A conserved uORF regulates APOBEC3G translation and is targeted by HIV-1 Vif protein to repress the antiviral factor

Abstract

The HIV-1 Vif protein is essential for viral fitness and pathogenicity. Vif decreases expression of cellular cytosine deaminases APOBEC3G (A3G), A3F, A3D and A3H, which inhibit HIV-1 replication by inducing hypermutations during reverse transcription. Vif counteracts A3G by several non-redundant mechanisms (transcription, translation and protein degradation) that concur in reducing the levels of A3G in cell and in preventing its incorporation into viral particles. How Vif affects A3G translation remains unclear. Here, we uncovered the importance of a short conserved uORF (upstream ORF) located within two critical stem-loop structures of the 5′ untranslated region (5′UTR) of A3G mRNA. Extensive mutagenesis of A3G 5′-UTR, combined with an analysis of their translational effect in transfected cells, indicated that the uORF represses A3G translation and that A3G mRNA is translated through a dual leaky-scanning and re-initiation mechanism. Interestingly, the uORF is also mandatory for the Vif-mediated repression of A3G translation. Furthermore, we showed that the redirection of A3G mRNA into stress granules was dependent not only on Vif, but also on the uORF. Overall, we discovered that A3G translation is regulated by a small uORF conserved in the human population and that Vif uses this specific motif to repress its translation.

Degradation-Independent Inhibition of APOBEC3G by the HIV-1 Vif Protein

“…This association leads to A3G mRNA translation inhibition, possibly by ribosome stalling, and its relocation to RNP granules (P-bodies, stress granules, etc.). A3F mRNA could be regulated similarly as their 5'UTRs are phylogenetically conserved [102]. The involvement of other cellular factors in this process is still unknown but it could favor, or inhibit, A3G translation.…”

“…While mechanisms leading to A3G translation inhibition still remain unclear, we recently uncovered the importance of a short and conserved upstream ORF (uORF) located within SL2-SL3 of the 5'UTR of A3G and A3F mRNAs (Figure 5) [102]. Interestingly, the uORF represses A3G translation itself (40%), as do many uORF located in 5'UTR of mRNAs [103][104][105], but it is also mandatory for the Vif-mediated repression of A3G translation and for the redirection of A3G mRNA into stress granules in the presence of Vif [102]. The fact that Vif was previously shown in vitro to interact with the lower and upper stems of SL3 [98] suggests that Vif may block ribosome scanning (stalling).…”