integrins) and the actin cytoskeleton and thus in an ideal position to coordinate a network of signals regulating cell motility. I aim to understand how vinculin and its multiple interaction partners (a) control cell adhesion and motility and (b) how they are involved in the transmission of forces. To date, protein interaction studies in FAs have relied mostly on biochemical analyses, which are unable to probe the enormously complex and dynamic events that occur in cells. We will capitalise on my knowledge on advanced fluorescence microscopy, including fluorescence recovery after photobleaching (FRAP), quantitative image correlation analysis (ICA), and fluorescence resonance energy transfer (FRET), as an essential tool for overcoming some of these barriers. We will complement these techniques with biochemisty and (nano-) technological devices that allow application and measurement of forces to elucidate: 1) How does vinculin control the large network of signals in focal adhesions? 2) How do interaction partners of vinculin vary in time and space? How is vinculin involved in sensing of extracellular forces, the generation of intracellular force, and the regulation of adhesion and migration?
Organisations
People |
ORCID iD |
Christoph Ballestrem (Principal Investigator) |
Publications
Goldyn AM
(2009)
Force-induced cell polarisation is linked to RhoA-driven microtubule-independent focal-adhesion sliding.
in Journal of cell science
Askari JA
(2010)
Focal adhesions are sites of integrin extension.
in The Journal of cell biology
Rooney C
(2010)
The Rac activator STEF (Tiam2) regulates cell migration by microtubule-mediated focal adhesion disassembly.
in EMBO reports
Goldyn AM
(2010)
The kinetics of force-induced cell reorganization depend on microtubules and actin.
in Cytoskeleton (Hoboken, N.J.)
Wang P
(2011)
The C terminus of talin links integrins to cell cycle progression.
in The Journal of cell biology
Stroud MJ
(2011)
Characterization of G2L3 (GAS2-like 3), a new microtubule- and actin-binding protein related to spectraplakins.
in The Journal of biological chemistry
Carisey A
(2011)
Fluorescence recovery after photobleaching.
in Methods in molecular biology (Clifton, N.J.)
Carisey A
(2011)
Vinculin, an adapter protein in control of cell adhesion signalling.
in European journal of cell biology
Zhao X
(2012)
Multi-layer phase analysis: quantifying the elastic properties of soft tissues and live cells with ultra-high-frequency scanning acoustic microscopy.
in IEEE transactions on ultrasonics, ferroelectrics, and frequency control
Morgan MR
(2013)
Syndecan-4 phosphorylation is a control point for integrin recycling.
in Developmental cell
Goult BT
(2013)
RIAM and vinculin binding to talin are mutually exclusive and regulate adhesion assembly and turnover.
in The Journal of biological chemistry
Carisey A
(2013)
Vinculin regulates the recruitment and release of core focal adhesion proteins in a force-dependent manner.
in Current biology : CB
Nijenhuis N
(2014)
Combining AFM and acoustic probes to reveal changes in the elastic stiffness tensor of living cells.
in Biophysical journal
Stroud MJ
(2014)
GAS2-like proteins mediate communication between microtubules and actin through interactions with end-binding proteins.
in Journal of cell science
Jansen KA
(2017)
Mechanotransduction at the cell-matrix interface.
in Seminars in cell & developmental biology
Nazgiewicz A
(2019)
GAS2-like 1 coordinates cell division through its association with end-binding proteins.
in Scientific reports
Title | Arts exhibition "Immortality II" |
Description | Participating with images of cells which were screen printed in collaboration with Sally Gilford at an arts gallery exhibition called "Immortality II" |
Type Of Art | Artistic/Creative Exhibition |
Year Produced | 2016 |
Impact | knowledge to the general public audience; stimulating thoughts about health and disease; artistic value; |
Title | Arts exhibition "Immortality" |
Description | Artistic prints or scientific images. |
Type Of Art | Artistic/Creative Exhibition |
Year Produced | 2015 |
Impact | Engaging the public; see movie on provided link with participation of C.Ballestrem. |
Title | Images of the micro world of cells. |
Description | Images were provided for an arts exhibition |
Type Of Art | Image |
Year Produced | 2010 |
Impact | Informing and stimulating thoughts about the role of cells and their ability to communicate with their environment in health and disease. |
Description | Our main aim was to deepen the understanding of how cells sense their environment, which enables them to react to mechanical and biochemical signals. Our data demonstrate that vinculin, a core protein of focal adhesions - the primary interaction sites of cells with their extracellular environment-, controls the link between adhesion receptors (integrins) and the actin cytoskeleton. Our data indicate that vinculin regulates coupling and uncoupling events between integrins and the actin cytoskeleton as well as the recruitment and release of focal adhesion components; both are required for coordinated cell migration. Our main achievements are detailed below. - We identified vinculin as a major coordinator of stretch induced signaling. We followed changes of intracellular structures such as actin, microtubules, and cell matrix interactions (focal adhesions, FA) in time and space and showed that mechanical forces activated signals that in conjunction with the actin cytoskeleton lead to force induced reorganization of cells and cell-matrix adhesion sites (Goldyn et al, JCS 2009; Goldyn et al, Cytoskeleton 2010). We then revealed that without vinculin the response of cells to forces is perturbed and the binding of vinculin to the actin cytoskeleton is important for its function in transducing mechanical signals (Carisey et al., JCB, manuscript under review). - Vinculin coordinates cell adhesion and migration through its activation of the talin/integrin complex. Using Atomic force spectroscopy to measure adhesion we show that vinculin through its interaction with talin regulates the activation status of integrins. Cells expressing constitutively active forms of vinculin adhere stronger and thereby inhibit cell migration (Carisey et al., Current Biology, 2013). - Vinculin controls the recruitment and the release of FA proteins. A variety of mutant forms of vinculin have enabled us to identify which proteins are recruited though direct binding or indirect signals to FAs (Carisey et al., Current Biology 2013). The interaction of vinculin with talin was key for vinculin recruitment itself (Wang et al, JCB 2011) and key for its downstream action in FAs. We identified which of thee clusters of binding sites for vinculin on talin are crucial for vinculin function (manuscript in preparation). - We elucidated the role of vinculin in regulating the dynamcis of a network of focal adhesion proteins. The dramatic impact of active vinculin on the dynamic turnover of its binding partner talin depended largely on the presence of two vinculin interaction sites. (Carisey et al. Methods Mol Biol. 2011; manuscript in preparation). - We determined the relationship of vinculin with paxillin and showed the differential interaction of vinculin with paxillin or Hic5 depending on the balance between Rac1 and RhoA activity (Deakin et al, PLOS one 2012). - We elucidated how vinculin regulates cellular elasticity a parameter that has been show be an important parameter to detect cancerous cells; cancer cells are softer than normal cells. Cells without vinculin or cells with vinulin lacking the link to the actin cytoskeleton are softer. (Nijenhuis et al, Biohysical Journal 2014). |
Exploitation Route | Academically: We revealed key aspects of how cells sense their environments and respond to extracellular mechanical and biochemical signals. These signals determine whether a cell migrates, stays in a resting phase or proliferates. We now build on our discoveries by exploring further molecular details that will enable us understand: 1) how the sensing machinery is regulating neuronal behaviour and their sensing of mechanical properties of their extracellular environment (grant proposal submitted to BBSRC Sept 2014). 2) how the dynamic system of molecular activation and deactivation processes control cell adhesion and migration (grant application to BBSRC for submission 2014in preparation). 3) how we can interfere with protein-protein interactions in order to manipulate cell adhesion and motility. For this we have set up collaborations that will lead to a multidisciplinary grant proposal involving the structural determination of the vinculin binding partner talin, the functional analysis of talin binding to its partners in cells, and the production of synthetic compounds which aim to modify molecular interactions thus enabling us to direct cell motility and proliferation. We hope that such cell permeable synthetic compounds will allow us to direct cell function and in future may thus serve as drug leads. For education: One of the major uses is in making the public understanding of how the body, i.e. cells in the body function. Students of the lab therefore participate in "Wellcome to the matrix events" for invited schools (http://www.wtccmr.manchester.ac.uk/forthepublic/events/matrix/). Our images of dynamic cells are not only of scientific interest, but also beautiful. Therefore we selected images for presentation at the arts exhibition "All About Us" in February 2011 in Bristol. Another arts exhibition is currently planned for March 2015. |
Sectors | Education,Healthcare |
URL | http://www.ballestremlab.com |
Description | Findings attracted: (1) Health care sector: the company "Smith and Nephews" had contacted us to examine how the molecular mechanism we revealed are implicated in Ultrasound mediated wound healing. This has lead to BBSRC supported CASE funding (ongoing project) where we aim to reveal signalling events that will lead to data that can improve wound healing. Our findings have further attracted BBSRC CASE funding from Intelligent Imaging Innovations (3i), with whom we helped to improve fluorescence energy transfer readouts using advanced microscopy. (2) Education: We engage in school education (Wellcome to the Matrix) where pupils form different schools are invited to learn about the cells interaction of cells with the matrix. Our lab also hosts selected highly gifted pupils from high school. Both events are coordinated by Ceri Harrop in the Wellcome Trust Centre for Cell-Matrix Research of which the our group is member of. (3) Society: We participate in arts exhibitions where we provide images to reveal the microscopic beauty of life on one hand and on the other hand important aspects in health and disease. |
First Year Of Impact | 2005 |
Sector | Education,Culture, Heritage, Museums and Collections |
Impact Types | Cultural,Societal |
Description | BBSRC studentship with Smith and Nephews CASE contribution: The Role of Ultrasound in Wound Healing |
Amount | £160,000 (GBP) |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2012 |
End | 09/2016 |
Description | DTP studentship: Regulation of actin, microtubules and focal adhesions during cell division - a specific role for GAS2-like proteins |
Amount | £100,000 (GBP) |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2010 |
End | 09/2014 |
Description | DTP studentship: The Regulation of the Cell-Matrix Adhesion Network |
Amount | £100,000 (GBP) |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2013 |
End | 09/2017 |
Description | DTP studentship; Investigating the differential dynamics of focal adhesion proteins: a key role for vinculin in their coordination |
Amount | £70,000 (GBP) |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2009 |
End | 09/2012 |
Description | DTP/CASE studentship together with and Intelligent Imaging Innovations (3i): The Role of Forces in Integrin Activation and Focal Adhesion Composition |
Amount | £100,000 (GBP) |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 01/2010 |
End | 12/2013 |
Description | Wellcome Trust 4 Year Studentship: Functional Characterisation of a Novel Microtubuel-Actin Interacting Protein, GAS2-like3 |
Amount | £150,000 (GBP) |
Organisation | Wellcome Trust |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 09/2006 |
End | 09/2010 |
Description | Lab shadowing sessions |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | Two pupils were visiting our lab shadowing the lab members' project work. This gave them insight the lab environment, how scientists work and in the topics that are investigated in our laboratory. |
Year(s) Of Engagement Activity | 2016 |
Description | Manchester Science Festival |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | Yes |
Geographic Reach | Local |
Primary Audience | Public/other audiences |
Results and Impact | was informative stimulating for non-scientists and sparked questions and discussion. Sparked interests of the public. |
Year(s) Of Engagement Activity | 2010,2011,2013,2014 |
Description | Primary School science-art masterclass - "what makes us human" |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Schools |
Results and Impact | Alongside scientists, children were looking at biology through an artistic lens. Pupils investigated how to make beautiful patterns from the unexpected. They looked through microscopes to identify patterns in the "nature" (e.g. cells), compare it with patterns in the museum and produce their own pattern design to print on cloth. At the end patterns of life was discussed with the children; how humans and how cells are affected by their environment in health and disease. |
Year(s) Of Engagement Activity | 2016 |
URL | http://www.whitworth.manchester.ac.uk/learn/schoolscollegesanduniversities/primary/masterclass/ |
Description | School Visit (Withington High School for Girls), Manchester UK |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | Lesson about "How cells sense their environment and what happens if this goes wrong". Sparked questions and discussions and the school increased interest in related subject areas and regular visits. |
Year(s) Of Engagement Activity | 2015,2016 |
Description | School visits (different Manchester schools) |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | Yes |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | mentoring sparked questions and discussions. Further visits of schools in coming years. |
Year(s) Of Engagement Activity | 2010,2011,2013,2014 |
URL | http://www.wellcome-matrix.org/foreveryone/for-schools/wellcome-to-the-matrix.html |
Description | School visits abroad (Singapore, Indonesia and Malaysia) and participation and recruitment fairs for international undergraduate students. |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Schools |
Results and Impact | Several high school classes were visited with a selection of talks around the topic of "How cells sense their environment and the role in health and disease". Around 20-50 students visited per school visit (some with parents); 4 school visits in 2015 (Singapore only); 10 in 2016 (Singapore, Indonesia, Malaysia). Besides engagement, the talks about my labs research aimed to attract international undergraduate students to the University of Manchester. |
Year(s) Of Engagement Activity | 2015,2016 |