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MxB binds to the HIV-1 core and prevents the uncoating process of HIV-1.

ABSTRACT: The IFN-?-inducible restriction factor MxB blocks HIV-1 infection after reverse transcription but prior to integration. Genetic evidence suggested that capsid is the viral determinant for restriction by MxB. This work explores the ability of MxB to bind to the HIV-1 core, and the role of capsid-binding in restriction.We showed that MxB binds to the HIV-1 core and that this interaction leads to inhibition of the uncoating process of HIV-1. These results identify MxB as an endogenously expressed protein with the ability to inhibit HIV-1 uncoating. In addition, we found that a benzimidazole-based compound known to have a binding pocket on the surface of the HIV-1 capsid prevents the binding of MxB to capsid. The use of this small-molecule identified the MxB binding region on the surface of the HIV-1 core. Domain mapping experiments revealed the following requirements for restriction: 1) MxB binding to the HIV-1 capsid, which requires the 20 N-terminal amino acids, and 2) oligomerization of MxB, which is mediated by the C-terminal domain provides the avidity for the interaction of MxB with the HIV-1 core.Overall our work establishes that MxB binds to the HIV-1 core and inhibits the uncoating process of HIV-1. Moreover, we demonstrated that HIV-1 restriction by MxB requires capsid binding and oligomerization.

SUBMITTER: Fricke T

PROVIDER: S-EPMC4145229 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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MxB binds to the HIV-1 core and prevents the uncoating process of HIV-1.

Fricke Thomas T White Tommy E TE Schulte Bianca B de Souza Aranha Vieira Daniel A DA Dharan Adarsh A Campbell Edward M EM Brandariz-Nuñez Alberto A Diaz-Griffero Felipe F

Retrovirology 20140814

<h4>Background</h4>The IFN-α-inducible restriction factor MxB blocks HIV-1 infection after reverse transcription but prior to integration. Genetic evidence suggested that capsid is the viral determinant for restriction by MxB. This work explores the ability of MxB to bind to the HIV-1 core, and the role of capsid-binding in restriction.<h4>Results</h4>We showed that MxB binds to the HIV-1 core and that this interaction leads to inhibition of the uncoating process of HIV-1. These results identify ...[more]

PMID: 25123063

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Project description:BackgroundInterferon-induced cellular proteins play important roles in the host response against viral infection. The Mx family of dynamin-like GTPases, which include MxA and MxB, target a wide variety of viruses. Despite considerable evidence demonstrating the breadth of antiviral activity of MxA, human MxB was only recently discovered to specifically inhibit lentiviruses. Here we assess both host and viral determinants that underlie MxB restriction of HIV-1 infection.ResultsHeterologous expression of MxB in human osteosarcoma cells potently inhibited HIV-1 infection (~12-fold), yet had little to no effect on divergent retroviruses. The anti-HIV effect manifested as a partial block in the formation of 2-long terminal repeat circle DNA and hence nuclear import, and we accordingly found evidence for an additional post-nuclear entry block. A large number of previously characterized capsid mutations, as well as mutations that abrogated integrase activity, counteracted MxB restriction. MxB expression suppressed integration into gene-enriched regions of chromosomes, similar to affects observed previously when cells were depleted for nuclear transport factors such as transportin 3. MxB activity did not require predicted GTPase active site residues or a series of unstructured loops within the stalk domain that confer functional oligomerization to related dynamin family proteins. In contrast, we observed an N-terminal stretch of residues in MxB to harbor key determinants. Protein localization conferred by a nuclear localization signal (NLS) within the N-terminal 25 residues, which was critical, was fully rescuable by a heterologous NLS. Consistent with this observation, a heterologous nuclear export sequence (NES) abolished full-length MxB activity. We additionally mapped sub-regions within amino acids 26-90 that contribute to MxB activity, finding sequences present within residues 27-50 particularly important.ConclusionsMxB inhibits HIV-1 by interfering with minimally two steps of infection, nuclear entry and post-nuclear trafficking and/or integration, without destabilizing the inherent catalytic activity of viral preintegration complexes. Putative MxB GTPase active site residues and stalk domain Loop 4 -- both previously shown to be necessary for MxA function -- were dispensable for MxB antiviral activity. Instead, we highlight subcellular localization and a yet-determined function(s) present in the unique MxB N-terminal region to be required for HIV-1 restriction.

Project description:Type I interferon inhibits viruses through inducing the expression of antiviral proteins, including the myxovirus resistance (Mx) proteins. Compared to the human MxA protein, which inhibits a wide range of viruses, the MxB protein has been reported to specifically inhibit primate lentiviruses, including HIV-1, and herpesviruses. Further, the role of endogenous MxB in alpha interferon-mediated inhibition of HIV-1 infection was questioned by a recent study showing that MxB knockout did not increase the level of infection by HIV-1 which carried the G protein of vesicular stomatitis virus (VSV), allowing infection of CD4-negative HT1080 cells. In order to further examine the anti-HIV-1 activity of endogenous MxB, we have used CRISPR/Cas9 to deplete MxB in different cell lines and observed a substantial restoration of HIV-1 infection in the presence of alpha interferon treatment. However, this rescue effect of MxB knockout became much less pronounced when infection was performed with HIV-1 carrying the VSV G protein. Interestingly, a CRISPR/Cas9 knockout screen of alpha interferon-stimulated genes in U87-MG cells revealed that the genes for interferon-induced transmembrane protein 2 (IFITM2) and IFITM3 inhibited VSV G-pseudotyped HIV-1 much more strongly than the rest of the genes tested, including the gene for MxB. Therefore, our results demonstrate the importance of MxB in alpha interferon-mediated inhibition of HIV-1 infection, which, however, can be underestimated if infection is performed with VSV G protein-pseudotyped HIV-1, due to the high sensitivity of VSV G-mediated infection to inhibition by IFITM proteins.IMPORTANCE The results of this study reconcile the controversial reports regarding the anti-HIV-1 function of alpha interferon-induced MxB protein. In addition to the different cell types that may have contributed to the different observations, our data also suggest that VSV G protein-pseudotyped HIV-1 is much less inhibited by alpha interferon-induced MxB than HIV-1 itself is. Our results clearly demonstrate an important contribution of MxB to alpha interferon-mediated inhibition of HIV-1 in CD4+ T cells, which calls for using HIV-1 target cells and wild-type virus to test the relevance of the anti-HIV-1 activity of endogenous MxB and other restriction factors.

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MxB binds to the HIV-1 core and prevents the uncoating process of HIV-1.

Publications

MxB binds to the HIV-1 core and prevents the uncoating process of HIV-1.

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