Suppression of ATX expression in human TM cells induces changes in actin cytoskeletal organization and myosin II phosphorylation.

<p>A. Primary cultures of human TM cells treated with ATX-specific siRNA revealed significant suppression of ATX expression (different isoforms) relative to cells treated with FITC-conjugated scrambled siRNA control. β-tubulin was used as a loading control. B. Protein levels of one of the variants (∼125 kDa) of ATX were significantly reduced (by >75%) in ATX siRNA treated HTM cells based on densitometric semi-quantification. C. The ATX siRNA treated human TM cells (after 72 hrs) maintained under serum starved conditions for 24 hrs revealed a marked decrease in actin stress fibers (labelled using rhodamine-phalloidin) relative to cells treated with scrambled siRNA (FITC-siRNA). D. ATX siRNA treated human TM cells (for 72 hrs) maintained under serum starved conditions for 24 hrs showed a significant decrease in the levels of phosphorylated myosin light chain (pMLC), a regulatory subunit of myosin II, as compared to cells treated with scrambled siRNA (FITC-siRNA). β-tubulin was blotted as a loading control. E. Changes in the levels of phospho-MLC in the ATX siRNA treated cells compared to the scrambled siRNA treated control, based on densitometric semi-quantitative analysis.