A Novel GSK-3 beta–C/EBP alpha–miR-122–Insulin-Like Growth Factor 1 Receptor Regulatory Circuitry in Human Hepatocellular Carcinoma

Abstract

1 miR-122 is a highly abundant, hepatocyte-specific microRNA. The biomedical significance and regulatory mechanisms of miR-122 remain obscure. We explored the role of miR-122 in tumorigenesis in the context of gene regulatory network. The miR-122 promoter and its transactivator were identified by way of luciferase reporter system, electrophoretic mobility shift, and chromatin immunoprecipitation assays. The miR-122 regulatory circuitry and its implication in hepatocarcinogenesis were identified using livers of different development stages, human hepatocellular carcinoma (HCC) tissues and cell lines, and aflatoxin B 1 (AFB 1 )-transformed cells. We characterized the 25.3 to 24.8 kb region upstream of miR-122 precursor as miR-122 promoter. Further investigation revealed that deletion of predicted CCAAT/enhancer-binding protein alpha (C/EBPa) binding sites C/EBPa knockdown significantly reduced miR-122 promoter activity and endogenous miR-122 expression; and C/ EBPa directly interacted with the miR-122 promoter in vitro and in vivo. These data suggest that C/EBPa is a transactivator for miR-122 transcription. We further demonstrated that miR-122 suppressed insulin-like growth factor 1 receptor (IGF-1R) translation and sustained glycogen synthase kinase-3 beta (GSK-3b) activity. The activated GSK-3b not only repressed cell proliferation, but also activated C/EBPa, which maintained miR-122 levels and thereby enforced IGF-1R suppression. Interestingly, down-regulation of miR-122 and C/EBPa, and up-regulation of IGF-1R were frequently observed in HCC tissues, and decreased miR-122 levels were associated with worse survival of HCC patients. Moreover, AFB 1 exposure resulted in decreased activity in GSK-3b, C/EBPa, and miR-122 and increased levels of IGF-1R, whereas restoration of miR-122 suppressed the tumorigenicity of HCC and AFB 1 -transformed cells. Conclusion: We have identified a novel GSK-3b-C/EBPa-miR-122-IGF-1R regulatory circuitry whose dysfunction may contribute to the development of HCC. Our findings provide new insight into miR-122's function and the mechanisms of hepatocarcinogenesis. (HEPATOLOGY 2010;52:1702-1712

Retracted: MicroRNA‐140‐5p elevates cerebral protection of dexmedetomidine against hypoxic–ischaemic brain damage via the Wnt/β‐catenin signalling pathway

“…The reversely transcribed cDNA were temporarily stored in a refrigerator at −20°C. The RT‐qPCR prime sequences are displayed in Table 1 using TaqMan probes 18. The reaction system was performed according to the instructions of TaqMan RT‐PCR kit (No.…”

MiR-122 in hepatic function and liver diseases

“…Using a genome-wide expression profiling approach, Burchard et al (2010) have recently shown decreased miR-122 expression and increased miR-122 predicted target genes in tumor tissues compared with nontumor tissues from HCC, which may possibly lead to impairment of mitochondrial metabolism. Loss of miR-122 expression in HCC, which is associated with poor prognosis and metastasis, has been confirmed by several other studies (Coulouarn et al, 2009;Bai et al, 2009;Zeng et al, 2010;Mizuguchi et al, 2011;Karakatsanis et a., 2011). In these studies, miR-122 is shown to be involved in cell proliferation, apoptosis, clonogenic survival, migration, in vivo invasion, and tumor formation in nude mice.…”

“…Loss of miR-122 facilitates cell tumorigenic properties such as cell migration and invasion, and restoration of miR-122 reverses this phenotype. Currently, several target genes of miR-122 have been identified to be involved in hepatocarcinogenesis, such as ADAM10 (a distintegrin and metalloprotease family 10), serum response factor (SRF) (Bai et al, 2009), insulin-like growth factor 1 receptor (Igf1R) (Zeng et al, 2010), cyclin G1 (Fornari et al, 2009), and Wnt1 (Xu et al, 2012). In addition, overexpression and restoration of miR-122 in HCC cells has been shown to sensitize HCC cells to chemotherapeutic agents (Bai et al, 2009;Xu et al, 2011;Yang et al, 2011).…”

Upregulation of miR-27a contributes to the malignant transformation of human bronchial epithelial cells induced by SV40 small T antigen

“…The existence of corresponsive PP2A B56g-miR-27a-Fbxw7 regulatory axis in human lung tumors further confirms that B56g subunit is indispensable in ST-induced upregulation of miR-27a. Recently, we have demonstrated that the dysfunction of a novel GSK-3b-C/EBPa-miR-122-IGF-1R regulatory circuitry contributes to the development of hepatocellular carcinoma (Zeng et al, 2010). We identified that C/EBPa, the activity of which is controlled by the status of phosphorylation, directly binds to the promoter of the miR-122 gene and initiates transcription.…”