3 years ago

MicroRNA-130a regulates both HCV and HBV replication through a central metabolic pathway.

Raymond T Chung, Baihai Jiao, Wenyu Lin, Zeng Tu, Jacinta A Holmes, Dachuan Cai, Wenting Li, Shilin Li, Esperance A Schaefer, Xiaoqiong Duan, Limin Chen, Chunhui Yang, Dahlene N Fusco, Shadi Salloum, Xiao Liu, Yujia Li
BACKGROUND: HCV infection has been shown to regulate miR-130a in patient biopsies and in cultured cells. We sought to identify miR-130a target genes and to explore the mechanisms by which miR-130a regulates HCV and HBV replication.METHODS: We used bioinformatics software including miRanda, TargetScan, PITA and RNAhybrid to predict potential miR-130a target genes. miR-130a and its target genes were overexpressed, or knocked down by siRNA or by CRISPR/Cas9 gRNA, respectively. Selected gene mRNAs and their proteins, together with HCV replication in OR6 cells, JFH1 HCV-infected Huh7.5.1 cells and JFH1 HCV-infected primary human hepatocytes (PHHs), and HBV replication in HepAD38 cells, HBV-infected NTCP-Huh7.5.1 cells and HBV-infected PHHs were measured by qRT-PCR and Western blot, respectively.RESULTS: We selected 116 predicted target genes whose expression was related to viral pathogenesis or immunity for qPCR validation. Of these, pyruvate kinase in liver and red blood cell (PKLR) was confirmed to be regulated by miR-130a overexpression. miR-130a overexpression (mimic) knocked down PKLR mRNA and protein levels. miR-130a inhibitor and gRNA increased PKLR expression, HCV replication and HBV replication, while miR-130a gRNA and PKLR overexpression increased HCV and HBV replication. Supplemental pyruvate increased HCV and HBV replication, and rescued the inhibition of HCV and HBV replication by miR-130a mimic and PKLR knockdown.CONCLUSION: miR-130a regulates HCV and HBV replication through its targeting of PKLR and subsequent pyruvate production. Our data provides novel insights into understanding key metabolic enzymatic pathway steps regulated by miR-130a, including PKLR and pyruvate, which are subverted by HCV and HBV replication.Importance:We identified that miR-130a regulates its target gene PKLR and its subsequent pyruvate production. Pyruvate is a key intermediate in several metabolic pathways, and we identified that pyruvate plays a key role in regulation of HCV and HBV replication. This previously unrecognized miRNA regulated antiviral mechanism has implications for the development of host-directed strategies to interrupt the viral lifecycle and prevent establishment of persistent infection for HCV, HBV and, potentially, other viral infections.

Publisher URL: http://doi.org/10.1128/JVI.02009-17

DOI: 10.1128/JVI.02009-17

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