JCS008037 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Relevance of Src family, ERK and Abl/Arg activities for ECM degradation. Lysates of A375MM cells pre-incubated with 5 µM SU6656, 10 µM U0126 or vehicle in the presence of BB94, and later incubated for further 3 hours with 5 µM SU6656, 10 µM U0126 or vehicle after BB94 washout, were subjected to SDS-PAGE, transferred to nitrocellulose and probed with anti-c-Src and anti-phospho-Src (A), or anti-phospho-ERK1/2 and anti-ERK1/2 (D). Representative experiments show that SU6656 and U0126 treatment inhibits phosphorylation of c-Src and both ERK1 (p42) and 2 (p44), respectively. (B) Inhibition of Src phosphorylation by SU6656 blocks invadopodia formation and therefore ECM degradation. (C) Inhibition of Abl/Arg family kinases do not affect invadopodia formation nor modify matrix degradation. (E) By contrast, U0126 reduces matrix degradation by 50%. Data represent the mean ± s.d. of three independent experiments. Statistical significances were evaluated by Student’s t-test: control vs SU6656- or UO126-treated P<0.0001.

• Supplemental Figure S2 -

  Fig. S2. ECM degradation after cotransfection of cortactin mutants. Non-phosphorylatable cortactin^{Y421,466,482F}, cortactin^S113A and cortactin^S405,^418A were cotransfected with pseudo-phosphorylated cortactin^S113D, cortactin^S405,418D and cortactin^S113D respectively. In all cases, the phosphomimicking mutants bypassed the effect of the non-phosphorylatable ones, suggesting that independent but convergent signaling pathways might regulate cortactin function at invadopodia. Data represent the mean ± s.d. of two independent experiments.