Interferon-stimulated gene (ISG12a) suppresses hepatitis B virus replication in Huh 7 cells line

Abstract

Hepatitis B virus (HBV) and its associated chronic liver disease pose a significant health hazard. Chronic HBV infection is a major contributor to the development of liver cirrhosis, liver fibrosis, and liver cancer. This study focused on ISG12a and its inhibitory effect in Huh7 cells on HBV gene expression and replication. The mammalian hepatoma cells Huh7 were transfected with the pHBV1.3 vector (pCAGGS) or HA-tagged ISG12a to determine the overexpression, to test this hypothesis further, we did knockdown or silencing by transducing Huh7 cells with lentiviruses containing shRNAs targeting ISG12a or scramble control shRNA (shControl). The expression of ISG12a was evaluated through western blot analysis. The HBsAg and HBeAg secretion in the Huh7 cells’ culture media was examined using an ELISA test. The qRT-PCR confirmed the mRNA and 3.5 mRNA Kb of ISG12a. The HBV total RNA was extracted and evaluated through a Northern blot. The isolated DNA was detected through qPCR. Additionally, a mechanistic study of enhancer II (EnhII/Cp) was examined through luciferase reporter testing. Our research findings indicate that ISG12a, an interferon-stimulated gene (ISG), plays a crucial role in suppressing the replication and gene expression of HBV. According to our study using the Huh7 cell system, overexpression of ISG12a resulted in a notable reduction in HBV protein levels as well as intracellular core-associated DNA and RNA levels. On the other hand, silencing ISG12a in Huh7 cells resulted in increased HBV RNA transcripts, DNA, and secreted proteins, indicating that ISG12a plays a role in suppressing HBV replication. Additionally, the research revealed that ISG12a inhibits the function of the EnhII/Cp promoter, which results in reduced HBV gene expression. The EnhII/Cp promoter is involved in regulating HBV gene expression, and the study showed that ISG12a restricts its activity. ISG12a may have a regulatory role in controlling the expression of HBV genes. These findings highlight the importance of ISG12a in HBV gene expression and provide valuable insights for understanding its antiviral function.

Keywords

• Hepatitis B virus
• ISG12a
• Huh 7 cells line
• DNA replication
• Gene silencing