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. Author manuscript; available in PMC: 2017 Jul 19.
Published in final edited form as: Nat Mater. 2016 Mar 14;15(7):792–801. doi: 10.1038/nmat4586

Figure 4.

Figure 4

Functional PTEN in non-invasive SBcl2 modulates its topotactic properties accounting for the responses shown in Fig. 1. (a) Non-invasive SBcl2 melanoma but not invasive 1205Lu cells express PTEN, a PI3K antagonist. Non-invasive melanoma cell lines, SBcl2 and WM1552C, display lower PI3K activity and higher ROCK activity vs. invasive 1205Lu and WM983B cell lines on GPDA coated with 1 μg/ml of FN; (b) Dependence of the PI3K and ROCK activity in SBcl2 cells on the ECM density measured by assaying their substrates, Akt and MLC correspondingly. The data were quantified and normalized using the analysis in panel (a) to show the relative PI3K and ROCK activities in SBcl2 (solid) and 1205Lu (dashed) cell lines. All error bars are S.E.M; (c) PI3K and ROCK signaling pathway activities in SBcl2 (solid) and 1205Lu (dashed) cell lines, inferred from panel (b), showing the predicted change in the balance of the relative PI3K and ROCK signaling activities and the resulting switch in the topotaxis directionality. This change in the activity balance is also schematically depicted in the diagram on the right; (d) The distributions of topotactic displacements of SBcl2 cells on 10 μg/ml FN and 50 μg/ml FN-coated GPDA over the course of 5 hours; (e) Reversal of the SBcl2 cell stiffness dependence on the post density in cells cultured on 50 μg/ml vs. 10 μg/ml FN coating density on GPDA. * = Statistical significance between the values on dense vs. sparse region. *P<0.05 and all paired two-sample Student's t-test. All error bars are S.E.M. (f) The fractions of SBcl2 cells that have relatively deeper FAs penetration into the substratum in the sparser vs. denser post array zones on 10 μg/ml FN and 50 μg/ml FN coated GPDA. * = Statistical significance of the deviation from the zero mean. n.s = no significance and ***P<0.005. Wilcoxon signed rank test for zero mean. (g) Confocal images showing differential vertical protrusions of SBcl2 cells depending on post density and ECM density. The analysis is equivalent to that shown in Fig. 2c.