Mutational analysis of the HIV-1 auxiliary protein Vif identifies independent domains important for the physical and functional interaction with HIV-1 reverse transcriptase.
The HIV-1 accessory protein Vif plays a dual role: it counteracts the natural restriction factors APOBEC3G and 3F and ensures efficient retrotranscription of the HIV-1 RNA genome. We have previously shown that Vif can act as an auxiliary factor for HIV-1 reverse transcriptase (RT), increasing its rate of association to RNA or DNA templates. Here, by using seven different Vif mutants, we provide in vitro evidences that Vif stimulates HIV-1 RT through direct protein-protein interaction, which is mediated by its C-terminal domain. Physical interaction appears to require the proline-rich region comprised between amino acid (aa) 161 and 164 of Vif, whereas the RT stimulatory activity requires, in addition, the extreme C-terminal region (aa 169-192) of the Vif protein. Neither the RNA interaction domain, nor the Zn(++)-binding domain of Vif are required for its interaction with the viral RT. Pseudotyped HIV-1 lentiviral vectors bearing Vif mutants deleted in the RNA- or RT-binding domains show defects in retrotranscription/integration processes in both permissive and nonpermissive cells. Our results broaden our knowledge on how three important functions of Vif (RNA binding, RT binding and stimulation and Zn(++) binding), are coordinated by different domains.
Kataropoulou
Alexandra
A
Institute of Molecular Genetics IGM-CNR, via Abbiategrasso 207, I-27100 Pavia, Italy.
Bovolenta
Chiara
C
Belfiore
Amalia
A
Trabatti
Sonia
S
Garbelli
Anna
A
Porcellini
Simona
S
Lupo
Rossella
R
Maga
Giovanni
G
D002460:Cell Line; D054303:HIV Reverse Transcriptase / Q000378:metabolism*; D006801:Humans; D009154:Mutation; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D012313:RNA / Q000378:metabolism; D048348:Reverse Transcription; D016662:Virus Integration; D016335:Zinc Fingers; D054320:vif Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D054320:vif Gene Products, Human Immunodeficiency Virus; C514847:vif protein, Human immunodeficiency virus 1; D012313:RNA; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase