SPDEF expression reduces in vitro tumor cell migration, invasion, and aggressive phenotype.
A, representative bioluminescent image quantitation of VC, SPDEF-expressing (SPDEF OE-Luc) PC3 cells, Scr shRNA, and SPDEF KD-Luc LNCaP cells, confirming luciferase expression. B, Western blot analysis indicating the expression levels of SPDEF protein in the SPDEF OE-Luc cells (both anti-SPDEF and anti-FLAG antibodies) and knockdown levels in two different shRNA LNCaP-Luc clones. Tubulin was used as a protein loading control. C, SPDEF overexpression reduces whereas knockdown increases cell migration. Shown are images and quantitation of wound healing assays performed between the four cell lines. D, SPDEF overexpression reduces and knockdown increases tumor cell invasion. SPDEF OE-Luc PC3 cells are less invasive, whereas SPDEF KD-Luc LNCaP cells are more invasive as determined using Matrigel-coated Boyden chambers as described under “Experimental Procedures.” E, representative images of three-dimensional Matrigel culture of PC3-Luc and LNCaP-Luc cells with and without SPDEF overexpression or knockdown. VC-Luc, VC-PC3-Luc; SPDEF OE-Luc, SPDEF-PC3-Luc; Scr-Luc, Scr-LNCaP-Luc; SPDEF KD-Luc, SPDEF-KD-LNCaP-Luc). *, statistical significance (p < 0.05) compared with VC-Luc or Scr-Luc cells.