The ESCRT-Associated Protein Alix Recruits the Ubiquitin Ligase Nedd4-1 To Facilitate HIV-1 Release through the LYPX n L L Domain Motif
| Autor: | Fadila Bouamr, Paola Sette, Vincent Dussupt, Joshua A. Jadwin, Nana F. Bello |
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| Rok vydání: | 2010 |
| Předmět: |
Immunoprecipitation
Nedd4 Ubiquitin Protein Ligases Ubiquitin-Protein Ligases Immunology Cell Cycle Proteins NEDD4 macromolecular substances gag Gene Products Human Immunodeficiency Virus Microbiology ESCRT Cell Line Ubiquitin Two-Hybrid System Techniques Virology Protein Interaction Mapping Humans TSG101 Virus Release Endosomal Sorting Complexes Required for Transport biology Structure and Assembly Calcium-Binding Proteins Molecular biology Ubiquitin ligase Cell biology Gene Knockdown Techniques Insect Science Host-Pathogen Interactions HIV-1 biology.protein Protein Binding |
| Zdroj: | Journal of Virology. 84:8181-8192 |
| ISSN: | 1098-5514 0022-538X |
| Popis: | The p6 region of HIV-1 Gag contains two late (L) domains, PTAP and LYPX n L, that bind Tsg101 and Alix, respectively. Interactions with these two cellular proteins recruit members of the host's fission machinery (ESCRT) to facilitate HIV-1 release. Other retroviruses gain access to the host ESCRT components by utilizing a PPXY-type L domain that interacts with cellular Nedd4-like ubiquitin ligases. Despite the absence of a PPXY motif in HIV-1 Gag, interaction with the ubiquitin ligase Nedd4-2 was recently shown to stimulate HIV-1 release. We show here that another Nedd4-like ubiquitin ligase, Nedd4-1, corrected release defects resulting from the disruption of PTAP (PTAP − ), suggesting that HIV-1 Gag also recruits Nedd4-1 to facilitate virus release. Notably, Nedd4-1 remediation of HIV-1 PTAP − budding defects is independent of cellular Tsg101, implying that Nedd4-1's function in HIV-1 release does not involve ESCRT-I components and is therefore distinct from that of Nedd4-2. Consistent with this finding, deletion of the p6 region decreased Nedd4-1-Gag interaction, and disruption of the LYPX n L motif eliminated Nedd4-1-mediated restoration of HIV-1 PTAP − . This result indicated that both Nedd4-1 interaction with Gag and function in virus release occur through the Alix-binding LYPX n L motif. Mutations of basic residues located in the NC domain of Gag that are critical for Alix's facilitation of HIV-1 release, also disrupted release mediated by Nedd4-1, further confirming a Nedd4-1-Alix functional interdependence. In fact we found that Nedd4-1 binds Alix in both immunoprecipitation and yeast-two-hybrid assays. In addition, Nedd4-1 requires its catalytic activity to promote virus release. Remarkably, RNAi knockdown of cellular Nedd4-1 eliminated Alix ubiquitination in the cell and impeded its ability to function in HIV-1 release. Together our data support a model in which Alix recruits Nedd4-1 to facilitate HIV-1 release mediated through the LYPX n L/Alix budding pathway via a mechanism that involves Alix ubiquitination. |
| Databáze: | OpenAIRE |
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