Epithelial cells within glandular tissues, like the breast, are organized into ducts and specialized spherical structures containing a monolayer of luminal epithelial cells surrounding a hollow lumen. Maintenance of this specialized architecture and suppression of aberrant cell proliferation is mediated by controls that restrict the survival of cells which proliferate into the hollow lumen or outside of their normal niche. Our previous studies have provided evidence that a major factor contributing to death of cells outside the epithelial monolayer is lack of appropriate matrix attachment. In addition, we found that both apoptotic and non-apoptotic death processes are activated in cells deprived of their appropriate matrix. Survival of aberrant proliferating cells is dependent on escape from both of these death processes. In this proposal, we describe studies, using three different experimental models- 1) Suspended, matrix-deprived breast epithelial cells in culture, 2) 3D acinar structures cultured in reconstituted basement membrane proteins, and 3) Terminal end buds (TEBs) of pubertal mouse mammary glands in vivo. Using these model systems, we will elucidate the mechanisms apoptotic and non- apoptotic death of cells lacking proper matrix attachment. In addition, we will examine the mechanisms involved in oncogene-mediated escape from these death mechanisms using both biased pathway dissection studies and unbiased siRNA screens. Lastly, we will examine the importance of one proapoptotic protein, Bim, in tumor initiation and progression. We have previously shown that Bim is required for apoptosis in all three experimental models. The studies proposed in this application will investigate the consequences of loss or gain of Bim expression in several mouse tumor models and examine Bim expression in human breast tumors. These studies will provide valuable information on the cellular pathways that are required for maintenance of tumor cell survival, the role of apoptotic proteins in breast tumor progression, and potentially identify targets for therapeutic elimination of tumor cells. ? ? ?
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