Principal Investigator Richard Hynes
Most cells adhere to their neighbors and to the extracellular matrix, a fibrillar meshwork surrounding or underlying most cells in the body. Cell adhesion plays important roles in the normal functions of cells, contributing to cellular organization and structure, proliferation and survival, metabolism and gene expression. During embryological development, cell adhesion is important for the correct movements of cells modeling the embryo. In the adult, appropriate cell adhesion is necessary for numerous physiological processes and can be deranged in many diseases, including thrombosis, inflammation and cancer.
The laboratory seeks to understand the proteins involved in cell adhesion and the way in which they control adhesion and migration of cells in both normal and pathological processes. Cell adhesion is mediated by several families of proteins, called adhesion receptors, specialized for adhesion between adjacent cells or between cells and the extracellular matrix. Adhesion receptors do much more for cells than merely sticking them down in the correct locations, although that, in itself, is important. They also form physical linkages between the extracellular environment and the internal structures of cells and thus control cell shape and motility. Adhesion receptors also act as two-way transducers of signals both into and out of cells. Therefore cells can control whether or not their adhesion receptors are functional; this is important to ensure appropriate cell adhesion. For example, blood platelets must adhere when a blood vessel is damaged in order to staunch bleeding – hemostasis – but must not adhere at the wrong time or place – that produces thrombosis. Similarly, leukocytes must adhere in appropriate places to fight infections but if they adhere at the wrong place or time, the result is inflammation. Alterations in cell adhesion also play important roles in the control of cell behavior during invasion and metastasis of malignant cancer cells and in angiogenesis. Thus, control of adhesion receptors is a matter of life and death. In their role as signal transducers into cells, adhesion receptors control cell proliferation, cell survival and the expression of specific genes. Our aim is to understand these processes at the molecular level.
One approach we use to decipher the roles of the proteins involved is to generate mice with mutations in the genes encoding them. In that way, we can discover which processes require specific adhesion proteins by "knocking out" or mutating the genes encoding them so that the resulting mice cannot make the proteins or make variant forms. We have used this approach to dissect the roles of various adhesion receptors in the recruitment of white blood cells to sites of inflammation and in the adhesion of platelets during hemostasis and thrombosis. We have also generated mouse models of human diseases affecting cell adhesion. For example, mice lacking the ÉøIIbÉ¿3 integrin exhibit bleeding just as do human patients with the same defect and we have shown that these mice also have defects in bone remodelling, because specialized bone cells (called osteoclasts) lack a related integrin, ÉøvÉ¿3, necessary for their function. Therefore, inhibition of the function of the ÉøvÉ¿3 integrin should ameliorate osteoporosis since that arises from overactivity of osteoclasts. Drugs blocking the function of platelet alpha-IIb-Beta3 integrin are already in use to reduce thrombosis after angioplasty.
Many of the mutations produce defects in the development of new blood vessels, or angiogenesis, which involves multiple cell adhesion and migration events. Our work has shown particularly important roles for the extracellular matrix proteins, fibronectins, and cell surface receptors for them. Fibronectins comprise a group of closely related proteins all encoded by a single gene and they promote cell adhesion and cell migration and affect many other cellular processes. Cells recognize fibronectins using cell surface integrin receptors. There are about two dozen different integrins. They are the major receptors for extracellular matrix and some integrins also participate in cell-cell adhesion; their extracellular portions recognize specific binding sites in proteins such as fibronectins. Their intracellular portions bind to cytoplasmic proteins, including both structural proteins of the cytoskeleton (which can be viewed as the “bones and muscles” of cells) and signaling proteins, which send messages into the cell affecting cell behavior. Thus, integrins serve as transmembrane linkers between the extracellular matrix outside and the cytoskeleton and signaling systems inside cells. Both fibronectin and several integrin receptors are essential for angiogenesis. We are currently analyzing the relative contributions of different splice isoforms of fibronectin and different integrins to vessel development. Some integrins have been suggested as targets for antiangiogenic drugs and it is important to determine which ones are the most crucially involved. Our work has lead to new interpretations concerning the efficacy of certain candidate antiangiogenic drugs.
Another example of the potential for novel anti-adhesive therapeutics comes from our research on other cell adhesion receptors, called selectins. These proteins mediate adhesion between circulating white blood cells or platelets and the walls of blood vessels. They, along with integrins, play important roles in ensuring that white blood cells circulate to their correct locations in the body and home on sites of infection or inflammation. We have made mice mutated in all possible permutations of their selectin genes. These mice are alive but have defects in their ability to recruit white blood cells. We have used these selectin-deficient mice to study the roles of selectins in various inflammatory responses.
We are currently applying our understanding of cell adhesion to analyses of cancer. Numerous steps in the progression of cancer, importantly including invasion and metastasis, involve altered adhesive properties of cells. Invasion and metastasis are what make tumors malignant and it would be invaluable to acquire a better understanding of these events. As one example, human carcinomas that express ligands for selectins have a poorer prognosis than do those that lack those ligands. This suggests that the malignant cells may use selectins during their progression. We have investigated this question, using our selectin-deficient mice, and find that selectins do contribute to metastatic spread of tumor cells expressing selectin ligands. We also use DNA arrays, which allow screening for large numbers of genes, to search for genes selectively expressed in metastatic cells. We have found that several genes involved in extracellular matrix assembly and in organization of the cytoskeleton are reproducibly upregulated and we went on to show that one of them, rhoC, is essential for metastasis. We are continuing with this approach to uncover other alterations contributing to invasion and metastasis. Recently, we have shown that a particular atypical G-protein-coupled receptor acts as a suppressor of tumor progression, apparently by acting as a receptor for suppressive signals from the tumor microenvironment. Now that the complete genomic sequences of humans and mice are available, it is possible to determine exactly how many genes contribute to cell adhesion – our current estimates are 2000-3000. Using approaches such as DNA arrays and proteomics we are investigating which of these genes are altered in their expression in different steps of invasion and metastasis. We are also interested in the large expansion of this complement of adhesion-related genes, the “adhesome,” during vertebrate evolution. Therefore, we are interested in analyzing the genomes of the deuterostome lineage leading to vertebrates and are involved in annotation of several recently completed non-vertebrate genomes from the deuterostome lineage. This analysis should lead to formulation of hypotheses as to what changes in cell adhesion and extracellular matrix were associated with particular evolutionary advances.
Work on adhesion molecules in intact animals and by genome-level approaches is complemented by studies in cell culture and with purified proteins, allowing more detailed analysis of the specific binding interactions between fibronectins and integrins and between integrins and their cytoskeletal connections. This strategy offers great promise for the design of drugs to block adhesion in the intact organism. In that way, one can hope to combat pathological processes involving adhesion such as thrombosis, inflammation, angiogenesis and cancer. Mouse models help to refine these strategies by showing which molecules are most important for particular processes.