A Novel Role of Fascin in Carcinoma Cells

Colorectal cancer (CRC) is the second deadliest cancer in the US according to the 2014 National Cancer Institute data. Despite recent academic and medical advances, our understanding of the progression of CRC and, therefore, new therapeutic approaches to improve patient outcomes remains poor. Fascin, an actin-bundling protein, although absent from the adult epithelial cells of the colon, is often overexpressed in CRC. In this study we investigated the role of fascin in CRC. We also examined the cancer-related functions of fascin in non-transformed Madin-Darby Canine Kidney cells (MDCK) to validate fascin specific functions in CRC cells. CRC cells (HT-29/19A) and MDCK cells with or without fascin were analyzed for the formation of multilayered epithelium, a phenotype in which cells grow on top of other cells forming several layers of cells. We investigated cell proliferation and contact inhibition in low and high density cultures. Tumors were grown using CRC cells with or without fascin to evaluate the role of fascin in proliferation of cells in vivo. We measured the ability of cells to attach to a substrate. Cells were transduced with mCherry-tagged LifeAct lentivirus to observe the dynamics of stress fibers. Mechanistically, we investigated the role of actin-binding by fascin in stress fibers formation. The results show that fascin promotes multilayering of cells by evading contact inhibition in CRC and in non-transformed MDCK cells. Decreased integrin 1 protein and reduction in cell-matrix attachment causes this multilayering. Fascin disrupts stress fibers which in turn prevents the localization of vinculin and -actinin to focal adhesion sites. Mechanistically, by binding to LIMK1 (p-Lin-11/Isl-1/Mec-3 kinase 1), fascin prevents the phosphorylation of cofilin, which reduces polymerization of filamentous actin in stress fibers. We conclude that fascin promotes evasion of contact inhibition by dysregulation of stress fibers to promote multilayering of carcinoma cells.Biology and Biochemistry, Department o