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. 2016 Jun 20;7(32):51908–51921. doi: 10.18632/oncotarget.10188

Figure 4. Roc-A alters the morphology of F-actin-based protrusions by an indirect effect on actin polymerization.

Figure 4

A. Roc-A alters the morphology of F-actin-based protrusions. PC-3, MDA-MB-231 and 293T cells were treated with Roc-A (15 nM for PC3, 60 nM for MDA-MB-231 and 30 nM for 293T) or solvent (DMSO) for 24h followed by staining of F-actin (green) and nuclei (blue). Three independent experiments were carried out per cell line. Representative images (Z-stacks) per cell line are shown. Scale bar = 20 μm (PC-3) or 10 μm (293T and MDA-MB-231). B. Roc-A does not directly affect actin polymerization. The influence of Roc-A on polymerization of pyrene-conjugated actin monomers was monitored by measuring the increase in fluorescence that occurs upon polymerization of pyrene-conjugated actin. Roc-A, solvent control or the known actin polymerization promoting agent Jasplakinolide were added to pyrene-conjugated actin monomers and 20 min later actin polymerization was initiated. Results are representative of three independent experiments. C. Roc-A does not directly affect actin depolymerization. The influence of Roc-A on depolymerization of pyrene-conjugated actin monomers was monitored by measuring the decrease in fluorescence that occurs upon depolymeization of pyrene-conjugated actin. Roc-A, solvent control or the known F-actin stabilizer phalloidin were added to polymerized pyrene-conjugated actin for 20 min, after which depolymerization of actin was initiated. Results are representative of three independent experiments.