Hepatitis B Virus-Encoded MicroRNA (HBV-miR-3) Regulates Host Gene PPM1A Related to Hepatocellular Carcinoma

Author(s): Tanit Chavalit, Pattaraporn Nimsamer, Kritsada Sirivassanametha, Songtham Anuntakarun, Suthat Saengchoowong, Pisit Tangkijvanich, Sunchai Payungporn*.

Journal Name: MicroRNA

Volume 9 , Issue 3 , 2020

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Abstract:

Background: Hepatitis B is a liver infection disease caused by the Hepatitis B Virus (HBV) that can become chronic and develop into hepatocellular carcinoma. HBV was classified as a double-stranded DNA virus. Currently, there is a report showing that HBV virus-encoded miRNA called HBV-miR-3 controls the replication of HBV. However, the regulation of HBV-miR-3 in host cells remains unclear.

Objective: This study aimed to investigate the regulation of HBV-miR-3 in host gene target which is related to chronic HBV infection and HCC process.

Methods: In this study, we analyzed the read count of HBV-miR-3 from next-generation sequencing of chronic hepatitis patients in Pegylated interferon alpha-2a (PEG-IFN-α-2a) treatment. To understand the regulation of HBV-miR-3 in host cells, the HBV-miR-3 recognition sites were predicted in host target genes using miRDB. The effect of HBV-miR-3 in host cells was examined using qPCR and 3′ UTR dual luciferase assay.

Results: The read count of HBV-miR-3 was found in chronic hepatitis patients before treatment. Moreover, the decrease of HBV-miR-3 was correlated with response group of chronic hepatitis patients after treatment. On the other hand, the abundance of HBV-miR-3 showed no difference in nonresponse group of chronic patients after PEG-IFN-α-2a treatment. To study the role of HBV-miR-3 in patients, four HBV-miR-3 target regions from Protein phosphatase 1A (PPM1A) and DIX domain containing 1 (DIXDC1) were identified in the human genome using miRDB. Interestingly, we found that HBV-miR-3 hybridized with PPM1A mRNA. The mRNA expression from RT-qPCR showed no difference between HepG2 transfected with pSilencer_scramble or pSilencer_HBV-miR-3. However, the reporter assay showed that PPM1A mRNA was suppressed by HBV-miR-3. The protein expression of PPM1A showed a decrease in cells overexpressing HBV-miR-3. Finally, the HBV-miR-3 can promote cell proliferation in cells overexpressing HBV-miR-3.

Conclusion: This study is the first report showed the HBV encoded miRNA can regulate host gene expression. HBV-miR-3 silenced PPM1A by inhibiting the translation process of PPM1A. The downregulation of PPM1A promotes cell proliferation related to HCC development.

Keywords: HBV-miR-3, HCC, HepG2 cells, miRNA, PPM1A, chronic.

[1]
Vojtechova Z, Tachezy R. The role of miRNAs in virus-mediated oncogenesis. Int J Mol Sci 2018; 19(4): 1217.
[http://dx.doi.org/10.3390/ijms19041217]
[2]
Schädler S, Hildt E. HBV life cycle: entry and morphogenesis. Viruses 2009; 1(2): 185-209.
[http://dx.doi.org/10.3390/v1020185] [PMID: 21994545]
[3]
Chen YJ, Chien PH, Chen WS, et al. Hepatitis B virus-encoded X protein downregulates EGFR expression via inducing microRNA-7 in hepatocellular carcinoma cells. Evid Based Complement Alternat Med 2013; 2013: 682380
[http://dx.doi.org/10.1155/2013/682380] [PMID: 23840262]
[4]
Ambros VJN. The functions of animal microRNAs. Nature 2004; 431(7006): 350-5.
[http://dx.doi.org/10.1038/nature02871]
[5]
Bartel DPJc. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004; 116(2): 281-97.
[6]
Jin Y, Chen Z, Liu X. Evaluating the microRNA targeting sites by luciferase reporter gene assay. Methods Mol Biol 2013; 936: 117-27.
[http://dx.doi.org/10.1007/978-1-62703-083-0_10]
[7]
Eichhorn SW, Guo H, McGeary SE, et al. mRNA destabilization is the dominant effect of mammalian microRNAs by the time substantial repression ensues. Mol Cell 2014; 56(1): 104-15.
[8]
Ardekani AM, Naeini MM. The role of microRNAs in human diseases. Avicenna J Med Biotechnol 2010; 2(4): 161-79.
[9]
Assalin HB, Gontijo JAR, Boer PAJPo. miRNAs, target genes expression and morphological analysis on the heart in gestational protein-restricted offspring. PLoS One 2019; 14(4): e0210454
[10]
Yang X, Li H, Sun H, et al. Hepatitis B virus-encoded microRNA controls viral replication. J Virol 2017; 91(10): e01919-16.
[http://dx.doi.org/10.1128/JVI.01919-16] [PMID: 28148795]
[11]
Kozomara A, Griffiths-Jones SJNar. miRBase: annotating high confidence microRNAs using deep sequencing data. Nucleic Acids Res 2014; 42(Database issue): D68-73.
[12]
Langmead B, Trapnell C, Pop M, Salzberg SL. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol 2009; 10(3): R25.
[http://dx.doi.org/10.1186/gb-2009-10-3-r25]
[13]
Rao M, Zeng Z, Tang L, Cheng G, Xia W, Zhu C. Next-generation sequencing-based microRNA profiling of mice testis subjected to transient heat stress. Oncotarget 2017; 8(67): 111672-82.
[http://dx.doi.org/10.18632/oncotarget.22900] [PMID: 29340083]
[14]
Khongnomnan K, Poomipak W, Praianantathavorn K, et al. Human microRNAs expression profiles in influenza B virus-infected cells based on illumina MiSeq platform. MicroRNA 2018; 7(3): 204-14.
[http://dx.doi.org/10.2174/2211536607666180515111048] [PMID: 29766833]
[15]
Jinato T, Chuaypen N, Poomipak W, et al. Original Research: Analysis of hepatic microRNA alterations in response to hepatitis B virus infection and pegylated interferon alpha-2a treatment. Exp Biol Med (Maywood) 2016; 241(16): 1803-10.
[http://dx.doi.org/10.1177/1535370216647184] [PMID: 27190255]
[16]
Wong N, Wang X. miRDB: an online resource for microRNA target prediction and functional annotations. Nucleic Acids Res 2015; 43(Database issue): D146-52.
[http://dx.doi.org/10.1093/nar/gku1104] [PMID: 25378301]
[17]
Rehmsmeier M, Steffen P, Hochsmann M, Giegerich R. Fast and effective prediction of microRNA/target duplexes. RNA 2004; 10(10): 1507-17.
[http://dx.doi.org/10.1261/rna.5248604] [PMID: 15383676]
[18]
Liu Y, Xu Y, Ma H, et al. Hepatitis B virus X protein amplifies TGF-β promotion on HCC motility through down-regulating PPM1a. Oncotarget 2016; 7(22): 33125-35.
[http://dx.doi.org/10.18632/oncotarget.8884] [PMID: 27121309]
[19]
Zhang Y, Tao R, Wu SS, et al. TRIM52 up-regulation in hepatocellular carcinoma cells promotes proliferation, migration and invasion through the ubiquitination of PPM1A. J Exp Clin Cancer Res 2018; 37(1): 116.
[20]
Zhou S, Shen J, Lin S, et al. Downregulated expression of DIXDC1 in hepatocellular carcinoma and its correlation with prognosis. Tumour Biol 2016; 37(10): 13607-16.
[http://dx.doi.org/10.1007/s13277-016-5213-9]
[21]
Seeger C, Mason WS. Hepatitis B virus biology. Microbiol Mol Biol Rev 2000; 64(1): 51-68.
[http://dx.doi.org/10.1128/MMBR.64.1.51-68.2000]
[22]
Ganem D, Prince AM. Hepatitis B virus infection--natural history and clinical consequences. N Engl J Med 2004; 350(11): 1118-29.
[23]
Sekiba K, Otsuka M, Ohno M, et al. Hepatitis B virus pathogenesis: Fresh insights into hepatitis B virus RNA. World J Gastroenterol 2018; 24(21): 2261-8.
[http://dx.doi.org/10.3748/wjg.v24.i21.2261] [PMID: 29881235]
[24]
Decorsière A, Mueller H, van Breugel PC, et al. Hepatitis B virus X protein identifies the Smc5/6 complex as a host restriction factor 2016; 531(7594): : 386.
[25]
Benhenda S, Cougot D, Buendia MA, Neuveut C. Hepatitis B virus X protein molecular functions and its role in virus life cycle and pathogenesis. Adv Cancer Res 2009; 103: 75-109.
[26]
Diab A, Foca A, Zoulim F, Durantel D, Andrisani O. The diverse functions of the hepatitis B core/capsid protein (HBc) in the viral life cycle: Implications for the development of HBc-targeting antivirals. Antiviral Res 2018; 149: 211-20.
[27]
Guo Y, Kang W, Lei X, et al. Hepatitis B viral core protein disrupts human host gene expression by binding to promoter regions. BMC Genomics 2012; 13(1): 563.
[http://dx.doi.org/10.1186/1471-2164-13-563]
[28]
Sekiba K, Otsuka M, Ohno M, et al. Hepatitis B virus pathogenesis: Fresh insights into hepatitis B virus RNA. World J Gastroenterol 2018; 24(21): 2261-8.
[http://dx.doi.org/10.3748/wjg.v24.i21.2261] [PMID: 29881235]
[29]
Yao L, Zhou Y, Sui Z, et al. HBV-encoded miR-2 functions as an oncogene by downregulating TRIM35 but upregulating RAN in liver cancer cells. EBioMedicine 2019; 48: 117-29.
[30]
Mann DJ, Campbell DG, McGowan CH, Cohen PT. Mammalian protein serine/threonine phosphatase 2C: cDNA cloning and comparative analysis of amino acid sequences. Biochim Biophys Acta 1992; 1130(1): 100-4.
[31]
Lin X, Duan X, Liang YY, et al. PPM1A functions as a Smad phosphatase to terminate TGFbeta signaling. Cell 2006; 125(5): 915-28.
[http://dx.doi.org/10.1016/j.cell.2006.03.044] [PMID: 16751101]
[32]
Fabregat I, Fernando J, Mainez J, Sancho P. TGF-beta signaling in cancer treatment. Curr Pharm Des 2014; 20(17): 2934-47.
[http://dx.doi.org/10.2174/13816128113199990591]
[33]
Massagué JJC. TGFβ in Cancer. Cell 2008; 134(2): 215-30.
[http://dx.doi.org/10.1016/j.cell.2008.07.001]
[34]
Zhou Y, Zhao Y, Gao Y, et al. Hepatitis C virus NS3 protein enhances hepatocellular carcinoma cell invasion by promoting PPM1A ubiquitination and degradation. J Exp Clin Cancer Res 2017; 36(1): 42.
[http://dx.doi.org/10.1186/s13046-017-0510-8] [PMID: 28283039]


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Article Details

VOLUME: 9
ISSUE: 3
Year: 2020
Page: [232 - 239]
Pages: 8
DOI: 10.2174/2211536608666191104105334

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