Non-metastatic Lewis lung carcinoma cells (LLC-C8) become more motile when protein phosphatases (PP-1 and -2A) are inhibited by okadaic acid, attaining the same level of motility as metastatic LLC (LLC-LN7) variants. This stimulation of LLC-C8 motility was tempered when protein kinase A activity was inhibited. We examined whether the okadaic acid-stimulated LLC-C8 motility was associated with alterations in the cytoskeletal organization so that these non-metastatic cells acquire the rounded morphology and diffuse cytoskeletal organization previously described for metastatic LLC-LN7 cells. Non-metastatic LLC-C8 are typically adherent during culture, achieving a spread morphology. Treatment of non-metastatic LLC-C8 cells with okadaic acid resulted in a contraction of most of their extended processes, formation of spikes and membrane blebs within 10 min, and complete cell rounding within 20 min for most of the cells. While the overall level of F-actin was minimally affected by the okadaic acid, its uniform distribution shifted to localization toward the periphery of the rounded cells, often concentrating at a single focus. Immunofluorescent staining for vimentin showed a similar shift to the cell periphery and similar capping. After okadaic acid treatment, the filamentous network of microtubules in non-metastatic LLC-C8 cells disappeared and was replaced with a diffusely staining distribution of beta-tubulin. These results show that PP-1 and -2A maintain cytoskeletal organization and that inhibition of this control reduces cytoskeletal organization and increases tumor cell motility.