Deleted in liver cancer 1 (DLC1) negatively regulates Rho/ROCK/MLC pathway in hepatocellular carcinoma.
OBJECTIVE
Deleted in liver cancer 1 (DLC1), a member of RhoGTPase activating protein (GAP) family, is known to have suppressive activities in tumorigenicity and cancer metastasis. However, the underlying molecular mechanisms of how DLC1 suppresses cell motility have not been fully elucidated. Rho-kinase (ROCK) is an immediate down-stream effector of RhoA in mediating cellular cytoskeletal events and cell motility. In the present study, we aimed to investigate the effects of DLC1 on Rho/ROCK signaling pathway in hepatocellular carcinoma (HCC).


RESULTS
We demonstrated that DLC1 negatively regulated ROCK-dependent actomyosin contractility. From immunofluorescence study, we found that ectopic expression of DLC1 abrogated Rho/ROCK-mediated cytoskeletal reorganization including formation of stress fibers and focal adhesions. It also downregulated cortical phosphorylation of myosin light chain 2 (MLC2). These inhibitory events by DLC1 were RhoGAP-dependent, as RhoGAP-deficient mutant of DLC1 (DLC1 K714E) abolished these inhibitory events. In addition, from western study, DLC1 inhibited ROCK-related myosin light chain phosphatase targeting unit 1 (MYPT1) phosphorylation at Threonine 853. By examining cell morphology under microscope, we found that ectopic expression of dominant-active ROCK released cells from DLC1-induced cytoskeletal collapse and cell shrinkage.


CONCLUSIONS
Our data suggest that DLC1 negatively regulates Rho/ROCK/MLC2. This implicates a ROCK-mediated pathway of DLC1 in suppressing metastasis of HCC cells and enriches our understanding in the molecular mechanisms involved in the progression of hepatocellular carcinoma.
Wong
Carmen Chak-Lui
CC
Department of Pathology, SH Ho Foundation Research Laboratory and Jockey Club Clinical Research Centre, The University of Hong Kong, Hong Kong, China.
Wong
Chun-Ming
CM
Ko
Frankie Chi-Fat
FC
Chan
Lo-Kong
LK
Ching
Yick-Pang
YP
Yam
Judy Wai-Ping
JW
Ng
Irene Oi-lin
IO
D000205:Actomyosin / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D024722:Cardiac Myosins / Q000378:metabolism*; D002448:Cell Adhesion; D002465:Cell Movement; D002522:Chlorocebus aethiops; D020690:GTPase-Activating Proteins; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism*; D018994:Myosin Light Chains / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000502:physiology*; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000378:metabolism*
C112352:DLC1 protein, human; D020690:GTPase-Activating Proteins; D018994:Myosin Light Chains; D025521:Tumor Suppressor Proteins; C107184:myosin light chain 2; D000205:Actomyosin; C515851:ROCK1 protein, human; D054460:rho-Associated Kinases; D024722:Cardiac Myosins; D020741:rho GTP-Binding Proteins