Regulation Of Human Carcinoma Cell Adhesion

National Institute Of Environmental Health Sciences

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Abstract

We have demonstrated that cis-polyunsaturated fatty acids stimulate adhesion of human breast tumor cells through a pathway that involves MAP kinases, protein kinases C and D, and beta-1 integrins. We have identified a MAP kinase, p38, as a specific protein that is phosphorylated during the activation of adhesion in these cells by the fatty acids. The activation of p38 requires upstream kinases, starting with TAK1, which is regulated by a protein complex including TAB1 and 2 and various TRAF molecules. We have determined that ubiquitination is required for activation of this pathway, and are currently examining these protein complexes to determine which proteins are the critical targets for ubiquitination after fatty acid exposure.[unreadable] [unreadable] Furthermore, we have shown that two kinases, protein kinase C epsilon and protein kinase D, are translocated to membrane fractions in response to exposure of breast tumor cells to a dietary fatty acid. The phosphorylation and proteolytic processing of PKD by calpain 1 is critical for adhesion of breast tumor cells on type IV collagen. We have also shown that dietary fatty acids induce changes in the localization of key cytoskeletal proteins, such as filamentous actin, vinculin, and alpha-actinin, and increase the colocalization of beta-1 integrins with these proteins. We are currently working to identify other members of signal transduction pathways whose activity is regulated by these fatty acids and to determine how these signals lead to changes in the cytoskeleton and, ultimately, in cell behaviors such as migration and invasion[unreadable] [unreadable] We have now shown that specific fatty acids induce increase invasion of human breast tumor cells in an in vitro invasion model, and that the small G proteins, Rho and Rac, are modulated in their activity upon exposure to arachidonic acid. These proteins are known to regulate cell motility and invasion and may explain the invasive phenotype exhbited by these cells. The activation of Rho depends on its association with p115RhoGEF, which is regulated by p38 activation. This pathway activates the Rho associated kinase, ROCK, whose activation is required for the phosphorylation and translocation of nucleolin, a nuclear protein critical for cell adhesion in our cells.[unreadable] [unreadable] To make a connection between the ability of dietary fats to influence tumor cell behavior in vitro, and the increased metastasis seen in animal models, we have examined the ability of dietary fats to induce changes in the gene expression profiles in human tumor cells and in mouse lungs, which serve as a target for metastasis. Using a cell line with high propensity to metastasize to the lung, we have examined the frequency and character of metastasis in mice fed various diets. We are examining the gene profiles and are identifying the pathways that are critical for effective metastasis in this system.[unreadable] [unreadable] We have also developed a mouse model with human gastric carcinoma cells that shows an increase in tumor cell growth when the mice are fed diets high in linoleic acid. We have now shown that the metastatic variant of these human tumor cells have increased invasive potential that depends on PI-3-Kinase activity. We are examining these cells to determine whether inhibition of PI-3-K can block the increased tumor growth in mice on high fat diets. We are also examining the signal transduction pathways responsible for the increase in invasive behavior in response to linoleic acid and the decreased invasion in response to omega-3 fatty acids, such as EPA and DHA.