Bone Marrow Stromal Antigen Research Articles

Bst2/Tetherin Is Induced in Neurons by Type I Interferon and Viral Infection but Is Dispensable for Protection against Neurotropic Viral Challenge.

In permissive mouse central nervous system (CNS) neurons, measles virus (MV) spreads in the absence of hallmark viral budding or neuronal death, with transmission occurring efficiently and exclusively via the synapse. MV infection also initiates a robust type I interferon (IFN) response, resulting in the synthesis of a large number of genes, including bone marrow stromal antigen 2 (Bst2)/tetherin/CD317. Bst2 restricts the release of some enveloped viruses, but to date, its role in viral infection of neurons has not been assessed. Consequently, we investigated how Bst2 was induced and what role it played in MV neuronal infection. The magnitude of induction of neuronal Bst2 RNA and protein following IFN exposure and viral infection was notably higher than in similarly treated mouse embryo fibroblasts (MEFs). Bst2 synthesis was both IFN and Stat1 dependent. Although Bst2 prevented MV release from nonneuronal cells, its deletion had no effect on viral pathogenesis in MV-challenged mice. Our findings underscore how cell-type-specific differences impact viral infection and pathogenesis. Viral infections of the central nervous system can lead to debilitating disease and death. Moreover, it is becoming increasingly clear that nonrenewable cells, including most central nervous system neurons, combat neurotropic viral infections in fundamentally different ways than other rapidly dividing and renewable cell populations. Here we identify type I interferon signaling as a key inducer of a known antiviral protein (Bst2) in neurons. Unexpectedly, the gene is dispensable for clearance of neurotropic viral infection despite its well-defined contribution to limiting the spread of enveloped viruses in proliferating cells. A deeper appreciation of the importance of cell type heterogeneity in antiviral immunity will aid in the identification of unique therapeutic targets for life-threatening viral infections.

Abstract 2487: The antitumor efficacy of anti-BST2 antibody is significantly increased in combination with CpG oligodeoxynucleotide in endometrial cancer

Abstract Background: CpG oligodeoxynucleotides (ODN) has been shown to exhibit a potent immunostimulatory activity. Therefore, CpG ODN has been considered to be a promising material for enhancing the therapeutic effect of anticancer monoclonal antibody with antibody-dependent cell-mediated cytotoxicity (ADCC) activity. We have previously identified bone marrow stromal antigen 2 (BST2) as a therapeutic target of endometrial cancer cells and also demonstrated that an anti-BST2 antibody showed a potent anti-tumor effect by ADCC and complement-dependent cytotoxicity (CDC) activities against BST2 positive cells. The aim of this study was to evaluate the synergistic anti-tumor effect by combination of anti-BST2 antibody and CpG ODN, and to clarify the mechanisms of its anti-tumor effect, using xenograft model of BST2 positive endometrial cancer cells. Material and Method: We implanted 7-8 week-old female SCID mice with 5×106 HEC-88 nu endometrial cancer cells subcutaneously. Mice with tumor were treated by intra-peritoneal injection of an anti-BST2 mAb alone (12.5 and 200 μg/mouse, twice a week for four weeks), intra-tumoral injection of CpG ODN alone (10 μg/mouse, twice a week for four weeks) or combination therapy (12.5 μg/mouse of anti-BST2 mAb and 10 μg/mouse of CpG ODN). Anti-tumor effect was evaluated by monitoring the changes of tumor volume. Finally, population of NK cells and macrophages infiltrated into the tumor were analyzed by the FACS analysis, how CpG ODN induces immunopotentiative action. Results: Intraperitoneal treatment by anti-BST2 antibody showed a significant anti-tumor effect, dose dependently. The combination therapy of anti-BST2 antibody and CpG ODN was more effective than the single-agent therapy (p = 0.0002). In addition, the combination therapy also resulted high population of NK cells and macrophages in tumor significantly (p = 0.00018). Conclusion: We showed that CpG ODN increases the efficacy of anti-tumor effect of anti-BST2 mAb induced ADCC activity via potentiating NK cells and macrophages mediated cancer cell injury. Combination of GpG ODN would be a promising approach for enhancing the activity of ADCC based monoclonal antibodies, which targeting anti-tumor antigen. Citation Format: Yusuke Takahashi, Satoshi Serada, Kosuke Hiramatsu, Kouji Kobiyama, Minoru Fujimoto, Ken Ishii, Tetsuji Naka. The antitumor efficacy of anti-BST2 antibody is significantly increased in combination with CpG oligodeoxynucleotide in endometrial cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2487. doi:10.1158/1538-7445.AM2015-2487

Abstract 4382: Anti-human LSR monoclonal antibody inhibits tumor growth of ovarian cancer directly

Abstract Ovarian cancer is the most lethal gynecologic malignancy; thus developing new treatment options is urgently required. Molecular targeted therapies for cancers, which are generally more tolerable than widely used cytotoxic agents, have shown highly specific inhibition of target molecules. We previously identified bone marrow stromal antigen 2 (BST2) as an endometrial cancer antigen using iTRAQ-based quantitative proteomic technology focused on cell surface membrane proteins, and also demonstrated the usefulness of an anti-BST2 monoclonal antibody (mAb) for endometrial cancer. In this study, we aimed to identify a new ovarian cancer antigen. We also aimed to develop a novel monoclonal antibody (mAb) and evaluate its preclinical efficacy against ovarian cancer. To identify a new ovarian cancer antigen, cell surface membrane proteins of normal ovarian epithelial and ovarian cancer cell lines were analyzed by iTRAQ-based proteomic technology. As the new therapeutic target for ovarian cancer, we identified lipolysis-stimulated lipoprotein receptor (LSR) which had one of the largest significant differences in protein level between normal ovarian epithelial and ovarian cancer cell lines. Immunohistochemical analysis showed that the overall survival of ovarian serous carcinoma patients with high LSR expression was significantly shorter than those with low LSR (p = 0.0293). We newly developed anti-LSR mAb and investigated its preclinical efficacy. Anti-LSR mAb showed significant in vivo inhibition of tumor growth against a xenograft model of hLSR-positive ovarian cancer in an antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) independent manner (p = 0.0001). And anti-LSR mAb also induced G0/G1 cell cycle arrest by regulation of MEK and p44/42 MAPK activities and expression levels of cell cycle related proteins in vitro. Furthermore, anti-hLSR mAb, which crossreacts with mouse LSR, had little detectable toxicity in mice. In summary, high expression of LSR in ovarian cancer was the poor prognostic factor. Our newly developed anti-LSR mAb showed significant tumor growth inhibition in ADCC and CDC independent manner in vivo. Anti-human LSR mAb also inhibited LSR function and showed direct tumor growth inhibition inducing G0/G1 cell cycle arrest in vitro. Our preclinical data demonstrated that targeting LSR by mAb is a promising therapy for patients with LSR-positive ovarian cancer. Citation Format: Kosuke Hiramatsu, Satoshi Serada, Takayuki Enomoto, Satoshi Nakagawa, Akiko Morimoto, Minoru Fujimoto, Takuhei Yokoyama, Yusuke Takahashi, Yutaka Ueda, Kiyoshi Yoshino, Eiichi Morii, Tadashi Kimura, Tetsuji Naka. Anti-human LSR monoclonal antibody inhibits tumor growth of ovarian cancer directly. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4382. doi:10.1158/1538-7445.AM2015-4382

Bone marrow stromal antigen 2 (BST-2) DNA is demethylated in breast tumors and breast cancer cells.

BackgroundBone marrow stromal antigen 2 (BST-2) is a known anti-viral gene that has been recently identified to be overexpressed in many cancers, including breast cancer. BST-2 is critical for the invasiveness of breast cancer cells and the formation of metastasis in vivo. Although the regulation of BST-2 in immune cells is unraveling, it is unknown how BST-2 expression is regulated in breast cancer. We hypothesized that meta-analyses of BST-2 gene expression and BST-2 DNA methylation profiles would illuminate mechanisms regulating elevated BST-2 expression in breast tumor tissues and cells.Materials and MethodsWe performed comprehensive meta-analyses of BST-2 gene expression and BST-2 DNA methylation in The Cancer Genome Atlas (TCGA) and various Gene Expression Omnibus (GEO) datasets. BST-2 expression levels and BST-2 DNA methylation status at specific CpG sites on the BST-2 gene were compared for various breast tumor molecular subtypes and breast cancer cell lines.ResultsWe show that BST-2 gene expression is inversely associated with the methylation status at specific CpG sites in primary breast cancer specimens and breast cancer cell lines. BST-2 demethylation is significantly more prevalent in primary tumors and cancer cells than in normal breast tissues or normal mammary epithelial cells. Demethylation of the BST-2 gene significantly correlates with its mRNA expression. These studies provide the initial evidence that significant differences exist in BST-2 DNA methylation patterns between breast tumors and normal breast tissues, and that BST-2 expression patterns in tumors and cancer cells correlate with hypomethylated BST-2 DNA.ConclusionOur study suggests that the DNA methylation pattern and expression of BST-2 may play a role in disease pathogenesis and could serve as a biomarker for the diagnosis of breast cancer.

Membrane Anchoring by a C-terminal Tryptophan Enables HIV-1 Vpu to Displace Bone Marrow Stromal Antigen 2 (BST2) from Sites of Viral Assembly

The restriction factor BST2 (tetherin) prevents the release of enveloped viruses from the host cell and is counteracted by HIV-1 Vpu. Vpu and BST2 interact directly via their transmembrane domains. This interaction enables Vpu to induce the surface down-regulation and the degradation of BST2, but neither of these activities fully accounts for the ability of Vpu to enhance virion release. During a study of naturally occurring Vpu proteins, we found that a tryptophan residue near the Vpu C terminus is particularly important for enhancing virion release. Vpu proteins with a W76G polymorphism degraded and down-regulated BST2 from the cell surface, yet they inefficiently stimulated virion release. Here we explore the mechanism of this anomaly. We find that Trp-76 is critical for the ability of Vpu to displace BST2 from sites of viral assembly in the plane of the plasma membrane. This effect does not appear to involve a general reorganization of the membrane microdomains associated with virion assembly, but rather is a specific effect of Vpu on BST2. Using NMR spectroscopy, we find that the cytoplasmic domain of Vpu and Trp-76 specifically interact with lipids. Moreover, paramagnetic relaxation enhancement studies show that Trp-76 inserts into the lipid. These data are consistent with a model whereby Trp-76 anchors the C terminus of the cytoplasmic tail of Vpu to the plasma membrane, enabling the movement of Vpu-bound BST2 away from viral assembly sites.

Bone Marrow Stromal Antigen 2 Is a Novel Plasma Biomarker and Prognosticator for Colorectal Carcinoma: A Secretome-Based Verification Study.

Background. The cancer cell secretome has been recognized as a valuable reservoir for identifying novel serum/plasma biomarkers for different cancers, including colorectal cancer (CRC). This study aimed to verify four CRC cell-secreted proteins (tumor-associated calcium signal transducer 2/trophoblast cell surface antigen 2 (TACSTD2/TROP2), tetraspanin-6 (TSPAN6), bone marrow stromal antigen 2 (BST2), and tumor necrosis factor receptor superfamily member 16 (NGFR)) as potential plasma CRC biomarkers. Methods. The study population comprises 152 CRC patients and 152 controls. Target protein levels in plasma and tissue samples were assessed by ELISA and immunohistochemistry, respectively. Results. Among the four candidate proteins examined by ELISA in a small sample set, only BST2 showed significantly elevated plasma levels in CRC patients versus controls. Immunohistochemical analysis revealed the overexpression of BST2 in CRC tissues, and higher BST2 expression levels correlated with poorer 5-year survival (46.47% versus 65.57%; p = 0.044). Further verification confirmed the elevated plasma BST2 levels in CRC patients (2.35 ± 0.13 ng/mL) versus controls (1.04 ± 0.03 ng/mL) (p < 0.01), with an area under the ROC curve (AUC) being 0.858 comparable to that of CEA (0.867). Conclusion. BST2, a membrane protein selectively detected in CRC cell secretome, may be a novel plasma biomarker and prognosticator for CRC.

Bone marrow stromal antigen 2 expressed in cancer cells promotes mammary tumor growth and metastasis.

IntroductionSeveral innate immunity genes are overexpressed in human cancers and their roles remain controversial. Bone marrow stromal antigen 2 (BST-2) is one such gene whose role in cancer is not clear. BST-2 is a unique innate immunity gene with both antiviral and pro-tumor functions and therefore can serve as a paradigm for understanding the roles of other innate immunity genes in cancers.MethodsMeta-analysis of tumors from breast cancer patients obtained from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) datasets were evaluated for levels of BST-2 expression and for tumor aggressiveness. In vivo, we examined the effect of knockdown of BST-2 in two different murine carcinoma cells on tumor growth, metastasis, and survival. In vitro, we assessed the effect of carcinoma cell BST-2 knockdown and/or overexpression on adhesion, anchorage-independent growth, migration, and invasion.ResultsBST-2 in breast tumors and mammary cancer cells is a strong predictor of tumor size, tumor aggressiveness, and host survival. In humans, BST-2 mRNA is elevated in metastatic and invasive breast tumors. In mice, orthotopic implantation of mammary tumor cells lacking BST-2 increased tumor latency, decreased primary tumor growth, reduced metastases to distal organs, and prolonged host survival. Furthermore, we found that the cellular basis for the role of BST-2 in promoting tumorigenesis include BST-2-directed enhancement in cancer cell adhesion, anchorage-independency, migration, and invasion.ConclusionsBST-2 contributes to the emergence of neoplasia and malignant progression of breast cancer. Thus, BST-2 may (1) serve as a biomarker for aggressive breast cancers, and (2) be a novel target for breast cancer therapeutics.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-014-0493-8) contains supplementary material, which is available to authorized users.

Abstract 2572: CpG oligodeoxynucleotide enhances the efficacy of anticancer monoclonal antibody in an in vivo xenograft model using human endometrial cancer cell

Abstract Background: CpG oligodeoxynucleotides (ODN) has been shown to exhibit a potent immunostimulatory activity. Therefore, CpG ODN has been considered to be a promising material for enhancing the therapeutic effect of anticancer monoclonal antibody. We have previously identified bone marrow stromal antigen 2 (BST2) as a therapeutic target of endometrial cancer cells and also demonstrated that an anti-BST2 antibody showed a potent anti-tumor effect by antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against BST2 positive cells. The aim of this study was to evaluate the synergistic anti-tumor effect by combination of anti-BST-2 antibody and CpG ODN, and to clarify the mechanisms of its anti-tumor effect, using xenograft model of anti-BST2 positive endometrial cancer cells. Material and Method: We implanted 7-8 week-old female SCID mice with 5×106 HEC-88 nu endometrial cancer cells subcutaneously. Mice with tumor were treated by intra-peritoneal injection of an anti-BST2 antibody alone (0 - 200 μg/mouse, twice a week for four weeks), intra-tumoral injection of CpG ODN alone (0 - 30 μg/mouse, twice a week for four weeks) or combination therapy (anti-BST2 antibody and CpG ODN). Anti-tumor effect was evaluated by monitoring the changes of tumor volume and also pathological analysis. In addition, activation of NK cells was evaluated. Results: Intraperitoneal treatment of anti-BST2 antibody showed a significant anti-tumor effect, dose dependently. Similarly, intratumoral injection of CpG ODN showed a significant anti-tumor effect, in a dose dependent manner. Interestingly, the combination therapy was more effective than the single-agent therapy. In addition, the combination therapy also resulted in a high activity of NK cells. Conclusion: We showed that CpG ODN increases the efficacy of anti-tumor effect of anti-BST2 antibody induced ADCC activity via potentiating NK cells mediated cancer cell injury. Combination of CpG ODN and anti-tumor monoclonal antibody is useful cancer therapy. Citation Format: Kosuke Hiramatsu, Satoshi Serada, Kouji Kobiyama, Akiko Morimoto, Toshihiro Kimura, Kiyoshi Yoshino, Minoru Fujimoto, Ken J. Ishii, Masami Fujita, Tetsuji Naka. CpG oligodeoxynucleotide enhances the efficacy of anticancer monoclonal antibody in an in vivo xenograft model using human endometrial cancer cell. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2572. doi:10.1158/1538-7445.AM2014-2572

Critical role for bone marrow stromal antigen 2 in acute Chikungunya virus infection.

Bone marrow stromal antigen 2 (BST-2; also known as tetherin or CD317) is an IFN-inducible gene that functions to block the release of a range of nascent enveloped virions from infected host cells. However, the role of BST-2 in viral pathogenesis remains poorly understood. BST-2 plays a multifaceted role in innate immunity, as it hinders retroviral infection and possibly promotes infection with some rhabdo- and orthomyxoviruses. This paradoxical role has probably hindered exploration of BST-2 antiviral function in vivo. We reported previously that BST-2 tethers Chikungunya virus (CHIKV)-like particles on the cell plasma membrane. To explore the role of BST-2 in CHIKV replication and host protection, we utilized CHIKV strain 181/25 to examine early events during CHIKV infection in a BST-2(-/-) mouse model. We observed an interesting dichotomy between WT and BST-2(-/-) mice. BST-2 deficiency increased inoculation site viral load, culminating in higher systemic viraemia and increased lymphoid tissues tropism. A suppressed inflammatory innate response demonstrated by impaired expression of IFN-α, IFN-γ and CD40 ligand was observed in BST-2(-/-) mice compared with the WT controls. These findings suggested that, in part, BST-2 protects lymphoid tissues from CHIKV infection and regulates CHIKV-induced inflammatory response by the host.

BST2/CD317 counteracts human coronavirus 229E productive infection by tethering virions at the cell surface

Bone marrow stromal antigen 2 (BST2), an interferon-inducible antiviral factor, has been shown to block the release of various enveloped viruses from cells. It has also been identified as an innate immune system component. Most enveloped viruses subject to BST2 restriction bud at the plasma membrane. Here we report our findings that (a) the production of human coronavirus 229E (HCoV-229E) progeny viruses, whose budding occurs at the ER-Golgi intermediate compartment (ERGIC), markedly decreases in the presence of BST2; and (b) BST2 knockdown expression results in enhanced HCoV-229E virion production. Electron microscopy analyses indicate that HCoV-229E virions are tethered to cell surfaces or intracellular membranes by BST2. Our results suggest that BST2 exerts a broad blocking effect against enveloped virus release, regardless of whether budding occurs at the plasma membrane or intracellular compartments.

BST-2/tetherin-mediated restriction of chikungunya (CHIKV) VLP budding is counteracted by CHIKV non-structural protein 1 (nsP1)

Chikungunya virus (CHIKV) is a re-emerging alphavirus transmitted by Aedes mosquitoes. Infection with CHIKV elicits a type I interferon response that facilities virus clearance, probably through the action of down-stream effectors such as antiviral IFN-stimulated genes (ISGs). Bone marrow stromal antigen 2 (BST-2) is an ISG shown to restrict HIV-1 replication by preventing the infection of bystander cells by tethering progeny virions on the surface of infected cells. Here we show that enrichment of cell surface BST-2 results in retention of CHIKV virus like particles (VLPs) on the cell membrane. BST-2 was found to co-localize with CHIKV structural protein E1 in the context of VLPs without any noticeable effect on BST-2 level. However, CHIKV nonstructural protein 1 (nsP1) overcomes BST-2-mediated VLPs tethering by down-regulating BST-2 expression. We conclude that BST-2 tethers CHIKV VLPs on the host cell plasma membrane and identify CHIKV nsP1 as a novel BST-2 antagonist.

Plasma membrane proteomics identifies bone marrow stromal antigen 2 as a potential therapeutic target in endometrial cancer

This report utilizes a novel proteomic method for discovering potential therapeutic targets in endometrial cancer. We used a biotinylation-based approach for cell-surface protein enrichment combined with isobaric tags for relative and absolute quantitation (iTRAQ) technology using nano liquid chromatography-tandem mass spectrometry analysis to identify specifically overexpressed proteins in endometrial cancer cells compared with normal endometrial cells. We identified a total of 272 proteins, including 11 plasma membrane proteins, whose expression increased more than twofold in at least four of seven endometrial cancer cell lines compared with a normal endometrial cell line. Overexpression of bone marrow stromal antigen 2 (BST2) was detected and the observation was supported by immunohistochemical analysis using clinical samples. The expression of BST2 was more characteristic of 118 endometrial cancer tissues compared with 59 normal endometrial tissues (p < 0.0001). The therapeutic effect of an anti-BST2 antibody was studied both in vitro and in vivo. An anti-BST2 monoclonal antibody showed in vitro cytotoxicity in BST2-positive endometrial cancer cells via antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity. In an in vivo xenograft model, anti-BST2 antibody treatment significantly inhibited tumor growth of BST2-positive endometrial cancer cells in an NK cell-dependent manner. The anti-BST2 antibody had a potent antitumor effect against endometrial cancer both in vitro and in vivo, indicating a strong potential for clinical use of anti-BST2 antibody for endometrial cancer treatment. The combination of biotinylation-based enrichment of cell-surface proteins and iTRAQ analysis should be a useful screening method for future discovery of potential therapeutic targets.

Intracellular Logistics of BST-2/Tetherin

Bone marrow stromal antigen 2 (BST-2) is a type II membrane protein with two targeting signals, one of which is located in the cytoplasmic domain and contains a non-canonical dual tyrosine-based motif responsible for its endocytosis from the plasma membrane, and the other is a C-terminal glycosylphosphatidylinositol anchor that facilitates its association with detergent-resistant membranes/lipid rafts and targeting to the apical domain in polarized epithelial cells. Due to its unusual topology at the membrane, BST-2 takes unique and complicated trafficking routes in cells. Recently, a physiological role for BST-2 as the "tetherin" molecule for viruses, especially for HIV-1, has been extensively examined. These studies have shown that the biosynthesis, intracellular trafficking, localization, and structure of human BST-2 are closely related to its antiviral activity. This review provides an overview of the intracellular logistics of human BST-2.

Role of the Endosomal ESCRT Machinery in HIV-1 Vpu-Induced Down- Regulation of BST2/Tetherin

The cellular protein "Bone marrow stromal antigen 2" (BST2 also called Tetherin, CD317, HM1.24) was identified as a major mediator of the innate immune defense against the dissemination of enveloped viruses. BST2 was shown to physically trap the de novo formed viral particles at the surface of infected cells, thereby reducing viral release. Lentiviruses have evolved specific strategies to down-regulate the expression level of BST2 from the surface of the cells and as such promote viral egress. In Human Immunodeficiency Virus-1 (HIV-1), the accessory protein Vpu counters BST2 antiviral activity. However, the cellular and molecular mechanisms involved are not fully understood. Vpu-mediated antagonism of BST2 antiviral activity seems to involve complex interplay between the viral protein and host components regulating protein turnover and vesicular trafficking. This review focuses on the interplay between Vpu and the ubiquitin/endosomal pathway in countermeasures of HIV-1 to BST2 restriction, with a particular emphasis on the "Endosomal Sorting Complexes Required for Transport" (ESCRT) machinery.

Sites of Action of HIV-1 Vpu in BST-2/Tetherin Downregulation

The interferon-inducible host restriction factor bone marrow stromal antigen 2 (BST-2/tetherin) blocks the release of human immunodeficiency virus type 1 (HIV-1) by directly cross-linking virions to the membrane of infected cells. This antiviral effect is counteracted by the HIV-1 accessory protein viral protein U (Vpu) through mechanisms that remain unclear. Accumulating evidence suggests that Vpu antagonizes BST-2 by removing it from the plasma membrane; however, neither the cellular sites of interaction nor the effector mechanisms that result in the downregulation of BST-2 cell-surface expression have been fully determined. Based on current evidence regarding the subcellular localization of Vpu and BST-2 and the latter's trafficking defects induced by their interaction, three models have been proposed. In the first, Vpu is hypothesized to block the traffic of newly synthesized BST-2 towards the cell surface by retaining it in the biosynthetic/secretory compartment. The second model suggests that Vpu sequesters BST-2 within intracellular compartments corresponding to recycling endosomes and the trans-Golgi network by blocking its recycling after endocytosis. In the third model, we and others have proposed that Vpu directly internalizes BST-2 from the plasma membrane and induces its enhanced endolysosomal trafficking and degradation. As for its intracellular fate, the viral antagonism of BST-2 is likely dependent on the intracellular sequestration, or the proteasomal/lysosomal degradation of the restriction factor. This review summarizes the current advances in our understanding of the cellular pathways and sites of action of Vpu in the downregulation of cell-surface BST-2.

Ancient origin of a deletion in human BST2/Tetherin that confers protection against viral zoonoses

Bone marrow stromal antigen 2 (BST2)/Tetherin is an antiviral factor that blocks the release of enveloped virions from infected cells. Recent data suggest that efficient BST2 antagonism was a prerequisite for the global spread of HIV/AIDS. Most simian immunodeficiency viruses (SIVs), including the direct precursors of HIV, use their Nef protein to antagonize BST2 of their respective host species. Human BST2, however, contains a five amino acid deletion in its cytoplasmic domain that confers resistance to Nef. Thus, this antiviral factor constitutes a major barrier to effective zoonotic transmissions of SIVs to humans. Here, we show that this protective deletion has already been present in Neanderthal and Denisovan BST2 and thus evolved at least 800,000 years ago. This ancient origin helps to explain why effectively spreading zoonotic transmissions of SIVs to humans have been rare, although SIVs are widespread in African nonhuman primates and humans must have been exposed to these viruses many times.

Structural Basis for the Antiviral Activity of BST-2/Tetherin and Its Viral Antagonism

The interferon-inducible host restriction factor bone marrow stromal antigen 2 (BST-2/tetherin) blocks the release of HIV-1 and other enveloped viruses. In turn, these viruses have evolved specific antagonists to counteract this host antiviral molecule, such as the HIV-1 protein Vpu. BST-2 is a type II transmembrane protein with an unusual topology consisting of an N-terminal cytoplasmic tail (CT) followed by a single transmembrane (TM) domain, a coiled-coil extracellular (EC) domain, and a glycosylphosphatidylinositol (GPI) anchor at the C terminus. We and others showed that BST-2 restricts enveloped virus release by bridging the host and virion membranes with its two opposing membrane anchors and that deletion of either one completely abrogates antiviral activity. The EC domain also shows conserved structural properties that are required for antiviral function. It contains several destabilizing amino acids that confer the molecule with conformational flexibility to sustain the protein’s function as a virion tether, and three conserved cysteine residues that mediate homodimerization of BST-2, as well as acting as a molecular ruler that separates the membrane anchors. Conversely, the efficient release of virions is promoted by the HIV-1 Vpu protein and other viral antagonists. Our group and others provided evidence from mutational analyses indicating that Vpu antagonism of BST-2-mediated viral restriction requires a highly specific interaction of their mutual TM domains. This interpretation is further supported and expanded by the findings of the latest structural modeling studies showing that critical amino acids in a conserved helical face of these TM domains are required for Vpu–BST-2 interaction and antagonism. In this review, we summarize the current advances in our understanding of the structural basis for BST-2 antiviral function as well as BST-2-specific viral antagonism.

The Interferon-Inducible Host Factor Bone Marrow Stromal Antigen 2/Tetherin Restricts Virion Release, but Is It Actually a Viral Restriction Factor?

Viruses face a variety of obstacles when infecting a new host. The past few years have brought exciting new insights into the function of restriction factors, which form part of the host's innate immune system. One of the most recently identified restriction factors is bone marrow stromal antigen 2 (BST-2)/tetherin. BST-2 is an interferon-inducible gene whose expression dramatically reduces the release of viruses from infected cells. This effect of BST-2 is not specific to human immunodeficiency virus but affects a broad range of enveloped viruses. Since the identification of BST-2 as a restriction factor in 2008, much progress has been made in understanding the molecular properties and functional characteristics of this host factor. The goal of this review was to provide an update on our current understanding of the role of BST-2 in regulating virus release and to discuss its role in controlling virus spread during productive infection with special emphasis on human immunodeficiency virus-1.

The Vpu protein: new concepts in virus release and CD4 down-modulation.

Human immunodeficiency virus type 1 (HIV-1) and several simian immunodeficiency viruses (SIV) encode for a transmembrane protein known as Vpu (viral protein U). While one of the smallest of the HIV-1 proteins, it has two important functions within virus-infected cells. The first of these functions is the down-regulation of the CD4 receptor to prevent its interaction with the HIV-1 envelope glycoprotein. Vpu interacts with the CD4 receptor in the rough endoplasmic reticulum (RER), resulting in its re-translocation across the RER and subsequent degradation via the proteasomal pathway. The second major function of the Vpu protein is to facilitate release of virus from infected cells. Previous studies have shown that virus release is cell type specific, suggesting that certain cells may express a restriction factor that inhibits virus release in the absence of Vpu. Recently, bone marrow stromal antigen 2 (BST-2/HM1.24/CD317/tetherin) has been identified as this factor. This review will focus on new findings within the last four years on the role of Vpu in CD4 down-regulation and the restriction of virus release from cells. We will relate these findings to virus pathogenesis and propose questions regarding BST-2 as a restriction factor.

HIV-1 Accessory Protein Vpu Internalizes Cell-surface BST-2/Tetherin through Transmembrane Interactions Leading to Lysosomes

Bone marrow stromal antigen 2 (BST-2, also known as tetherin) is a recently identified interferon-inducible host restriction factor that can block the production of enveloped viruses by trapping virus particles at the cell surface. This antiviral effect is counteracted by the human immunodeficiency virus type 1 (HIV-1) accessory protein viral protein U (Vpu). Here we show that HIV-1 Vpu physically interacts with BST-2 through their mutual transmembrane domains and leads to the degradation of this host factor via a lysosomal, not proteasomal, pathway. The degradation is partially controlled by a cellular protein, beta-transducin repeat-containing protein (betaTrCP), which is known to be required for the Vpu-induced degradation of CD4. Importantly, targeting of BST-2 by Vpu occurs at the plasma membrane followed by the active internalization of this host protein by Vpu independently of constitutive endocytosis. Thus, the primary site of action of Vpu is the plasma membrane, where Vpu targets and internalizes cell-surface BST-2 through transmembrane interactions, leading to lysosomal degradation, partially in a betaTrCP-dependent manner. Also, we propose the following configuration of BST-2 in tethering virions to the cell surface; each of the dimerized BST-2 molecules acts as a bridge between viral and cell membranes.