Background:  Infection with GB virus C (GBV-C) is associated with prolonged survival among HIV-infected individuals in 10 of 13 studies. GBV-C co-infection of primary lymphocytes with HIV in vitro results in inhibition of HIV replication, and this is mediated by induction of chemokines and downregulation of CCR5. Chemokine regulation appears to be partially mediated by the GBV-C envelope glycoprotein E2, although inhibition is not as great as with viral infection. To determine whether other GBV-C proteins play a role in HIV inhibition, we evaluated the effect of the GBV-C NS5A phosphoprotein on HIV replication.

Methods:  A stable Jurkat cell line that inducibly expressed GBV-C NS5A protein (by removing tetracycline) was generated. A control cell line stably transfected with the vector served as the control. Cells were maintained in tetracycline or tetracycline was removed to induce NS5A expression. HIV replication was measured by p24 antigen in Jurkat cells supernatants.

Results:  Jurkat cells grown without tetracycline expressed NS5A as determined by Western blot. In addition to the phosphorylated form, a hyperphosphorylated NS5A was identified. Jurkat cells expressing NS5A had up to 40% inhibition of HIV replication. This reduction in HIV p24 Ag release was significant when compared to cells expressing the vector control with or without tetracycline (p = 0.008 and 0.02, respectively), and when compared to the stably transfected cells in which NS5A expression was inhibited with tetracycline (p = 0.014). The extent of HIV inhibition was related to the amount of NS5A expression observed using Immunoblot analysis.

Conclusions:  GBV-C appears to inhibit HIV replication by multiple pathways including GBV-C replication, E2 protein interactions with cell surface receptors, and by effects of NS5A on cellular milieu. HIV inhibition was greatest in peripheral blood mononuclear cells (PBMC) co-infected with GBV-C and HIV; however, expression of NS5A alone resulted in diminished replication. Understanding how GBV-C inhibits HIV replication may identify novel HIV therapeutic strategies, and further evaluation of the effects of GBV-C NS5A protein on cellular gene expression is underway.

Keywords: GB Virus C; Hepatitis G; NS5A protein