Signaling and Regulation The Activation of MEK/ERK Signaling Pathway by Bone Morphogenetic Protein 4 to Increase Hepatocellular Carcinoma Cell Proliferation and Migration

Hepatocellular carcinoma (HCC) is one of the most common visceral malignancies worldwide, with a very high incidence and poor prognosis. Bonemorphogenesis protein 4 (BMP4), which belongs to the TGF-b superfamily of proteins, is a multifunctional cytokine, which exerts its biologic effects through SMADand non-SMADdependent pathways, and is also known to be involved in human carcinogenesis. However, the effects of the BMP4 signaling in liver carcinogenesis are not yet clearly defined. Here, we first show that BMP4 and its receptor, BMPR1A, are overexpressed in amajority of primaryHCCs and that it promotes the growth andmigration ofHCC cell lines in vitro. We also establish that BMP4 can induce HCC cyclin-dependent kinase (CDK)1 and cyclin B1 upregulation to accelerate cell-cycle progression. Our study indicates that the induction ofHCC cell proliferation is independent of the SMADsignaling pathway, as Smad4 knockdown ofHCC cell lines still leads to the upregulation of CDK1 and cyclin B1 expression after BMP4 treatment. Using mitogen-activated protein/extracellular signalregulated kinase (MEK) selective inhibitors, the induction of CDK1, cyclin B1 mRNA and protein were shown to be dependent on the activation of MEK/extracellular signal-regulated kinase (ERK) signaling. In vivo xenograft studies confirmed that the BMPR1A-knockdown cells were significantly less tumorigenic than the control groups. Our findings show that the upregulation of BMP4 andBMPR1A inHCCpromotes the proliferation andmetastasis of HCC cells and that CDK1 and cyclin B1 are important SMAD-independent molecular targets in BMP4 signaling pathways, during the HCC tumorigenesis. It is proposed that BMP4 signaling pathways may have potential as new therapeutic targets in HCC treatment. Mol Cancer Res; 10(3); 415–27. 2012 AACR.