Tetherin, also known as bone marrow stromal antigen 2 (BST-2), inhibits the release of a wide range of enveloped viruses, including human immunodeficiency virus, type 1 (HIV-1) by directly tethering nascent virions to the surface of infected cells. The HIV-1 accessary protein Vpu counteracts tetherin restriction via sequestration, down-regulation, and/or displacement mechanisms to remove tetherin from sites of virus budding. However, the exact mechanism of Vpu-mediated antagonism of tetherin restriction remains to be fully understood. Here we report a novel role for the actin cross-linking regulator filamin A (FLNa) in Vpu anti-tetherin activities. We demonstrate that FLNa associates with tetherin and that FLNa modulates tetherin turnover. FLNa deficiency was found to enhance cell surface and steady-state levels of tetherin expression. In contrast, we observed that overexpression of FLNa reduced tetherin expression levels both on the plasma membrane and in intracellular compartments. Although FLNb shows high amino acid sequence similarity with FLNa, we reveal that only FLNa, but not FLNb, plays an essential role in tetherin turnover. We further showed that FLNa deficiency inhibited Vpu-mediated enhancement of virus release through interfering with the activity of Vpu to down-regulate cellular tetherin. Taken together, our studies suggest that Vpu hijacks the FLNa function in the modulation of tetherin to neutralize the antiviral factor tetherin. These findings may provide novel strategies for the treatment of HIV-1 infection.

Schistosomes are parasitic helminths of the genus Schistosoma that cause the vascular disease Schistosomiasis. Previous studies show that the major antibody responses to schistosome infections are directed against glycans, making these molecules potential targets for the development of vaccines. We are developing monoclonal antibodies to these antigens from spleens of infected mice. Twelve mice were infected with 25–30 cercariae and the antibody responses were monitored over the period of 14 weeks. IgM responses start at week 4 post infection, peak at week 8, and remained high during the course of the study. IgG responses start at week 6, peak at week 11, and remained high during the course of the study. Interestingly, the IgG responses to antigens were sensitive to periodate oxidation only up to week 10 post infection and became progressively resistant to periodate oxidation thereafter. This was confirmed by analyzing infected mice sera against three defined Schistosome glycan antigens. Taken together, IgM responses to glycan antigens persist throughout the course of infection while IgG responses switch from anti‐glycan responses to non‐glycan responses in the later stages of infection. Thus, there is a narrow window of time for harvesting spleens from infected mice for generation of IgG monoclonal antibodies to glycan antigens. Supported by grants T34GM008411 and 5 S06 GM078091‐04.

The interferon‐induced tetherin is a restriction factor that inhibits the release of HIV‐1 and other enveloped viruses from infected cells. The unique topology of tetherin, featured with two different membrane anchors at the opposite ends, enables it to capture the mature virions at the cell surface through a direct tethering mechanism. HIV‐1 has evolved the strategy to overcome tetherin restriction by encoding an accessory protein, Vpu. HIV‐1 Vpu neutralizes tetherin through multiple mechanisms. Recently, we identified that the host cytoskeletal component, filamin A (FLNa) acts as a cofactor in HIV‐1 assembly and release. By cell‐based assays, we demonstrated that FLNa was capable of binding tetherin. FLNa depletion increased steady‐state and cell surface levels of tethrin expression, whereas FLNa overexpression reduced tetherin expression. Furthermore, FLNa depletion or overexpression redistributed tetherin subcellular localization. These findings suggest that FLNa is involved in tetherin turnover by mediating tetherin trafficking pathways. Transmission electron microscopy studies illustrated that FLNa depletion induces a defect in HIV‐1 particle release, characterized with virions tethered to each other as well as to cell surface. Finally, virus release experiments revealed that FLNa depletion abolished the activity of Vpu to enhance particle release. Taken together, our results suggest that HIV‐1 Vpu takes advantage of the FLNa‐dependent tetherin trafficking pathways to neutralize tetherin and promote virus release. These studies shed new insights into the mechanisms of Vpu‐mediated counteraction of tetherin restriction. Grant Funding Source: AI089330, GM059994

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