Epigenetically Silenced miR-1224 Suppresses the Proliferation of Human Hepatocellular Carcinoma via the CREB-Mediated YAP Signaling Pathway
41 Pages Posted: 18 Dec 2018More...
Background: MicroRNAs (miRNAs) are involved in a range of biological processes and their dysregulation have emerged as a critical regulatory mechanism in the progression of multiple human cancers, including hepatocellular carcinoma (HCC). Mounting evidence indicates that miR-1224 is commonly down-regulated in various malignant tumors including glioma and metastatic colorectal cancer, and acts as a tumor suppressor by inhibiting cell proliferation, invasion and by promoting apoptosis. However, the function and mechanisms responsible for the role of miR-1224 in HCC are still unknown.
Methods: miR-1224 expression levels were detected in HCC cells and tissues by quantitative real-time PCR (qPCR) and fluorescent in situ hybridization (FISH). Bisulfite sequencing PCR (BSP) was used for DNA methylation analysis. miR-1224 shRNAs and an overexpression vector were transfected into HCC cells in order to down-regulate or up-regulate the expression of miR-1224. In vitro and in vivo assays were performed to investigate the functional role of miR-1224 in the cell cycle progression of HCC cells. Dual-luciferase reporter assays were also used to determine the underlying targets and mechanisms regulated by miR-1224.
Results: miR-1224 was down-regulated in human HCC tissues. The methylation status of CpG islands was also found to be involved in the modulation of miR-1224 expression. Additionally, we found that the overexpression of miR-1224 induced HCC cells to arrest in G0/G1 phase and suppressed the proliferation of HCC cells while knockdown of miR-1224 promoted HCC cell proliferation both in vitro and in vivo. Moreover, mechanistic investigation showed that miR-1224 could repress YAP transcription and induce the HIPPO signaling pathway via binding with cAMP-response element binding protein (CREB).
Conclusions: Taken together, these findings show that miR-1224 not only inhibits the proliferation of HCC cells in vitro, but also inhibits the proliferation of these cells in vivo, at least in part, by suppressing the activation of Hippo/YAP signaling by targeting CREB.
Funding Statement: This study was supported by the Foundation of Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, the Priority Academic Program Development of Jiangsu Higher Education Institutions, the National Natural Science Foundation of China (81871259), Six talent peaks project in Jiangsu Province (WSW-019).
Declaration of Interests: The authors declare that they have no conflict of interest.
Ethics Approval Statement: This study was approved by the Ethics Committee of Nanjing Medical University and written informed consent was obtained from all enrolled patients prior to surgery. The recorded data did not contain any information that could identify individual patients.
Keywords: miR-1224, HCC, proliferation, Hippo/YAP signaling
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