Hepatitis C virus NS3 protein modulates the biological behaviors of malignant hepatocytes by altering the expression of host cell microRNA

Abstract

Abstract. Chronic hepatitis C virus (HCV) infection is a major cause of hepatocellular carcinoma (HCC) worldwide. The HCV non-structural protein 3 (NS3) protein is considered to affect normal cellular functions and to be involved in HCV carcinogenesis. The expression of microRNA (miRNA) is altered in human HCC, thus implicating its role in hepatocarcinogenesis. To investigate the mechanisms by which the HCV NS3 protein affects the expression of miRNA in malignant hepatocytes, if any, the present study constructed expression vectors encoding the HCV NS3 and NS3/4A proteins, which were stably transfected into HepG2 cells. The biological behaviors of the HepG2 transfectants and their differential expression levels of miRNA expression were investigated. Compared with the HepG2-vector cells, the HepG2-NS3 cells grew at a slower rate, were arrested in the G0/G1 cell cycle phase, formed more colonies and developed larger tumors at a faster rate. Co-expression of HCV NS4A resulted in the inhibition of HCV NS3-stimulated tumorigenicity. A total of 35 miRNAs were dysregulated, 26 of which were downregulated and nine of which were upregulated, in the HepG2-NS3 cells, and 75 miRNAs were altered in HepG2-NS3/4A cells, of which 20 were downregulated and 55 were upregulated). In addition, significant decreases in the mRNA levels of p53 and p21 were observed, which confirmed differential expression of miRNA. These results suggested that differential miRNA profiling in malignant hepatocytes may account for the variable pathophysiological manifestations associated with the HCV NS3 protein. These differentially expressed miRNAs may offer potential as candidates for the development of miRNA-based therapeutics.

HCV Proteins Modulate the Host Cell miRNA Expression Contributing to Hepatitis C Pathogenesis and Hepatocellular Carcinoma Development

“…In 2015, Zhang and colleagues studied the miRNome of HepG2 cells transfected either with a plasmid expressing the full-length NS3 protein or the NS3/4A protein. A total of 35 miRNAs were dysregulated, NS3 expressing HepG2 while 75 miRNAs were altered in HepG2 NS3/4A cells [100]. With the exceptions of miR-143, miR-181c, miR-181d, and miR153, the NS3 and the NS3/4A-expressing HepG2 had a differential miRNA profile with miR-130a and miR-153 being the most significantly altered miRNAs in HepG2-NS3 and HepG2-NS3/4A cells, respectively.…”

“…With the exceptions of miR-143, miR-181c, miR-181d, and miR153, the NS3 and the NS3/4A-expressing HepG2 had a differential miRNA profile with miR-130a and miR-153 being the most significantly altered miRNAs in HepG2-NS3 and HepG2-NS3/4A cells, respectively. The differences in the miRNA profiles of HepG2-NS3 and HepG2-NS3/4A may explain the differential cellular behavior, HepG2-NS3 having a more marked colony formation capacity and tumorigenicity in vivo than HepG2-NS3/4A cells [100]. Indeed, the up-regulation of miR-122, a well-known oncosuppressor, was only observed in NS3/4A-expressing cells [100].…”

“…Besides the effects of structural HCV proteins on the expression of miRNAs, some studies also reported the involvement of non-structural proteins in the modulation of miRNAs. In particular, NS3 and NS5 proteins showed numerous interactions with various cellular components that determine the alteration of intracellular miRNAs involved in immune response [55], fibrosis [99], and tumorigenesis [100].…”

“…The differences in the miRNA profiles of HepG2-NS3 and HepG2-NS3/4A may explain the differential cellular behavior, HepG2-NS3 having a more marked colony formation capacity and tumorigenicity in vivo than HepG2-NS3/4A cells [100]. Indeed, the up-regulation of miR-122, a well-known oncosuppressor, was only observed in NS3/4A-expressing cells [100]. MiR-122 represses HCC by targeting multiple genes involved in carcinogenesis [117], such as cyclin G1 [118] and ADAM Metallopeptidase Domain 17 (ADAM17) [119], involved in proliferation and EMT, respectively.…”

“…Interestingly, these results remain consistent with the observations of Zhang and colleagues that reported downregulation of miR-181c in HepG2 cells transfected either with NS3 protein or NS3/4A protein, thus evidencing the importance of a miR-181c inhibition in infected cells. However, it is not clear what the advantage is of this downregulation for the virus [100].…”