Fig 1.

Syx is necessary to maintain U251 static cell polarity. (A) Western blot analysis of Syx expression in a panel of conventional glioblastoma multiforme (GBM) cancer cell lines. HUVECs are included as a positive control. (B) Western blot analysis of Syx expression in human GBM tumors serially propagated in the flanks of mice. (C) For the left panel, a quantitative real-time PCR analysis of Syx mRNA levels was performed in cells expressing control shRNA versus either of two nonoverlapping syx shRNAs, normalized to GAPDH (glyceraldehyde-3-phosphate dehydrogenase) mRNA levels. The right panels presents the results of a Western blot analysis showing the efficiency of Syx protein depletion in U251 cells expressing the same shRNAs. (D) Immunofluorescence staining of vinculin (green) and f-actin (phalloidin; red) in U251 cells expressing control shRNA versus either of two nonoverlapping syx shRNAs. Note the progressive cell rounding and loss of front-rear polarity from control shRNA expressing cells to Syx shRNA2-expressing cells. (E) As in panel D, cells expressing indicated shRNAs were stained for α-tubulin. Arrows point to examples of unbundled microtubules.