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10 April 2017 : Laboratory Research

miR-4417 Targets Tripartite Motif-Containing 35 (TRIM35) and Regulates Pyruvate Kinase Muscle 2 (PKM2) Phosphorylation to Promote Proliferation and Suppress Apoptosis in Hepatocellular Carcinoma Cells

Lijie Song1ABCDEF, Weijie Zhang1ABCDEF, Zhiwei Chang1BCDF, Yanfeng Pan1BCD, Hong Zong1BCD, Qingxia Fan1BCF, Liuxing Wang1ACDG*

DOI: 10.12659/MSM.900296

Med Sci Monit 2017; 23:1741-1750

Abstract

BACKGROUND: MicroRNAs (miRNAs) are a class of small non-coding RNAs that are strongly involved in various types of carcinogenesis, including hepatocellular carcinoma (HCC). This study aimed to clarify whether miR-4417 promotes HCC growth by targeting TRIM35 and regulating PKM2 phosphorylation.

MATERIAL AND METHODS: Online software, including TargetScan and miRanda, was used to predict the potential target of miR-4417. Real-Time PCR (qRT-PCR) and Western blot assays were performed to detect the expression levels of mRNA and protein, respectively. Cell proliferation was measured by MTT assay and apoptosis in A549 cells was examined by flow cytometry.

RESULTS: Bioinformatics reveal that TRIM35 mRNA contains 1 conserved target site of miR-4417. High level of miR-4417 and low levels of TRIM35 mRNA and protein were observed in HCC cells compared with a normal liver cell line. Biological function analysis showed that miR-4417 inhibitor inhibits cell proliferation and promotes apoptosis in HCC cells. Furthermore, we verified that TRIM35 is a functional target of miR-4417 by use of luciferase reporter assay, and TRIM35 overexpressing showed an elevation of proliferation and a reduction of apoptosis in HCC cells. We subsequently investigated whether miR-4417 and TRIM35 regulate HCC cell proliferation and apoptosis through PKM2 Y105 phosphorylation, and the results supported our speculation that miR-4417 targets TRIM35 and regulates the Y105 phosphorylation of PKM2 to promote hepatocarcinogenesis.

CONCLUSIONS: Our findings indicate that miR-4417 may function as an oncogene in HCC and is a potential alternative therapeutic target for this deadly disease.

Keywords: Lim Kinases, Pyruvate Kinase

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Medical Science Monitor eISSN: 1643-3750
Medical Science Monitor eISSN: 1643-3750