3417

Time-lapse video microscopy of detached mammary epithelial cells detected the extension of long, dynamic protrusions of the plasma membrane, and we have characterized the cytoskeletal mechanism underlying this novel observation. Nearly 90% of human solid tumors arise as carcinomas from epithelial cells, so such an aggressive motility response to detachment could have broad implications for metastatic spread. Epithelial cells are tasked with maintaining barrier function in the body through the formation of continuous sheets, but are particularly prone to apoptotic cell death when detached from the extracellular matrix. The dynamic protrusions that we observe may provide a selective advantage by promoting survival and the maintenance of the epithelial barriers through reattachment. Nontumorigenic and tumorigenic mammary epithelial cell lines of both human and mouse origin produce such protrusions, indicating that it is an inherent characteristic of the untransformed cells. This response is relatively short-lived in nontumorigenic cell lines, since they die by apoptosis within 24 hours in suspension. However, cell lines that resist apoptosis through expression of either Bcl-2 or Bcl-xL will persistently form protrusions for at least ten days. Our previous studies indicate that such apoptotically-resistant cell lines will not form tumors in mice and likely lie dormant in distant tissues. Given the persistent motility response in these detached cells, they may not be as dormant as previously suggested. These protrusions can be inhibited with tubulin depolymerizing agents, but not those affecting actin polymerization. Immunofluorescence microscopy and Western blotting demonstrate that the protrusions are largely composed of detyrosinated Glu-tubulin, a post-translationally modified form of tubulin that is found in stabilized microtubules. These microtubule processes can extend more than 5 times the cell diameter, and undergo a rapid probing motion more than once per second. Formation of these microtubule protrusions is necessary for efficient cell-cell and cell-substratum attachment. It is possible that this inherent microtubule response of mammary epithelial cells provides some of the invasive characteristics of breast cancer, independent of any tumor-specific genetic alteration.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]