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. 2011 Nov 14;287(1):148–158. doi: 10.1074/jbc.M111.298117

FIGURE 8.

FIGURE 8.

PTK6 promotes cell migration through p130CAS and ERK5 signaling. A, PC3 cell migration is diminished upon PTK6 knockdown in Transwell chamber assays using 20% FBS as a chemoattractant. B, knockdown of PTK6 by siRNA in PC3 cells lasts 4 days. Cells were transfected with control siRNA (si-Cont) or PTK6 siRNA for 24 h and harvested at 2 (d2) or 4 (d4) days after transfection. Total cell lysates were analyzed by immunoblotting with anti-PTK6, p130CAS, phospho-p130CAS (PY-165), ERK5, P-ERK5, and β-actin antibodies. C, knockdown of PTK6 results in decreased ERK5 phosphorylation in PC3 cells. Relative band densities from B were quantified with NIH ImageJ software (44). Data were collected from two individual experiments, and S.D. is shown. D, PC3 cells stably expressing Palm-PTK6-YF show increased migration in wound healing assays. Representative images at 0 and 48 h are shown. Size bars denote 50 μm. E, membrane-targeted PTK6 promotes cell migration through phosphorylation of p130CAS and activation of ERK5. PC3 cells stably expressing Palm-PTK6-YF show greater cell migratory ability than control cells in Transwell chamber assays using 20% FBS as a chemoattractant. Knockdown of FAK causes a modest decrease, whereas knockdown of p130CAS or ERK5 (E5) results in a substantial reduction of cell migration induced by Palm-PTK6-YF. Error bars in A, C, and E represent standard deviation. F, a proposed model shows how PTK6 regulates peripheral adhesion complex formation and promotes migration. Integrin and growth factor receptor receptors are upstream of PTK6. PTK6 phosphorylates p130CAS near the plasma membrane (PM) and then activates ERK5 signaling, inducing formation of peripheral adhesion complexes and promoting migration. PTK6 has been shown to directly target AKT and promote AKT activation (9). Palm-PTK6-YF-expressing PC3 cells show increased AKT activation upon FBS stimulation that can be impaired by knockdown of p130CAS. AKT was reported to promote migratory and invasive ability of squamous carcinoma cells (62). Solid line arrows indicate direct regulation, whereas dotted line arrows represent proposed or indirect regulation. Vec, vector.