90K, an interferon-stimulated gene product, reduces the infectivity of HIV-1

BACKGROUND: In response to viral infections, interferons induce the transcription of several hundred genes in mammalian cells. Specific antiviral functions, however, have only been attributed to a few of them. 90K/LGALS3BP has been reported to be an interferon-stimulated gene that is upregulated in individuals with cancer or HIV-1 infection. RESULTS: Here, we show that 90K expression dose-dependently decreased the particle infectivity of HIV-1 progeny. The lower infectivity of released particles correlated with reduced virion incorporation of mature envelope glycoproteins gp120 and gp41. Further, proteolytic processing of the gp160 precursor and surface expression of gp120 in the producer cell were impaired in the presence of 90K expression. In contrast, expression of Gag, Nef and Vpu, and virus release were not grossly affected by 90K expression. 90K-imposed restriction occurred in the absence of direct interaction of 90K with HIV-1 Env or entrapment of Env in the ER. The cell-associated, but not the secreted species of 90K, mediated the antiviral effect. A truncated version of human 90K, solely consisting of the two intermediate domains, displayed a similar antiviral activity as the full-length wildtype 90K, indicating that the N-terminal SRCR-like domain and the C-terminal domain are dispensable for 90K’s antiviral activity. The murine homolog of 90K, CypCAP (Cyclophilin C-associated protein), neither modulated particle infectivity of HIV-1 nor lowered the virion incorporation of mature gp120, suggesting a species-specific mode of action. 90K was expressed at basal levels in TZM-bl cells and in primary macrophages, and at low levels in CD4(+) T-cells and PBMCs. 90K’s susceptibility to IFN-mediated stimulation of expression was cell type-specific. siRNA-mediated knockdown of 90K in TZM-bl cells and primary macrophages enhanced the incorporation of Env glycoproteins into progeny virions, boosted the particle infectivity of released HIV-1, and accelerated HIV-1 spread. Conversely, treatment of HIV-1 infected macrophages with IFN-α induced 90K expression and lowered the particle infectivity of HIV-1. CONCLUSIONS: Thus, 90K constitutes a novel antiviral factor that reduces the particle infectivity of HIV-1, involving interference with the maturation and incorporation of HIV-1 Env molecules into virions

Characterization of 90K/LGALS3BP as antiviral factor

Cellular restriction factors inhibit HIV-1 at different stages of its replication cycle. One hallmark of these antiviral proteins is their interferon-inducible expression. The secreted glycoprotein 90K is also upregulated by interferon and induced in HIV-1 infected people. Herein, 90K was characterized as novel antiviral factor against HIV-1. Cellular 90K reduced the particle infectivity of HIV-1 progeny. Flow cytometry analysis revealed reduced levels of gp120 at the cell surface in presence of 90K expression leading to a lack in viral envelope incorporation. Additionally, a 90K induced processing defect of the gp160 precursor to mature gp120 and gp41 was observed, but seems to be uncoupled from the envelope incorporation defect. 90K did not act as a general inhibitor of furin cleavage nor did it trap Envelope in the endoplasmic reticulum. Envelope and 90K are both trafficked through the secretory pathway and therefore highly co-localize within cellular compartments, but direct interaction was not observed by co-immunoprecipitation. The antiviral activity of 90K was mapped to the two central protein binding domains BTB/POZ and IVR. 90K´s antiviral potential was conserved between 90K orthologs from six non-human primates. In contrast, 90K derived from rhesus macaques and the mouse homolog CypCAP failed to inhibit HIV-1, indicating 90K´s species specificity. Importantly, 90K is expressed in HIV-1 relevant CD4+ T cells, PBMCs and primary macrophages. In the latter, 90K expression is further stimulated by type I interferons. siRNA mediated knockdown of 90K in primary macrophage and PBMC cultures led to improved virus production. Silencing of 90K expression in primary macrophages boosted particle infectivity and enhanced Envelope incorporation into HIV-1 progeny virions. 90K also inhibited HIV-2 and SIVmac239, pointing toward a broader antiviral spectrum. Conclusively, 90K constitutes a relevant novel antiviral factor against HIV-1

Ion channel activity of HIV-1 Vpu is dispensable for counteraction of CD317.

While the C-terminal domain of HIV-1 Vpu is critical for CD4 degradation, the transmembrane domain (TM) mediates ion channel activity, enhances virus release and is essential for counteracting CD317/Bst-2/Tetherin. Here we analyzed whether the ion channel activity of Vpu is required to antagonize CD317-mediated restriction of virion release. We examined TM-mutants of three conserved residues: the S23A mutation, which was previously shown to abrogate ion channel function, did not affect Vpu mediated augmentation of virus release. In contrast, the A14N and A18N mutation did not affect ion channel activity of Vpu, but substantially reduced its ability to support virus release and to down-regulate CD317 from the cell surface. Altogether, our data suggest that not the ion channel activity of Vpu, but its ability to remove CD317 from the cell surface is required to augment HIV-1 release