MicroRNAs (miRNAs) and methionine adenosyltransferase 1A (MAT1A) are dysregulated in hepatocellular carcinoma (HCC), and reduced MAT1A expression correlates with worse HCC prognosis. Expression of miR-664, miR-485-3p, and miR-495, potential regulatory miRNAs of MAT1A, is increased in HCC. Knockdown of these miRNAs individually in Hep3B and HepG2 cells induced MAT1A expression, reduced growth, and increased apoptosis, while combined knockdown exerted additional effects on all parameters. Subcutaneous and intraparenchymal injection of Hep3B cells stably overexpressing each of this trio of miRNAs promoted tumorigenesis and metastasis in mice. Treatment with miRNA-664 (miR-664), miR-485-3p, and miR-495 siRNAs reduced tumor growth, invasion, and metastasis in an orthotopic liver cancer model. Blocking MAT1A induction significantly reduced the antitumorigenic effect of miR-495 siRNA, whereas maintaining MAT1A expression prevented miRNA-mediated enhancement of growth and metastasis. Knockdown of these miRNAs increased total and nuclear level of MAT1A protein, global CpG methylation, lin-28 homolog B (Caenorhabditis elegans) (LIN28B) promoter methylation, and reduced LIN28B expression. The opposite occurred with forced expression of these miRNAs. In conclusion, upregulation of miR-664, miR-485-3p, and miR-495 contributes to lower MAT1A expression in HCC, and enhanced tumorigenesis may provide potential targets for HCC therapy.
Heping Yang, Michele E. Cho, Tony W.H. Li, Hui Peng, Kwang Suk Ko, Jose M. Mato, Shelly C. Lu
(A) Northern blot analysis showing expression of select miRNAs in HCC compared with adjacent nontumorous (NT) tissue. (B) Northern blot analysis confirming siRNA knockdown efficiency of miR-664, miR-485-3p, and miR-495 in HepG2 and Hep3B cells as compared with scramble siRNA (SC) control. (C and D) Northern (top) and Western (bottom) blot analyses showing the effect of siRNA knockdown of miR-664, miR-485-3p and miR-495, alone or in combination, on MAT1A expression in HepG2 (C) and Hep3B cells (D). Numbers below the blots represent densitometric values expressed as percentage of respective controls. Representative blots are shown for C and D from 3 experiments, *P < 0.01 vs. SC; †P < 0.05 vs. SC, and triple knockdown; ‡P < 0.001 vs. SC and single knockdown.