Preliminary study on effect of hepatitis C virus nonstructural protein 5A on HIV LTR

Objective To investigate the effect of nonstructural protein 5A ( NS5A) encoded by the human hepatitis C virus ( HCV) RNA genome on human immunodeficiency virus ( HIV) long terminal repeat ( LTR) and to provide an experimental basis for the study on the effect of HCV on HIV. Methods Hepatocellular carcinoma Huh7 cells were divided into blank group, control group, and experimental group to be transfected with plasmid pGL3- LTR- Luc ( containing luciferase reporter gene driven by the LTR promoter) , plasmid pRc / CMV plus plasmid pGL3- LTR- Luc, and plasmid pCNS5A ( containing HCV NS5A gene) plus plasmid pGL3- LTR- Luc, respectively; Huh7 cells were collected 48 h later. The protein and mRNA expression levels of HCV NS5A were measured by immunocytochemistry, Western blot, and RT- PCR. The relative luciferase activity was measured to evaluate the HIV LTR activity and the effect of HCV NS5A on HIV LTR. The activity values were expressed as mean ± SD, and Levene' s test of homogeneity of variance was used; comparison between all groups was made by one- way analysis of variance ( ANOVA) , and comparison between two groups was made by least significant difference ( LSD) test. Results The mRNA and protein expression of HCV NS5A was detected in the cytoplasm of Huh7 cells in experimental group. The one- way ANOVA showed that there were significant differences in LTR luciferase activity between the three groups ( F = 7. 876, P = 0. 002) . The LSD test showed that the experimental group had a significantly higher relative luciferase activity than the blank group and control group ( 22476 ± 4471 vs 15887 ± 3039, P = 0. 002; 22476 ± 4471 vs 16321 ± 4162, P = 0. 008) . Conclusion Huh7 cells can be transfected with the HCV NS5A expression plasmid ( pCNS5A) . HCV NS5A can activate HIV LTR, which suggests that HCV NS5A may be one of the molecular mechanisms of HCV promoting HIV replication.