ROCK2 (Rho-associated, coiled-coil containing protein kinase 2)

2012-12-01   Carmen Chak-Lui Wong , Irene Oi-Lin Ng 

Department of Pathology, State Key Laboratory for Liver Research, The University of, Hong Kong, Hong Kong, China





ROCK2 was first identified as a target of active RhoA in an expression screening assay from a rat brain cDNA library (Leung et al. 1995).


The ROCK2 gene is 4164 bp long which encodes 1388 amino acids producing a protein of 160 kDa (UniGene: Hs.681743). The ROCK2 gene comprises 33 exons.



ROCK2 is a 160 kDa serine/threonine kinase which is composed of a kinase domain (44-415 aa) at the amino-terminus, a rho-binding domain (981-1046 aa), and a pleckstrin homology (PH) domain with a cysteine-rich region at the carboxyl-terminus (1152-1350 aa). A coiled-coil inhibitory structure was predicted to form between the kinase domain and the PH domain. ROCK2 exists in an auto-inhibitory form which could be released when active form of RhoA (RhoA-GTP) binds to the RBD domain of ROCK2. The functional domains of ROCK2 have unique roles such as protein-protein interaction, cellular localization, and regulation of ROCK2 activity. Kinase domain of ROCK2 is the key domain conferring the most important function of ROCK2 which is the phosphorylation and activation of its downstream substrates (Leung et al., 1995). PH domain of ROCK2 is responsible for lipid binding and membrane localization of ROCK2 (Miyazaki et al., 2006). PH domain of a ROCK2 homolog also facilitates the interaction with other proteins such as filamin A, a protein critical to actin cytoskeleton remodeling (Ueda et al., 2003).
Atlas Image
Schematic diagram of ROCK2. RBD: rho-binding domain; PH: pleckstrin homology; C1: cysteine-rich region.


Very few studies have demonstrated the distinctive roles of ROCK1 and ROCK2; however, their differential tissue distribution indicates that they participate in different physiological functions. ROCK1 and ROCK2 are ubiquitously expressed in all tissues. Earlier studies using Northern blot analysis demonstrated that ROCK1 mRNA expression was especially abundant in testis, liver, and lung while ROCK2 mRNA expression was especially abundant in brain and muscle (Leung et al., 1996).
Regulation: The most common regulatory mechanism of ROCK2 is mediated by RhoGTPases whose activity exchanges between a GTP-bound active form and a GDP-bound inactive form. RhoA-GTP binds to the RBD domain of ROCK and releases the kinase domain of ROCK leading to the activation of ROCK (Leung et al., 1995). In addition to this classical model of activity regulation, ROCK2 can also be activated by lipids such as arachidonic acid (Feng et al., 1999). The ROCK2 kinase domain can also be released when the C-terminus of ROCK2 is cleaved by granzyme B (Sebbagh et al., 2005). Furthermore, Polo-like kinase-1 (Plk1) interacts with and phosphorylates ROCK2 at Threonine 967, Serine 1099, Serine 1133, and S1374 and activates ROCK2 (Lowery et al., 2007).
Recent studies have demonstrated that ROCK2 is also regulated at the expression level. Two microRNAs (miRNAs), miR-139 and miR-124, have independently been shown to interact with the 3untranslated region of ROCK2 and subsequently suppress ROCK2 expression in hepatocellular carcinoma (HCC) cell lines (Wong et al., 2011; Zheng et al., 2012). MiR-139 and miR-124 have been shown to be downregulated in human HCCs and their expression levels inversely correlated with ROCK2 protein expression in human HCC samples (Wong et al., 2011; Zheng et al., 2012).


ROCK2 is mainly found in the cytoplasm in most cell types. It was reported that overexpression of a dominant-active form of RhoA re-localized ROCK2 to the membranous actin filaments in a cervical cancer cell line, HeLa (Leung et al., 1995). A study has also shown that ROCK2 could be found in the nuclei in multiple cell types such as human keratinocytes, mouse mammary epithelial cells, osteoblasts, and mouse fibroblasts (Tanaka et al., 2006). ROCK2 was also found to be located in the centrosome in fibroblasts (Ma et al., 2006; Wang et al., 2011).


ROCK2 are responsible for many key cellular processes including cell migration and invasion (Croft et al., 2004), apoptosis (Sebbagh et al., 2005), centrosome duplication (Ma et al., 2006), and cytokinesis (Lowery et al., 2007).
The best well-characterized function of ROCK is contributed by the kinase domain for its ability to phosphorylate the serine/threonine residues of downstream substrates that are important for actin cytoskeleton organization. The most well-known substrates of ROCK include myosin light chain 2 (MLC2) and myosin phosphatase 1 (MYPT1) which regulate actomyosin contractility in cells (Amano et al., 1996). Other ROCK substrates include cofilin and LIM kinases, responsible for actin polymerization and depolymerization (Yang et al., 1998; Sumi et al., 1999; Ohashi et al., 2000; Sumi et al., 2001); vimentin (Goto et al., 1998), responsible for actin filament formation; adducin, responsible for membrane ruffles formation (Fukata et al., 1999); calponin, responsible for actin filament binding (Kaneko et al., 2000); ezrin, responsible for anchoring the cytoskeleton to the cell membrane and the formation of focal adhesion molecules (Matsui et al., 1998; Tran Quang et al., 2000).
In addition to cell movement, ROCK2 was shown to be important in keratinocyte differentiation (McMullan et al., 2003). Also, Plk1-mediated phosphorylation and activation of ROCK2 induced cytokinesis (Lowery et al., 2007). Nuclear function of ROCK2 was first reported when ROCK2 was found to be a binding partner of p300 acetyltransferase (Tanaka et al., 2006). Nuclear ROCK2 phosphorylated p300 and activated p300-mediated transcription (Tanaka et al., 2006). Centrosomal ROCK2 interacted with nucleophosmin/B23 and BRCA2 proteins for centrosome duplication (Ma et al., 2006; Wang et al., 2011).
ROCK2 also regulates the extracellular matrix. Expression of conditional active ROCK2 construct has been shown to activate MYPT and MLC and drive actomyosin contraction in mouse skin, thereby modifying the ECM through increased collagen deposition and tissue stiffness (Samuel et al., 2011). Along with the ECM modification, activation of ROCK2 may also increase nuclear accumulation and activity of β-catenin. Activation of β-catenin can elevate epidermal cell proliferation rate, subsequently contributing to epidermal hyperplasia and tumor growth (Samuel et al., 2011).


ROCK2 shares close homology with another protein called ROCK1. ROCK1 and ROCK2 are 65% homologous in human; particularly, their kinase domains are 87% homologous (Leung et al., 1996). Due to their high degree of homology, ROCK1 and ROCK2 are regulated by common mechanisms and share many common substrates.

Implicated in

Entity name
Solid cancers
ROCK, in general, has been shown to be implicated in various cancer cell line models; however, only a few studies focus on the expression and roles of ROCK2 in cancers.
Entity name
Colorectal cancer
In a subcutaneous tumor model in nude mice, conditional expression of active ROCK2 construct in colorectal cancer cell lines has been demonstrated to enhance angiogenesis and cancer cell invasion into the surrounding stromal tissues (Croft et al., 2004).
Entity name
Hepatocellular carcinoma
ROCK2 protein was shown to be over-expressed in 54% (22/41 cases) of human hepatocellular carcinoma (HCC) as compared with their corresponding non-tumorous liver tissues by Western blot analysis (Wong et al., 2009). Over-expression of ROCK2 in HCC was associated with the presence of tumor microsatellite formation, an important clinicopathological feature of aggressive HCC and an indicator of intrahepatic metastasis in human HCC (Wong et al., 2009). Knockdown of ROCK2 in human HCC cell lines suppressed stress fiber and focal adhesion formation, as well as actomyosin contractility in HCC cells, thereby retarding HCC cell invasion in vitro and in vivo (Wong et al., 2009).
Entity name
Squamous skin carcinomas
Immunohistochemical study using an antibody with reactivity towards both ROCK1 and ROCK2 indicates that high ROCK expression was detected in 40 cases of human squamous skin carcinomas (Samuel et al., 2011). Along with this finding, phosphorylation of MYPT, as a reflection of ROCK activity, was also detected in these cases of carcinoma (Samuel et al., 2011).
Entity name
Testicular cancer
ROCK2 protein was shown in Western blot analysis to be overexpressed in testicular cancers as compared to non-tumorous tissues in a cohort of 57 patients (Kamai et al., 2004). ROCK2 protein expression was significantly higher in patients with recurrent testicular cancers than those without sign of recurrence after treatment (Kamai et al., 2004).
Entity name
Breast cancer
It has long been challenging to detect the ROCK activity in human clinical specimens owing to the fact that ROCK activity declines rapidly in clinical specimens after resection. Also, the phosphorylation sites in ROCK, which truly reflect its activity status, have not been identified or extensively studied. Recently, a study has demonstrated that the activity status of ROCK2 can be reflected by the phosphorylation level of ROCK2 at Ser1366. Immunohistochemical staining shows that phosphorylation of ROCK2 at Ser1366 was increased in 2 cases of breast cancer tissues as compared to their normal counterparts (Chuang et al., 2012).


Pubmed IDLast YearTitleAuthors
87027561996Phosphorylation and activation of myosin by Rho-associated kinase (Rho-kinase).Amano M et al
222731452012ROCKII Ser1366 phosphorylation reflects the activation status.Chuang HH et al
156042642004Conditional ROCK activation in vivo induces tumor cell dissemination and angiogenesis.Croft DR et al
109983512000Specificity and mechanism of action of some commonly used protein kinase inhibitors.Davies SP et al
99209271999Rho-associated kinase of chicken gizzard smooth muscle.Feng J et al
102090291999Phosphorylation of adducin by Rho-kinase plays a crucial role in cell motility.Fukata Y et al
95655951998Phosphorylation of vimentin by Rho-associated kinase at a unique amino-terminal site that is specifically phosphorylated during cytokinesis.Goto H et al
107793822000Pharmacological properties of Y-27632, a specific inhibitor of rho-associated kinases.Ishizaki T et al
99308721999An essential part for Rho-associated kinase in the transcellular invasion of tumor cells.Itoh K et al
152691552004Overexpression of RhoA, Rac1, and Cdc42 GTPases is associated with progression in testicular cancer.Kamai T et al
108735722000Identification of calponin as a novel substrate of Rho-kinase.Kaneko T et al
88164431996The p160 RhoA-binding kinase ROK alpha is a member of a kinase family and is involved in the reorganization of the cytoskeleton.Leung T et al
74939231995A novel serine/threonine kinase binding the Ras-related RhoA GTPase which translocates the kinase to peripheral membranes.Leung T et al
174468642007Proteomic screen defines the Polo-box domain interactome and identifies Rock2 as a Plk1 substrate.Lowery DM et al
170154632006Interaction between ROCK II and nucleophosmin/B23 in the regulation of centrosome duplication.Ma Z et al
94563241998Rho-kinase phosphorylates COOH-terminal threonines of ezrin/radixin/moesin (ERM) proteins and regulates their head-to-tail association.Matsui T et al
146806352003Keratinocyte differentiation is regulated by the Rho and ROCK signaling pathway.McMullan R et al
168451712006Dynamics of RhoA and ROKalpha translocation in single living cells.Miyazaki K et al
106523532000Rho-associated kinase ROCK activates LIM-kinase 1 by phosphorylation at threonine 508 within the activation loop.Ohashi K et al
216651512011Actomyosin-mediated cellular tension drives increased tissue stiffness and β-catenin activation to induce epidermal hyperplasia and tumor growth.Samuel MS et al
156990752005Direct cleavage of ROCK II by granzyme B induces target cell membrane blebbing in a caspase-independent manner.Sebbagh M et al
82174081993[Treatment of cerebral vasospasm by a protein kinase inhibitor AT 877].Shibuya M et al
110180422001Specific activation of LIM kinase 2 via phosphorylation of threonine 505 by ROCK, a Rho-dependent protein kinase.Sumi T et al
112307372001Inhibition of intrahepatic metastasis of human hepatocellular carcinoma by Rho-associated protein kinase inhibitor Y-27632.Takamura M et al
165746622006Nuclear Rho kinase, ROCK2, targets p300 acetyltransferase.Tanaka T et al
109708502000Ezrin function is required for ROCK-mediated fibroblast transformation by the Net and Dbl oncogenes.Tran Quang C et al
125897952003The carboxy-terminal pleckstrin homology domain of ROCK interacts with filamin-A.Ueda K et al
210842792011BRCA2 and nucleophosmin coregulate centrosome amplification and form a complex with the Rho effector kinase ROCK2.Wang HF et al
209516992011The microRNA miR-139 suppresses metastasis and progression of hepatocellular carcinoma by down-regulating Rho-kinase 2.Wong CC et al
96553981998Cofilin phosphorylation by LIM-kinase 1 and its role in Rac-mediated actin reorganization.Yang N et al
216729402012The putative tumour suppressor microRNA-124 modulates hepatocellular carcinoma cell aggressiveness by repressing ROCK2 and EZH2.Zheng F et al

Other Information

Locus ID:

NCBI: 9475
MIM: 604002
HGNC: 10252
Ensembl: ENSG00000134318


dbSNP: 9475
ClinVar: 9475
TCGA: ENSG00000134318


Gene IDTranscript IDUniprot

Expression (GTEx)



PathwaySourceExternal ID
Wnt signaling pathwayKEGGko04310
Axon guidanceKEGGko04360
Focal adhesionKEGGko04510
Tight junctionKEGGko04530
Leukocyte transendothelial migrationKEGGko04670
Regulation of actin cytoskeletonKEGGko04810
Pathogenic Escherichia coli infectionKEGGko05130
Wnt signaling pathwayKEGGhsa04310
Axon guidanceKEGGhsa04360
Focal adhesionKEGGhsa04510
Tight junctionKEGGhsa04530
Leukocyte transendothelial migrationKEGGhsa04670
Regulation of actin cytoskeletonKEGGhsa04810
Pathogenic Escherichia coli infectionKEGGhsa05130
Pathways in cancerKEGGhsa05200
Vascular smooth muscle contractionKEGGhsa04270
Chemokine signaling pathwayKEGGko04062
Vascular smooth muscle contractionKEGGko04270
Chemokine signaling pathwayKEGGhsa04062
Salmonella infectionKEGGko05132
Salmonella infectionKEGGhsa05132
Proteoglycans in cancerKEGGhsa05205
Proteoglycans in cancerKEGGko05205
Platelet activationKEGGhsa04611
Oxytocin signaling pathwayKEGGhsa04921
Oxytocin signaling pathwayKEGGko04921
cGMP-PKG signaling pathwayKEGGhsa04022
cGMP-PKG signaling pathwayKEGGko04022
cAMP signaling pathwayKEGGhsa04024
cAMP signaling pathwayKEGGko04024
Sphingolipid signaling pathwayKEGGhsa04071
Sphingolipid signaling pathwayKEGGko04071
Signal TransductionREACTOMER-HSA-162582
Signaling by VEGFREACTOMER-HSA-194138
Signaling by Rho GTPasesREACTOMER-HSA-194315
RHO GTPase EffectorsREACTOMER-HSA-195258
RHO GTPases Activate ROCKsREACTOMER-HSA-5627117
Signaling by GPCRREACTOMER-HSA-372790
GPCR downstream signalingREACTOMER-HSA-388396
G alpha (12/13) signalling eventsREACTOMER-HSA-416482
Developmental BiologyREACTOMER-HSA-1266738
Axon guidanceREACTOMER-HSA-422475
Semaphorin interactionsREACTOMER-HSA-373755
Sema4D in semaphorin signalingREACTOMER-HSA-400685
Sema4D induced cell migration and growth-cone collapseREACTOMER-HSA-416572
EPH-Ephrin signalingREACTOMER-HSA-2682334
EPHA-mediated growth cone collapseREACTOMER-HSA-3928663
EPHB-mediated forward signalingREACTOMER-HSA-3928662

Protein levels (Protein atlas)

Not detected


Pubmed IDYearTitleCitations
173444172007Notch1 is a p53 target gene involved in human keratinocyte tumor suppression through negative regulation of ROCK1/2 and MRCKalpha kinases.125
209516992011The microRNA miR-139 suppresses metastasis and progression of hepatocellular carcinoma by down-regulating Rho-kinase 2.115
174468642007Proteomic screen defines the Polo-box domain interactome and identifies Rock2 as a Plk1 substrate.103
189228922008Phosphorylated caveolin-1 regulates Rho/ROCK-dependent focal adhesion dynamics and tumor cell migration and invasion.94
191316462009ROCK isoform regulation of myosin phosphatase and contractility in vascular smooth muscle cells.86
199131212009Gene-centric association signals for lipids and apolipoproteins identified via the HumanCVD BeadChip.85
202323932010Downregulation of the Rho GTPase signaling pathway is involved in the microRNA-138-mediated inhibition of cell migration and invasion in tongue squamous cell carcinoma.70
156990752005Direct cleavage of ROCK II by granzyme B induces target cell membrane blebbing in a caspase-independent manner.64
175965092007Rho/Rho-associated kinase-II signaling mediates disassembly of epithelial apical junctions.60
186958902008The expression and prognostic value of ROCK I and ROCK II and their role in human breast cancer.51


Carmen Chak-Lui Wong ; Irene Oi-Lin Ng

ROCK2 (Rho-associated, coiled-coil containing protein kinase 2)

Atlas Genet Cytogenet Oncol Haematol. 2012-12-01

Online version: