Requirement of PEA3 for Transcriptional Activation of FAK Gene in Tumor Metastasis
Figure 1
Increased FAK expression in highly metastatic B16F10 melanoma cells.
(A–B) Comparison of the in vivo and in vitro metastatic potential of B16F10 and B16F1 melanoma cells. B16F10 and B16F1 cells (1×106) were injected intravenously into C57BL/6J mice via tail vein. After 20 days, lungs from the mice were resected and analyzed for metastasis. The representative lung in mice injected with F1 or F10 cells are shown (A-1). After fixation in Bouin’s solution, metastatic nodules found on lung surface are counted and averaged (A-2). Data are presented as mean ± SEM of five mice. ***P<0.01. (B-1) Migration of F10 and F1 in vitro. F1 and F10 cells were incubated in the transwell insert with 10% FBS in the lower chamber for 12 h. Photo show representative membrane attached migrated cells. Scale bar = 100 µm. Quantitative analysis of the number of the cells migrated to the down side of the membrane (B-2). Data are mean ±SEM of three independent experiments. ***P<0.01. (C–D) Northern blot and RT-PCR analyses of FAK mRNA in B16F10 and B16F1 cells. β-actin was amplified in RT-PCR and used as an internal control. (E-1) Western blot analysis of FAK expression level and p-FAK level in B16F10 and B16F1 cells. Whole cell lysates from B16F10 and B16F1 cells were blotted with antibodies to FAK, phosphorylated FAK (Y397 p-FAK) or tubulin as a control. (E-2) Densitometric quantification revealed a significant increase in expression of FAK protein in F10 cell line. Densitometric units were normalized to Tubulin and then divided by F1 group results. (n = 3; ***P<0.01, **p<0.05 ).