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Background:  Cullin-based E3 ligases target substrates for ubiquitin-dependent proteasome-mediated degradation. The SCF (Skp1-Cul1-F box) and ECS (ElonginC-Cul2-SOCS box) complexes are well characterized cullin-based ligases. Cellular adaptor proteins assemble with ECS ligase through a SOCS box. While HIV-1 Vif uses a SOCS-box to recruit the Cul5-based E3 ligase, adenovirus E4orf6 and Vif proteins from most SIV lineages do not have a SOCS box. This poses the question as to whether these viral proteins can assemble with Cul5, ElonginB, and ElonginC to form a functional E3 ligase and degrade their respective targets.

Methods:  To examine whether these viral proteins were functional E3 ligase adaptors, we first determined whether lentiviral Vif proteins and adenovirus E4orf6 were able to co-immunoprecipitate with Cul5, ElonginB, and ElonginC. We then examined the effect of lentiviral Vif proteins on APOBEC3G stability and virion incorporation. Vif mutants were generated to identify the residues required for functional assembly with the E3 ligase. Cul5 dominant negative mutants were generated to examine the requirement for a Cul5 containing E3 ligase. We then examined the effect of E4orf6 wild type and mutant proteins on p53 stability.

Results:  We show that a viral-specific BC-box in primate lentiviral Vif is required to recruit Cul5-ElonginB-ElonginC and to suppress autologous APOBEC3G. Furthermore, we demonstrate that a similar BC-box motif in adenovirus E4orf6 is essential for its interaction with Cul5-ElonginB-ElonginC and the degradation of p53.

Conclusions:  These viral-specific BC-box motifs are highly divergent from the cellular SOCS boxes. Therefore, lentiviral Vif proteins and adenovirus E4orf6 represent novel families of viral BC-box adaptor proteins. The cellular targets of Cul5-containing E3 ligases remain elusive. The identification of these novel BC-box motifs that recruit the Cul5 complex may facilitate the identification of additional adaptor proteins and shed light on the cellular function of Cul5-containing E3 ligase.

Keywords: Cullin E3 Ubiquitin ligase; HIV-SIV Vif; APOBEC3G