Positive Quantitative Relationship between EMT and Contact-Initiated Sliding on Fiber-like Tracks

Abstract

Epithelial-mesenchymal transition (EMT) is a complex process by which cells acquire invasive properties that enable escape from the primary tumor. Complete EMT, however, is not required for metastasis: circulating tumor cells exhibit hybrid epithelial-mesenchymal states, and genetic perturbations promoting partial EMT induce metastasis in vivo. An open question is whether and to what extent intermediate stages of EMT promote invasiveness. Here, we investigate this question, building on recent observation of a new invasive property. Migrating cancer cell lines and cells transduced with prometastatic genes slide around other cells on spatially confined, fiberlike micropatterns. We show here that low-dosage/short-duration exposure to transforming growth factor beta (TGFb) induces partial EMT and enables sliding on narrower (26 mm) micropatterns than untreated counterparts (41 mm). High-dosage/long-duration exposure induces more complete EMT, including disrupted cell-cell contacts and reduced E-cadherin expression, and promotes sliding on the narrowest (15 mm) micropatterns. These results identify a direct and quantitative relationship between EMT and cell sliding and show that EMT-associated invasive sliding is progressive, with cells that undergo partial EMT exhibiting intermediate sliding behavior and cells that transition more completely through EMT displaying maximal sliding. Our findings suggest a model in which fiber maturation and EMT work synergistically to promote invasiveness during cancer progression.

Golgi stabilization, not its front-rear bias, is associated with EMT-enhanced fibrillar migration

“…During the 12 days, cells were passaged at the regular frequency using the aforementioned standard methods, with the exception that cells were maintained in 20 ng/ml TGFβ-containing growth medium. This protocol matches exactly that used in our earlier work to quantify morphological and protein-expression changes induced by TGFβ treatment (11).…”

“…Third, the combined effect of EMT and fibrillar maturation on motility is seen not only at the level of single-cell motility, but also at the level of migration response to pairwise cell-cell interactions. We recently showed that when non-transformed epithelial cells encounter another cell along a fiber-like track, they reverse direction (11,15). Progression through TGFβ-mediated EMT shifts this behavior, enabling cells to slide past each other and continue to move in their original direction.…”

“…We recently used fiber-like micropatterns to show that EMT and fibrillar environment cooperate to regulate migration response to cell-cell interactions (11,15). When migrating nontransformed mammary epithelial cells encounter each other on fiber-like tracks, they reverse direction and move apart.…”

Multivariate relationships among nucleus and Golgi properties during fibrillar migration are robust to and unchanged by epithelial-to-mesenchymal transition

“…Migration of non-transformed human mammary epithelial cells (MCF10A) expressing histone 2B-GFP (nuclear marker) and GM130-RFP (Golgi marker) was imaged on micropatterned lines of collagen as detailed in [ 15 ]. MCF10A cells were maintained and passaged using standard growth medium and tissue culture protocols [ 16 , 17 ]. Additionally, to analyze how EMT affects relationships among nucleus and Golgi properties, MCF10A cells were treated with 20 ng/mL TGF β in growth medium for 12 days.…”

“…Most unexpectedly, our analysis shows that a PCA model built with data from untreated cells is remarkably similar to that built from data collected with TGF β -treated cells. Despite significant changes in gene expression, morphology and motile phenotype induced by TGF β -mediated EMT in these cells [ 15 , 16 ], treatment with TGF β does not significantly affect the relationships among nucleus size, aspect ratio and orientation with migration direction or among Golgi size and nucleus-Golgi separation distance. These results suggest that migration along spatially-confined fiber-like tracks employs a conserved nucleus-Golgi arrangement that is independent of EMT state.…”