Catch me if you can – the crosstalk of ZIKV and the restriction factor Tetherin

Zika virus (ZIKV) is a flavivirus that is mainly transmitted by Aedes mosquitos and normally causes mild symptoms. During the outbreak in the Americas in 2015, it was associated with more severe implications, like microcephaly in new-borns and the Gullain-Barré syndrome. The lack of specific vaccines and cures strengthen the need for a deeper understanding of the virus life cycle and virus-host interactions. The restriction factor tetherin (THN) is an interferon-inducible cellular protein with broad antiviral properties. It is known to inhibit the release of various enveloped viruses by tethering them to each other and to the cell membrane, thereby preventing their further spread. On the other hand, different viruses have developed various escape strategies against THN. Analysis of the crosstalk between ZIKV and THN revealed that in spite of a strong induction of THN mRNA expression in ZIKV-infected cells, this is not reflected by an elevated protein level of THN. Contrariwise, the THN protein level is decreased due to a reduced half-life. The increased degradation of THN in ZIKV infected cells involves the endo-lysosomal system, but does not depend on the early steps of autophagy. Enrichment of THN by depletion of the ESCRT-0 protein HRS diminishes ZIKV release and spread, which points out the capacity of THN to restrict ZIKV and explains the enhanced THN degradation in infected cells as an effective viral escape strategy. Importance Although tetherin expression is strongly induced by ZIKV infection there is a reduction in the amount of tetherin protein. This is due to an enhanced lysosomal degradation. However, if tetherin level is rescued release of ZIKV is impaired. This shows that tetherin is a restriction factor for ZIKV and the induction of an efficient degradation represents a viral escape strategy. To our knowledge this is the first study that describes and characterizes tetherin as an restriction factor for ZIKV life cycle.

The viral protein U (Vpu)-interacting host protein ATP6V0C down-regulates cell-surface expression of tetherin and thereby contributes to HIV-1 release

Host proteins with antiviral activity have evolved as first-line defenses to suppress viral replication. The HIV-1 accessory protein viral protein U (Vpu) enhances release of the virus from host cells by down-regulating the cell-surface expression of the host restriction factor tetherin. However, the exact mechanism of Vpu-mediated suppression of antiviral host responses is unclear. To further understand the role of host proteins in Vpu's function, here we carried out yeast two-hybrid screening and identified the V0 subunit C of vacuolar ATPase (ATP6V0C) as a Vpu-binding protein. To examine the role of ATP6V0C in Vpu-mediated tetherin degradation and HIV-1 release, we knocked down ATP6V0C expression in HeLa cells and observed that ATP6V0C depletion impairs Vpu-mediated tetherin degradation, resulting in defective HIV-1 release. We also observed that ATP6V0C overexpression stabilizes tetherin expression. This stabilization effect was specific to ATP6V0C, as overexpression of another subunit of the vacuolar ATPase, ATP6V0C″, had no effect on tetherin expression. ATP6V0C overexpression did not stabilize CD4, another target of Vpu-mediated degradation. Immunofluorescence localization experiments revealed that the ATP6V0C-stabilized tetherin is sequestered in a CD63– and lysosome-associated membrane protein 1 (LAMP1)–positive intracellular compartment. These results indicate that the Vpu-interacting protein ATP6V0C plays a role in down-regulating cell-surface expression of tetherin and thereby contributes to HIV-1 assembly and release.

The Vpu-interacting protein ATP6V0C regulates expression of tetherin and HIV-1 release

The HIV-1 accessory protein Vpu enhances virus release by down-regulating cell surface expression of the host restriction factor tetherin. To further understand the role of host proteins in Vpu function, we carried out yeast two-hybrid screening and identified the V0 subunit C of vacuolar ATPase (ATP6V0C) as a Vpu-binding protein. To examine the role of ATP6V0C in Vpu-mediated tetherin degradation and HIV-1 release, we knocked down ATP6V0C expression in HeLa cells and observed that ATP6V0C depletion impairs Vpu-mediated tetherin degradation, resulting in a defect in HIV-1 release. We also observed that overexpression of ATP6V0C stabilizes tetherin expression. This stabilization is specific to ATP6V0C, as overexpression of another subunit of the vacuolar ATPase, ATP6V0C”, had no effect on tetherin expression. ATP6V0C overexpression did not stabilize CD4, another target of Vpu-mediated degradation. Immunofluorescence localization studies showed that the ATP6V0C-stabilized tetherin is sequestered in a CD63- and LAMP1-positive intracellular compartment. These data demonstrate that the Vpu-interacting protein ATP6V0C plays a role in regulating tetherin expression and HIV-1 assembly and release.

Tetherin Inhibits Nipah Virus but Not Ebola Virus Replication in Fruit Bat Cells

ABSTRACT Ebola virus (EBOV) and Nipah virus (NiV) infection of humans can cause fatal disease and constitutes a public health threat. In contrast, EBOV and NiV infection of fruit bats, the putative (EBOV) or proven (NiV) natural reservoir, is not associated with disease, and it is currently unknown how these animals control the virus. The human interferon (IFN)-stimulated antiviral effector protein tetherin (CD317, BST-2) blocks release of EBOV- and NiV-like particles from cells and is counteracted by the EBOV glycoprotein (GP). In contrast, it is unknown whether fruit bat tetherin restricts virus infection and is susceptible to GP-driven antagonism. Here, we report the sequence of fruit bat tetherin and show that its expression is IFN stimulated and associated with strong antiviral activity. Moreover, we demonstrate that EBOV-GP antagonizes tetherin orthologues of diverse species but fails to efficiently counteract fruit bat tetherin in virus-like particle (VLP) release assays. However, unexpectedly, tetherin was dispensable for robust IFN-mediated inhibition of EBOV spread in fruit bat cells. Thus, the VLP-based model systems mimicking tetherin-mediated inhibition of EBOV release and its counteraction by GP seem not to adequately reflect all aspects of EBOV release from IFN-stimulated fruit bat cells, potentially due to differences in tetherin expression levels that could not be resolved by the present study. In contrast, tetherin expression was essential for IFN-dependent inhibition of NiV infection, demonstrating that IFN-induced fruit bat tetherin exerts antiviral activity and may critically contribute to control of NiV and potentially other highly virulent viruses in infected animals. IMPORTANCE Ebola virus and Nipah virus (EBOV and NiV) can cause fatal disease in humans. In contrast, infected fruit bats do not develop symptoms but can transmit the virus to humans. Why fruit bats but not humans control infection is largely unknown. Tetherin is an antiviral host cell protein and is counteracted by the EBOV glycoprotein in human cells. Here, employing model systems, we show that tetherin of fruit bats displays higher antiviral activity than human tetherin and is largely resistant against counteraction by the Ebola virus glycoprotein. Moreover, we demonstrate that induction of tetherin expression is critical for interferon-mediated inhibition of NiV but, for at present unknown reasons, not EBOV spread in fruit bat cells. Collectively, our findings identify tetherin as an antiviral effector of innate immune responses in fruit bats, which might allow these animals to control infection with NiV and potentially other viruses that cause severe disease in humans.

Tetherin Sensitivity of Influenza A Viruses Is Strain Specific: Role of Hemagglutinin and Neuraminidase

ABSTRACTThe expression of the antiviral host cell factor tetherin is induced by interferon and can inhibit the release of enveloped viruses from infected cells. The Vpu protein of HIV-1 antagonizes the antiviral activity of tetherin, and tetherin antagonists with Vpu-like activity have been identified in other viruses. In contrast, it is incompletely understood whether tetherin inhibits influenza A virus (FLUAV) release and whether FLUAV encodes tetherin antagonists. Here, we show that release of several laboratory-adapted FLUAV strains and a seasonal FLUAV strain is inhibited by tetherin, while pandemic FLUAV A/Hamburg/4/2009 is resistant. Studies with a virus-like particle system and analysis of reassortant viruses provided evidence that the viral hemagglutinin (HA) is an important determinant of tetherin antagonism but requires the presence of its cognate neuraminidase (NA) to inhibit tetherin. Finally, tetherin antagonism by FLUAV was dependent on the virion context, since retrovirus release from tetherin-positive cells was not rescued, and correlated with an HA- and NA-dependent reduction in tetherin expression. In sum, our study identifies HA and NA proteins of certain pandemic FLUAV as tetherin antagonists, which has important implications for understanding FLUAV pathogenesis.IMPORTANCEInfluenza A virus (FLUAV) infection is responsible for substantial global morbidity and mortality, and understanding how the virus evades the immune defenses of the host may uncover novel targets for antiviral intervention. Tetherin is an antiviral effector molecule of the innate immune system which can contribute to control of viral invasion. However, it has been unclear whether FLUAV is inhibited by tetherin and whether these viruses encode tetherin-antagonizing proteins. Our observation that several pandemic FLUAV strains can counteract tetherin via their HA and NA proteins identifies these proteins as novel tetherin antagonists and indicates that HA/NA-dependent inactivation of innate defenses may contribute to the efficient spread of pandemic FLUAV.