Reduction of <i>fad104</i> facilitates migration and invasion of A375C6 cells.

<p>A, Knockdown efficiency of <i>fad104</i> in A375C6 cells. A375C6 cells were transfected with two different siRNAs targeting <i>fad104</i> (si<i>fad104</i>-A, si<i>fad104</i>-B). Luciferase siRNA was used as a control. FAD104 protein expression was examined by Western blotting. 7.5 μg of protein was loaded per lane. β-actin expression was used as a control. The ratio of protein level of FAD104/β-actin, as determined by NIH-Image software, is shown under each lane. B, Transwell migration assay of <i>fad104</i> knockdown A375C6 cells and control cells. A total of 2.5x10⁴ cells were plated in the upper chamber of the filters that had been coated with fibronectin. Cells that migrated to the underside of the transwell insert were measured after 24 hours. Representative images of migrated cells are shown (upper panel). The mean number of migrated cells in the field was calculated (lower panel). C, Transwell invasion assay of A375C6 cells transfected with <i>fad104</i>siRNAs. A total of 1x10⁵ cells were plated in the upper chamber of the filters that had been coated with Matrigel. The cells that invaded the underside of the transwell insert were measured after 24 hours. Representative images of invaded cells are shown (upper panel). The mean number of invaded cells in the field was calculated (lower panel). D, The expression levels of <i>mmp2</i> mRNA in A375C6 cells transfected with si<i>fad104</i>-A were determined by qRT-PCR and normalized with 18S rRNA expression. Each column represents the mean with standard deviation (error bars) (n = 3). **p<0.01. Similar results were obtained in at least two independent experiments. Western blot shows the representative results in at least two independent experiments. Original uncropped images of blots are shown in Figure B in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0117197#pone.0117197.s001" target="_blank">S1 File</a>.