RNA and DNA binding properties of HIV-1 Vif protein: a fluorescence study.
The HIV-1 viral infectivity factor (Vif) is a small basic protein essential for viral fitness and pathogenicity. Some "non-permissive" cell lines cannot sustain replication of Vif(-) HIV-1 virions. In these cells, Vif counteracts the natural antiretroviral activity of the DNA-editing enzymes APOBEC3G/3F. Moreover, Vif is packaged into viral particles through a strong interaction with genomic RNA in viral nucleoprotein complexes. To gain insights into determinants of this binding process, we performed the first characterization of Vif/nucleic acid interactions using Vif intrinsic fluorescence. We determined the affinity of Vif for RNA fragments corresponding to various regions of the HIV-1 genome. Our results demonstrated preferential and moderately cooperative binding for RNAs corresponding to the 5'-untranslated region of HIV-1 (5'-untranslated region) and gag (cooperativity parameter omega approximately 65-80, and K(d) = 45-55 nM). In addition, fluorescence spectroscopy allowed us to point out the TAR apical loop and a short region in gag as primary strong affinity binding sites (K(d) = 9.5-14 nM). Interestingly, beside its RNA binding properties, the Vif protein can also bind the corresponding DNA oligonucleotides and their complementary counterparts with an affinity similar to the one observed for the RNA sequences, while other DNA sequences displayed reduced affinity. Taken together, our results suggest that Vif binding to RNA and DNA offers several non-exclusive ways to counteract APOBEC3G/3F factors, in addition to the well documented Vif-induced degradation by the proteasome and to the Vif-mediated repression of translation of these antiviral factors.
Bernacchi
Serena
S
Architecture et Réactivité de l'ARN, Université Louis Pasteur de Strasbourg, CNRS, IBMC, 15 Rue René Descartes, 67084 Strasbourg, France.
Henriet
Simon
S
Dumas
Philippe
P
Paillart
Jean-Christophe
JC
Marquet
Roland
R
D020121:5' Untranslated Regions / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D000071480:APOBEC-3G Deaminase; D001665:Binding Sites / Q000502:physiology; D003564:Cytidine Deaminase; D043525:Cytosine Deaminase / Q000276:immunology / Q000378:metabolism; D004279:DNA, Viral / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D016346:Gene Products, vif / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016679:Genome, Viral / Q000502:physiology; D016325:HIV Long Terminal Repeat / Q000502:physiology; D015497:HIV-1 / Q000235:genetics / Q000276:immunology / Q000378:metabolism* / Q000472:pathogenicity; D006801:Humans; D007113:Immunity, Innate / Q000502:physiology; D009700:Nucleoside Deaminases / Q000276:immunology / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D014176:Protein Biosynthesis / Q000502:physiology; D012367:RNA, Viral / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D054320:vif Gene Products, Human Immunodeficiency Virus
D020121:5' Untranslated Regions; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D015683:Gene Products, gag; D016346:Gene Products, vif; D009841:Oligonucleotides; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; D054320:vif Gene Products, Human Immunodeficiency Virus; D009700:Nucleoside Deaminases; C492906:APOBEC3F protein, human; D043525:Cytosine Deaminase; D000071480:APOBEC-3G Deaminase; C464924:APOBEC3G protein, human; D003564:Cytidine Deaminase