Roles and regulations of Rho-kinases in hepatocellular carcinoma

Abstract

Hepatocellular carcinoma (HCC) is the most common liver cancer and is one of the most prevalent cancers in the human population which causes 500, 000 deaths per annum. However, a complete understanding of the underlying molecular mechanisms involved in HCC is still missing. Since intrahepatic metastasis is the main cause of lethality in HCC patients, study of the molecular mechanisms involved in intrahepatic extension engages a high significance. Rho GTPases (Rho) are responsible for reorganizing actin cytoskeleton in cells and hyperactivation of Rho is implicated in cancer cell metastasis and invasion. Rho-associated serine-threonine protein kinase (ROCK), a downstream effector of Rho, can be activated by the active GTP bound form of Rho, and ROCK is found to be related to the regulations of cell migration. However, the functional roles and regulations of ROCK in HCC still remain to be elusive and hence were investigated herein.

In this study, we demonstrated that ROCK was overexpressed in 59% of primary HCC by at least 2 fold comparing with their corresponding non-tumorous livers in protein level. Besides, ROCK was highly expressed in HCC cell lines especially those with high metastatic potential. Furthermore, ROCK specific inhibitor, Y27632, exhibited substantial inhibitory effect in HCC cell migration via suppressing the formation of stress fibers and focal adhesions in HCC cells.

Rho/ROCK upstream regulators are important in determining the fate of the cell by controlling Rho/ROCK pathway. Members of GTPase Activating Proteins (GAPs) family, well-known negative regulators of Rho, inactivate Rho by promoting Rho to stay in its inactive GDP bound form. Deleted in Liver Cancer 1 (DLC1), encodes a RhoGAP protein, is a potential upstream negative regulator of Rho/ROCK pathway. In this study, we verified the regulatory role of DLC1 in ROCK activity. Ectopic expression of DLC1 inhibited ROCK activity and this inhibitory effect was completely lost in DLC1 RhoGAP deficient mutant, demonstrating that DLC1 negatively regulated ROCK activity via RhoGAP. In conclusion, we showed that ROCK is implicated in HCC cell migration and ROCK activity is negatively regulated by DLC1 in a RhoGAP dependent manner.