Kindlin and EGFR control convergent pathways to regulate epithelial cell function

This research relates to two proteins that we know are not correctly controlled in patients with skin blistering disease. These proteins are called Kindlin-1 and Epidermal Growth Factor Receptor (or EGFR). We already know that lack of Kindlin-1 in the skin leads to Kindler Syndrome - a rare genetic disorder that results in skin blistering and sometimes leads to skin cancer. We have also recently identified a new group of patients that have defective EGFR protein and this similarly results in fragile skin. Our recent research on these two proteins strongly suggests that the observed clinical defects in the patients are partly caused by similar processes going wrong. This defect is mainly in the ability of cells to stick, or 'adhere' to the proteins that surround them in the skin. Normal functioning of the adhesion process is critical to health, and if it goes wrong, the sort of diseases that might occur include skin fragility, psoriasis, cancer, reduced immunity and inflammation. The research we would like to do now involves analysing Kindlin-1 and EGFR function in more detail. We firstly want to know precisely how these proteins work together to contribute to normal skin function. We plan to use a number of biochemical and microscopy techniques to answer questions such as: do these two proteins bind together in cells; how does loss of one or the other upset the ability of cells to adhere to proteins around them, and how do the common 'targets' of this protein complex also control this process? In our research we would also like to create more complex models to allow us to study Kindlin-1 and EGFR function in an intact organism. The reason we want to do this is because we would like to find out how these proteins behave in a more 'real' setting where there are other cells and tissues that might alter how these proteins behave. We will use a very well characterized model of epidermal blistering in the Fruitfly for this purpose. These animals are much easier, cheaper and faster to work with than mice and have versions of all of the genes/proteins we are planning to study, so are ideal for our experiments. By combining the information we get from experiments in cells, human tissues and the Fruitfly model, we can also start to plan potential corrective treatments with genes, proteins and drugs, work that could lead to improvements for patients in the future. Such work could lead to less skin blistering and improved quality of life in this and other families that have defects in Kindlin-1 or EGFR. The work is expected to expand our knowledge of the fundamentally important basic science topic of cell adhesion to surrounding tissues as well as the physiological maintenance of healthy skin.

The goal of this project is to integrate the identification of new mutations in patients with skin blistering with a cell biological analysis of the defects in mammalian cells, and exploitation of the genetics of Drosophila to group components into novel pathways. Our serendipitous findings to date provide us with a unique opportunity to interrogate the co-operation and convergent molecular signatures of EGFR and kindlin-1-dependent skin disease. Our first goal is to confirm that Kindlin-1 regulation of EGFR is a key part of its unique function that cannot be substituted for by Kindlin-2. Second, we wish to identify the mechanism by which Kindlin-1 regulates EGFR levels. Third, we aim to identify the common mediators of targets of EGFR ad Kindlin-1 signalling that mediate the epithelial cell adhesion. These studies will characterise novel candidate genes and mechanisms that contribute to skin fragility disorders using a powerful combination of in vitro and in vivo models: human keratinocytes (from healthy volunteers and KS patients) and the genetically tractable organism Drosophila. We anticipate that the knowledge gained will improve our understanding of how changes in levels or activation of each of the pathway components gives rise to the disease pathology, suggests possible treatments, and aids in the identification of less serious skin conditions that may arise from a more mild impairment of these pathways.

Data arising from this study will be of significant interest to a wide range of cell and developmental biologists. The questions we are approaching in this study are of immediate relevance to researchers in the cell adhesion and migration field, and adhesion/growth factor receptor crosstalk is well recognized as being centrally important to a wide range of basic biological processes such as proliferation and migration. Adhesion and growth factor signalling are evolutionarily conserved events and therefore our data stand to benefit a wide collection of investigators in basic and translational research, including biologists, microscopists, immunologists, infectious disease specialists, neurobiologists, and oncologists. The Kindlin proteins are currently emerging as key players in other disease settings including cancer, immune dysfunction and wound healing. Moreover, EGFR is well-known to be dysregulated in many cancers so provision of novel information relating to potential upstream or downstream regulators will help inform on potential combined treatments in EGFR mutated disease settings. The outcomes will also be important to clinical dermatologists both from a diagnostic and treatment perspective, as unraveling these pathways provides a platform for future translational research including gene, protein, cell and drug therapies aimed at restoring function in patient skin and reducing the skin blistering.
The collaboration between basic cell and developmental biology and clinical researchers proposed here provides an exciting and powerful multidisciplinary team to dissect the full implications of Kindlin-EGFR signaling in the context of epithelial homeostasis and disease. The workplan offers excellent training opportunities for the new team members and provides a platform for open discussions between all three labs that will not only benefit the current study but likely lead to future collaborative projects together. Moreover, our novel discoveries and related studies are expected to open the door to new collaborations with both European and global investigative research teams over the next 1-3 years.
Outside the immediate academic beneficiaries, data obtained in this study will also be of benefit to the patient charitable organisation that assists individuals with inherited blistering skin diseases. This organisation is called the Dystrophic Epidermolysis Bullosa Research Association (DebRA) and it provides support for more than 5,000 people in the UK with different forms of genetic blistering diseases involving mutations in 14 different genes. The data arising from this study will also be of direct benefit to the NHS diagnostic laboratory for epidermolysis bullosa based at St Thomas' Hospital in London. This laboratory (The Robin Eady National Diagnostic Epidermolysis Bullosa Laboratory) will now be able to add additional diagnostic tests (immunohistochemistry for EGFR, FRMPD1, UNC93, mutation analysis of associated genes) to make its diagnostic services even more comprehensive and inclusive.