Vinculin: a key to deciphering mechanotransduction

Cells have a sense of touch, and this research proposal aims to discover the underlying molecular machinery that allows cells to respond to mechanical forces. For example, as muscles enlarge and are able to contract with greater force, it is critical that the attachments of the ends of the muscles to the tendon/tendon matrix also become strengthened. We know that this is achieved by a mechanosensitive attachment machinery, but we dont yet know how this machinery works. We therefore have focused our research on a protein, vinculin, that we think will provide the "Rosetta Stone" of mechanotransduction, by providing a key that will reveal the underlying mechanisms. This is because vinculin has the exceptional property of becoming concentrated at different sites within the cell, when these sites are being pulled on by contractile forces. These sites includes the places where cells attach to other cells or to the scaffolding that surrounds cells, the extracellular matrix. The recruitment of vinculin to these sites of adhesion when they are under force helps to strengthen them, but it is not known how vinculin does that.

Vinculin exists in an "off" state, where it is tightly curled up, and an uncurled "on" state where it can bind to other proteins. Having too much vinculin in the on state in fact causes more problems to the cell than if vinculin is removed from the cell, but again we dont yet understand why this is the case. Our goal is therefore to discover:
1) how vinculin is recruited in response to force.
2) how vinculin functions, and what other proteins may be providing a similar function, such that the removal of vinculin can be tolerated surprisingly well.
3) how having too much "on" vinculin causes problems to the organism.

Our discoveries will impact on human health in multiple ways. Some human diseases result from the weakening of cell adhesion, which could be improved by developing methods to mimic the force signal, thus strengthening adhesion. Similarly, movement of cancer cells, or metastasis, renders cancers much more difficult to treat, and strengthening adhesion will restrain cell movement. Furthermore, understanding the molecular details of mechanotransduction may lead to the design of new nanomachines that can respond to force with local drug delivery. While our goal is to understand how mechanotransduction works in humans, we can answer these questions most effectively by using the experimental advantages of the model organism Drosophila melanogaster, the fruit fly. The proteins involved with vinculin function are also found in Drosophila, and we can use the sophisticated molecular genetics of Drosophila, combined with state of the art microscopy to use vinculin to discover how cells sense and respond to mechanical force.

Mechanotransduction pathways are emerging as central to cellular behaviour. We will use the protein vinculin as the key to discovering mechanisms of mechanotransduction at sites of cell adhesion. Vinculin is a complex molecule, with closed and open conformational states that alter its ability to bind many protein partners. Vinculin is a mechanoeffector, as it is recruited by mechanosensors at both cell-ECM and cell-cell junctions in response to acto-myosin force, but it is also a mechanosensor, as force opens up vinculin to permit new protein interactions. Mutations that open up the vinculin conformation cause strong effects within cells and lethality in the organism. The proposed research will use the experimental advantages of Drosophila to examine events upstream and downstream of vinculin. We will characterise the multiple force-dependent pathways of vinculin recruitment, using complementary approaches testing whether other integrin-associated proteins are necessary or sufficient to recruit vinculin in its different conformational states. We will discover how vinculin functions within the animal by 1) isolating mutations in genes encoding proteins that compensate for the lack of vinculin, 2) using cell biological approaches to elucidate the pathway by which overactive vinculin mediates its dramatic effects, and 3) using proteomic approaches to identify proteins recruited by vinculin in its open conformation. We will integrate the data from these approaches to generate an integrated model of how vinculin contributes to mechanotransduction pathways. The new knowledge of vinculin function will permit us to design effective assays to screen for molecules that can modify vinculin function in a clinical setting, to increase or decrease cell adhesion, and will provide a valuable paradigm for force-dependent nanomachines.

The beneficiaries of this project will be:

1. Pharmaceutical and Biotech industries. The role of Vinculin as a key mechanosensor, paired with the increasing recognition of the importance of mechanotransduction in disease, means that our work will be relevant for Pharmaceutical and Biotech industries. In particular, they will potentially be interested in using our findings to develop new drugs and technologies for the patients described below (2. and 4.). The better understanding of mechanosensing we will get from this project could lead to the design of nanoparticles that sense and respond to mechanical force. We will contact the Cambridge Nanoscience Centre to develop such technologies, benefiting the economy of the United Kingdom and strengthen our position as a world leader in drug development. Timescale 5-10 years.

2. Patients suffering from invasive cancers. A vast majority of cancer deaths are caused by metastasis, the process by which cancer cells spread within the body. The migration and invasive behaviour of cancer cells is critically regulated by mechanotransduction. We believe that our work will lead to a better understanding of how vinculin could be used as a tool or target to adversely affect the metastatic capabilities of cancer cells. For example, as we plan to identify proteins and mechanisms regulating vinculin function, these novel factors could be used to develop new drugs designed to reduce or disrupt the metastatic capability of cancer cells. Timescale 10-15 years.

3. Medical diagnosis of cancers. The identification of novel regulators and mechanisms of action for vinculin function may help to design biomarkers aimed at identifying cancerous cells, because changes to these may precede the invasive behaviour of certain cell types that mis-regulate vinculin expression. As cancers are predominantly diagnosed in older adults, better diagnostics and treatment could contribute to healthy aging and allow sufferers to live and remain active longer, increasing in turn the quality of the nation's health. Timescale 5-10 years.

4. Patients suffering from genetic disorders that affect cell adhesion, such as muscle dystrophies or the Kindler syndrome. We aim to understand how vinculin is regulated in live cells and this could for example lead to the identification of small molecules that activate vinculin and therefore strengthen cell adhesions. New drugs would therefore be developed and applied to strengthen the cell adhesions of patients with such disorders. Timescale 10-15 years.

5. Patients suffering from neurodegenerative diseases, such as Alzheimer's or Parkinson's disease, which are associated with the presence of protein aggregations in the affected tissues. Our proposed project aims to elucidate the mechanism by which constitutively open vinculin forms aggregates within cells. Therefore, such an understanding could be used to develop drugs preventing protein aggregation. Timescale 10-15 years.

6. Organisations and Companies recruiting scientifically trained staff, including both public and private sectors. The two postdoctoral researchers funded by this work will develop their training and expertise, as well as supervising A-level and undergraduate students. Thus the work will benefit a new generation of scientists. After the completion of the work, the postdoctoral researchers will be able to contribute to the scientific economy of the United Kingdom by applying the skills gained in the project, whether in public or private sectors.

7. The general public. Through engagement with the public through talks, websites, and general audience publications we seek to communicate the excitement and beauty of scientific research. Our work will involve a substantial amount of compelling images and movies that serve as an important starting point for public engagement with biomedicine. We will submit such images to competitions (e.g. Nikon/Wellcome Trust) to reach the widest audience.