A Carboxy-Terminally Truncated Human CPSF6 Lacking Residues Encoded by Exon 6 Inhibits HIV-1 cDNA Synthesis and Promotes Capsid Disassembly

In a previous screen of putative interferon-stimulated genes, SUN2 was shown to inhibit HIV-1 infection in an uncharacterized manner. SUN2 is an inner nuclear membrane protein belonging to the linker of nucleoskeleton and cytoskeleton complex. We have analyzed here the role of SUN2 in HIV infection. We report that in contrast to what was initially thought, SUN2 is not induced by type I interferon, and that SUN2 silencing does not modulate HIV infection. However, SUN2 overexpression in cell lines and in primary monocyte-derived dendritic cells inhibits the replication of HIV but not murine leukemia virus or chikungunya virus. We identified HIV-1 and HIV-2 strains that are unaffected by SUN2, suggesting that the effect is specific to particular viral components or cofactors. Intriguingly, SUN2 overexpression induces a multilobular flower-like nuclear shape that does not impact cell viability and is similar to that of cells isolated from patients with HTLV-I-associated adult T-cell leukemia or with progeria. Nuclear shape changes and HIV inhibition both mapped to the nucleoplasmic domain of SUN2 that interacts with the nuclear lamina. This block to HIV replication occurs between reverse transcription and nuclear entry, and passaging experiments selected for a single-amino-acid change in capsid (CA) that leads to resistance to overexpressed SUN2. Furthermore, using chemical inhibition or silencing of cyclophilin A (CypA), as well as CA mutant viruses, we implicated CypA in the SUN2-imposed block to HIV infection. Our results demonstrate that SUN2 overexpression perturbs both nuclear shape and early events of HIV infection. IMPORTANCECells encode proteins that interfere with viral replication, a number of which have been identified in overexpression screens. SUN2 is a nuclear membrane protein that was shown to inhibit HIV infection in such a screen, but how it blocked HIV infection was not known. We show that SUN2 overexpression blocks the infection of certain strains of HIV before nuclear entry. Mutation of the viral capsid protein yielded SUN2-resistant HIV. Additionally, the inhibition of HIV infection by SUN2 involves cyclophilin A, a protein that binds the HIV capsid and directs subsequent steps of infection. We also found that SUN2 overexpression substantially changes the shape of the cell's nucleus, resulting in many flower-like nuclei. Both HIV inhibition and deformation of nuclear shape required the domain of SUN2 that interacts with the nuclear lamina. Our results demonstrate that SUN2 interferes with HIV infection and highlight novel links between nuclear shape and viral infection.

Section: Discussionmentioning

confidence: 80%

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confidence: 99%

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confidence: 99%

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SUN2 Overexpression Deforms Nuclear Shape and Inhibits HIV

Donahue

Amraoui

Nunzio

et al. 2016

“…The cellular restriction factors TRIM5␣ and TRIMCyp have been shown to inhibit infection of multiple retroviruses by binding the capsid and inducing premature uncoating (7)(8)(9). Conversely, when CPSF6, which is normally located in the nucleus, is aberrantly localized in the cytoplasm, the uncoating of some retroviral capsids is slowed, resulting in decreased infection (10,11). Some of the inhibitory effects on HIV-1 infection associated with knockdown of TNP03, a nuclear import factor, may be due to secondary effects on CPSF6 localization (12)(13)(14).…”

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confidence: 99%

Contribution of PDZD8 to Stabilization of the Human Immunodeficiency Virus Type 1 Capsid

Guth

Sodroski

2014

“…The MLV CA interacts with the cellular Friend virus susceptibility factor 1 (Fv1) (30). Binding of these proteins to capsids in the cytosol affects several postentry events (22,23,(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40), suggesting that the FL serves both structural and regulatory roles in the establishment of a new infection. As yet, no CA-binding host proteins have been identified in the alpharetroviruses.…”

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confidence: 99%

Contributions of Charged Residues in Structurally Dynamic Capsid Surface Loops to Rous Sarcoma Virus Assembly

Heyrana

Goh

Perilla

et al. 2016

Extensive studies of orthoretroviral capsids have shown that many regions of the CA protein play unique roles at different points in the virus life cycle. The N-terminal domain (NTD) flexible-loop (FL) region is one such example: exposed on the outer capsid surface, it has been implicated in Gag-mediated particle assembly, capsid maturation, and early replication events. We have now defined the contributions of charged residues in the FL region of the Rous sarcoma virus (RSV) CA to particle assembly. Effects of mutations on assembly were assessed in vivo and in vitro and analyzed in light of new RSV Gag lattice models. Virus replication was strongly dependent on the preservation of charge at a few critical positions in Gag-Gag interfaces. In particular, a cluster of charges at the beginning of FL contributes to an extensive electrostatic network that is important for robust Gag assembly and subsequent capsid maturation. Second-site suppressor analysis suggests that one of these charged residues, D87, has distal influence on interhexamer interactions involving helix ␣7. Overall, the tolerance of FL to most mutations is consistent with current models of Gag lattice structures. However, the results support the interpretation that virus evolution has achieved a charge distribution across the capsid surface that (i) permits the packing of NTD domains in the outer layer of the Gag shell, (ii) directs the maturational rearrangements of the NTDs that yield a functional core structure, and (iii) supports capsid function during the early stages of virus infection. IMPORTANCEThe production of infectious retrovirus particles is a complex process, a choreography of protein and nucleic acid that occurs in two distinct stages: formation and release from the cell of an immature particle followed by an extracellular maturation phase during which the virion proteins and nucleic acids undergo major rearrangements that activate the infectious potential of the virion. This study examines the contributions of charged amino acids on the surface of the Rous sarcoma virus capsid protein in the assembly of appropriately formed immature particles and the maturational transitions that create a functional virion. The results provide important biological evidence in support of recent structural models of the RSV immature virions and further suggest that immature particle assembly and virion maturation are controlled by an extensive network of electrostatic interactions and long-range communication across the capsid surface. Infectious retrovirus particles are the end products of a complex multistage assembly process. In the early phase, the Gag polyprotein binds virion genomic RNA and is targeted to the plasma membrane, where it assembles a near-spherical immature protein lattice. As the virion leaves the cell by a budding process, the virally encoded protease PR is activated, initiating maturation. Cleavage of the Gag polyprotein into the structural proteins MA (matrix), CA (capsid), and NC (nucleocapsid) results in a large-scale re...