Quantification of the HIV-1 Vif - ElonginC interaction by a new on bimolecular fluorescence complementation (BiFC) based assay

The human immunodeficiency virus type 1 (HIV-1) virion infectivity factor (Vif) is known to rescue viral infectivity by triggering the ubiquitin-mediated proteasomal degradation of the antiviral active cellular cytidine deaminases (APOBEC3 protein family; A3D, E, F and G). During this Vif induced degradation mechanism of A3 proteins, the cellular adaptor protein Elongin C plays a key role in connecting the Vif bound A3 to a SCF-like E3 ubiquitin ligase complex composed of Rbx2, Cullin 5, Elongin B and Elongin C. Degradation of A3 proteins can only take place when this complex is properly assembled. Hence, blocking the Vif-ElonginC interaction is predicted to rescue the antiviral activity of the APOBEC3 deaminases. In order to quantify the direct interaction between Vif and ElonginC, we developed a bimolecular fluorescence complementation (BiFC) based cellular assay, that can be expanded to a HTS platform, to detect potential novel anti-retroviral (-HIV) compounds. To properly validate our test system, we investigated the effect of different Vif and ElonginC mutants on the interaction between both proteins. The here described BiFC assay can be used to quantitatively characterize the Vif-ElonginC interaction and to screen compound libraries for potential Vif-ElonginC interaction antagonists. Hit compounds may serve as lead candidates for the development of novel antiretroviral agents based on the Vif-ElonginC interaction.