Adhesion and cancer
Aims in short
Cancer progression involves uncontrolled proliferation and motility of cells. Integrins, transmembrane cell surface adhesion receptors, are proteins known to regulate cell behaviour by transducing extracellular signals to cytoplasmic protein complexes. At VTT we have performed genome-wide screens to identify integrin-binding intracellular proteins and demonstrated that recruitment of specific protein complexes by the cytoplasmic domains of integrins is important in tumorigenesis. Our aim is to extend our studies on the novel integrin binding proteins to gain insight on the diverse biological roles of integrins in cancer
Invasive and metastatic behaviour of malignant cells is the major cause of mortality in all cancer patients. Migration of cancer cells is critically regulated by physical adhesion of cells to each other and to their non-cellular surroundings (i.e. extracellular matrix). Cell-matrix interactions are mediated by a class of proteins called the integrins. Adhesion dependent migration in tissue is a prerequisites for lymphatic or hematogenous cancer cell dissemination. Epithelial tissue is the source of more than 80% of human cancers and epithelial-to-mesenchymal transition (EMT), where epithelial cells loose polarization is an important event during tumor progression and metastasis. Integrin mediated adhesion regulates in part epithelial cell polarity. The endo-exocytic traffic of integrins is important for directional cell motility of transformed cells. However, the molecular mechanisms regulating this process are still poorly characterized. In addition, it is unknown whether integrin traffic plays a role in the homeostasis of epithelial cells and if alterations in integrin traffic are necessary for cell transformation to occur.
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In the past few years, we have performed genome-wide screens to identify integrin-binding intracellular proteins. The novel protein-protein interactions studied thus far regulate receptor tyrosine kinase (RTK) signalling and cell proliferation as well as integrin traffic and motility of cancer cells. Recently, we have also shown that integrin endocytic traffic is critical for normal cytokinesis. This suggests that integrin traffic may play a role in maintenance of homeostasis of normal tissue.
In addition, we have recently studied in detail cell migration and EMT. We have shown that a novel interaction between a5 integrin and ZO-1 tight junction protein functions in the lamellipodia of transformed cells to support cell migration and EMT in vitro and in vivo. We have also investigated the role of mesenchymal protein vimentin in EMT.
Our aim is to extend our studies on the identified integrin binding proteins to gain novel insight on the diverse and sometimes unexpected biological roles of integrins. The role of integrin traffic and intermediate filaments in the regulation cell architecture and transformation will be of particular interest. We aim to characterize the molecular mechanisms regulating integrin traffic in different cell types and to dissect in detail the role of Rab21-integrin association in this process. We also aim to use genome-wide siRNA screening to identify novel regulators of integrin function, relevant to metastasis.
Screening for novel integrin regulators
Integrins usually switch between an active and an inactive state. In the active state they are able to bind extracellular ligands like collagen or fibronectin, which leads to strengthened local cell adhesion. This state is induced by a propagation of conformational changes from the intracellular domains of integrins to the extracellular domains leading to an increase in affinity to the ligand. The proposed final step in this activation process is the binding of talin to the cytoplasmic tail of beta-integrin (beta-cyt). However, kindlin proteins, mutated in the Kindler syndrome, were recently shown to activate integrins by binding to beta-cyt similarly to talin. Furthermore, it was shown that the initial integrin-driven steps of cell spreading are independent of talin. These results indicate that there have to be additional proteins regulating integrin activity.
Existing literature suggests an important role for integrins in cancer metastasis and especially in the process of extravasation, which include arrest to endothelium, migration through it, as well as invasive growth in the metastatic foci. Therefore, we aim to identify novel regulators of integrin activity that are likely to regulate cancer spread. This project will utilize a live cell microarray platform developed at VTT that enables reverse siRNA transfection and microscopy analysis of live cells within a three-dimensional matrix (see Figure). We have developed this novel, innovative siRNA and live cell-array to detect integrin activity in cells growing on matrigel. Up to 46,000 individual siRNA transfected cell samples can be screened for integrin activation, cell multiplication and changes in the cytoskeleton.