The C-terminal p6 domain of the HIV-1 Pr55(Gag) precursor is required for specific binding to the genomic RNA
| Autor: | Serena Bernacchi, Philippe Wolff, William J. McKinstry, Roland Marquet, Noé Dubois, Jean-Christophe Paillart, Tanja Seissler, Keith K. Khoo, Shannon Ghossein, Johnson Mak |
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| Přispěvatelé: | Université Lumière - Lyon 2 (UL2), Centre for Virology, Macfarlane Burnet Institute, Architecture et réactivité de l'ARN (ARN), Centre National de la Recherche Scientifique (CNRS)-Université Louis Pasteur - Strasbourg I |
| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
[SDV]Life Sciences [q-bio] viruses Genome Viral Biology gag Gene Products Human Immunodeficiency Virus Virus Domain (software engineering) law.invention 03 medical and health sciences law Humans Protein Interaction Domains and Motifs Guide RNA Protein Precursors Molecular Biology Binding selectivity ComputingMilieux_MISCELLANEOUS Fluorescent Dyes Binding Sites 030102 biochemistry & molecular biology Wild type RNA Cell Biology Recombinant Proteins 3. Good health Cell biology 030104 developmental biology Mutation Recombinant DNA HIV-1 RNA Viral Genomic rna Research Paper Plasmids Protein Binding |
| Zdroj: | RNA Biology RNA Biology, Taylor & Francis, 2018, 15 (7), pp.923-936. ⟨10.1080/15476286.2018.1481696⟩ |
| ISSN: | 1547-6286 1555-8584 |
| Popis: | The Pr55(Gag) precursor specifically selects the HIV-1 genomic RNA (gRNA) from a large excess of cellular and partially or fully spliced viral RNAs and drives the virus assembly at the plasma membrane. During these processes, the NC domain of Pr55(Gag) interacts with the gRNA, while its C-terminal p6 domain binds cellular and viral factors and orchestrates viral particle release. Gag∆p6 is a truncated form of Pr55(Gag) lacking the p6 domain usually used as a default surrogate for wild type Pr55(Gag) for in vitro analysis. With recent advance in production of full-length recombinant Pr55(Gag), here, we tested whether the p6 domain also contributes to the RNA binding specificity of Pr55(Gag) by systematically comparing binding of Pr55(Gag) and Gag∆p6 to a panel of viral and cellular RNAs. Unexpectedly, our fluorescence data reveal that the p6 domain is absolutely required for specific binding of Pr55(Gag) to the HIV-1 gRNA. Its deletion resulted not only in a decreased affinity for gRNA, but also in an increased affinity for spliced viral and cellular RNAs. In contrast Gag∆p6 displayed a similar affinity for all tested RNAs. Removal of the C-terminal His-tag from Pr55(Gag) and Gag∆p6 uniformly increased the Kd values of the RNA-protein complexes by ~ 2.5 fold but did not affect the binding specificities of these proteins. Altogether, our results demonstrate a novel role of the p6 domain in the specificity of Pr55(Gag)-RNA interactions, and strongly suggest that the p6 domain contributes to the discrimination of HIV-1 gRNA from cellular and spliced viral mRNAs, which is necessary for its selective encapsidation. |
| Databáze: | OpenAIRE |
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