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Signaling by ErbB/HER Family Receptors in Breast Cancer Cell Membranes
Tumor Cell Biology II
Epidermal growth factor receptor (EGFR) family members (also termed ErbB family members) are often dramatically over-expressed in mammary tumor cells, which can enhance breast cancer growth and metastasis. Consequently, ErbB receptors are the targets for exciting new cancer therapeutics. Recent studies indicate that ErbB receptors reside within submicroscopic domains of the cellular membrane (caveolae, lipid rafts, or termed generally, membrane microdomains) having a unique composition. Various lipids and proteins identified in membrane microdomains are involved in growth factor receptor signaling, which suggests that these domains might constitute submicroscopic signaling platforms. This proposal hypothesizes that ErbB receptors are localized within breast cancer cell membrane microdomains that serve to coordinate their signaling activities. Our preliminary data indicate that ErbB receptors are indeed localized within microdomains in breast cancer cell membranes, and we seek to determine how this micro-localization impacts upon ErbB receptor function. The specific aims of the proposed research are to (1) characterize the spatial organization of ErbB receptors in breast cancer cell membranes and identify the microdomains in which they reside, and (2) determine how membrane microdomain localization coordinates specific ErbB receptor signaling events. These aims would be addressed via a combination of complementary experimental approaches. Biochemical approaches include the fractionation of cellular membranes by newer detergent-free methods, so that isolated membrane microdomains can be analyzed for the presence of ErbB receptors and specific signaling targets, and to determine in which membrane compartments specific signaling events occur. Alternatively, fluorescence microscopy will be used to visualize microdomains in the cellular membrane and the potential localization of ErbB receptors and their signaling targets within them. A high-resolution immuno-electron microscopy method will be used to image the spatial distribution of ErbB receptors on the membrane surface and to examine the potential colocalization of ErbB receptors with various membrane microdomain components and signaling molecules. The spatial organization of receptor signaling events within signaling microdomains likely represents the next level of complexity in receptor signal transduction. We expect through these studies to determine how ErbB receptors are localized in breast cancer cell membranes with submicron resolution, identify/characterize the microdomains in which ErbB receptors reside, and determine how specific signaling events are influenced by membrane microenvironment. The results of these studies would be of particular significance in the context of human breast cancer, as it is unclear how the aberrant expression of ErbB receptors perturbs their normal membrane localization, trafficking, and mitogenic signaling.
Frequently in breast cancer tumors, the individual cancer cells have unusually high numbers of cell surface proteins termed ErbB receptors. ErbB receptors are both markers for breast cancer diagnosis and the targets of new anti-cancer agents designed to recognize and bind to them. ErbB receptors also promote the growth and spread of breast cancer, making it important to understand how the high numbers of ErbB receptors in cancer cells stimulate their growth. ErbB receptors normally function as growth factor receptors, which send biochemical signals to the cell nucleus that trigger cell division. Recent studies suggest that ErbB receptors are localized in small microdomains on the membrane of the cell surface, which could be the sites in which these biochemical signals are initiated. We hypothesize that the micro-organization of ErbB receptors and biochemical signaling events on the cell membrane is perturbed in the case of breast cancer. In the proposed research, we would work to develop the biochemical and microscopy techniques necessary to study the micro-organization of ErbB receptors and biochemical signaling events in breast cancer cell membranes, to biochemically characterize the microdomains in which ErbB receptors reside, and to determine how the micro-organization of ErbB receptors impacts upon their signaling functions. Because the size of the membrane microdomains in which ErbB receptors are believed to reside is smaller the limit of resolution of conventional light microscopy, we propose to use a combination of biochemical, fluorescence microscopy, and electron microscopy techniques to characterize the micro-organization of ErbB receptors and receptor signaling events. Electron microscopy, the highest resolution of these methods, has the particular advantage in that with it we can image the micro-organization of individual ErbB receptor molecules on the cell surface. The findings of our proposed research would be important with respect to our understanding of breast cancer and in developing new treatments. New anti-cancer agents targeting ErbB receptors show much promise, but unfortunately do not work in many diagnosed cases. It is thus important to understand how ErbB receptors are localized on the cell membrane surface, and exactly how they function to control breast cancer cell growth.