The Experts below are selected from a list of 2526 Experts worldwide ranked by ideXlab platform
Abraham L Brass - One of the best experts on this subject based on the ideXlab platform.
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IFITM3 proximity to viral fusion proteins allows virus restriction.
2019Co-Authors: Krishna C. Suddala, Paul Meraner, Abraham L Brass, Ruben M. Markosyan, Christine C. Lee, Mariana Marin, Tanay M. Desai, Fredric S. Cohen, Gregory B. MelikyanAbstract:(A-D) IFITM3 incorporates into virions and restricts viral fusion in A549 cells. (A) Western blot of control and IFITM3-containing IAVpp prepared through co-transfection of 293T/17 cells with additional plasmids–either empty vector (Vector) or IFITM3 (IFITM3), respectively. Blots were probed with anti-IFITM3, anti-p24, and anti-influenza HA antibodies. (B) IAVpp containing BlaM-Vpr and lacking or containing IFITM3 in the presence or absence of AmphoB were used to infect A549 cells and the resulting viral fusion measured by the BlaM assay. Data are means and SEM based on three experiments performed in triplicate (except for IAVpp/IFITM3 treated with AmphoB, which was performed twice in triplicate). (C) LASV GPc induced cell-cell fusion is inhibited by IFITMs. Co-cultures of 293T cells stably expressing IFITM1, IFITM2 or IFITM3 with Cos7 transiently expressing LASV GPc were co-cultured for 30 min at room temperature and exposed to a pH 5.0 citrate buffer for 20 min at 37°C to induce LASV fusion. The extend of fusion was measured by microscopy-based dye transfer assay, as described in Materials and Methods. Data are mean with SEM from 3 independent experiments. (See also S6 Fig).
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The IFITMs Inhibit Zika Virus Replication
Cell Reports, 2016Co-Authors: George Savidis, Jill M. Perreira, Paul Meraner, Zhiru Guo, Sharone Green, Jocelyn M. Portmann, Abraham L BrassAbstract:Summary Zika virus has emerged as a severe health threat with a rapidly expanding range. The IFITM family of restriction factors inhibits the replication of a broad range of viruses, including the closely related flaviruses West Nile virus and dengue virus. Here, we show that IFITM1 and IFITM3 inhibit Zika virus infection early in the viral life cycle. Moreover, IFITM3 can prevent Zika-virus-induced cell death. These results suggest that strategies to boost the actions and/or levels of the IFITMs might be useful for inhibiting a broad range of emerging viruses.
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IFITM3 restricts influenza a virus entry by blocking the formation of fusion pores following virus endosome hemifusion
PLOS Pathogens, 2014Co-Authors: Tanay M. Desai, George Savidis, Abraham L Brass, Christopher R Chin, Mariana Marin, Gregory B. MelikyanAbstract:Interferon-induced transmembrane proteins (IFITMs) inhibit infection of diverse enveloped viruses, including the influenza A virus (IAV) which is thought to enter from late endosomes. Recent evidence suggests that IFITMs block virus hemifusion (lipid mixing in the absence of viral content release) by altering the properties of cell membranes. Consistent with this mechanism, excess cholesterol in late endosomes of IFITM-expressing cells has been reported to inhibit IAV entry. Here, we examined IAV restriction by IFITM3 protein using direct virus-cell fusion assay and single virus imaging in live cells. IFITM3 over-expression did not inhibit lipid mixing, but abrogated the release of viral content into the cytoplasm. Although late endosomes of IFITM3-expressing cells accumulated cholesterol, other interventions leading to aberrantly high levels of this lipid did not inhibit virus fusion. These results imply that excess cholesterol in late endosomes is not the mechanism by which IFITM3 inhibits the transition from hemifusion to full fusion. The IFITM3's ability to block fusion pore formation at a post-hemifusion stage shows that this protein stabilizes the cytoplasmic leaflet of endosomal membranes without adversely affecting the lumenal leaflet. We propose that IFITM3 interferes with pore formation either directly, through partitioning into the cytoplasmic leaflet of a hemifusion intermediate, or indirectly, by modulating the lipid/protein composition of this leaflet. Alternatively, IFITM3 may redirect IAV fusion to a non-productive pathway, perhaps by promoting fusion with intralumenal vesicles within multivesicular bodies/late endosomes.
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The Viral Restriction Factor IFITM3 Promotes Hemifusion but Blocks Full Fusion of Influenza Virus
Biophysical Journal, 2014Co-Authors: Tanay M. Desai, George Savidis, Abraham L Brass, Christopher R Chin, Mariana Marin, Gregory B. MelikyanAbstract:Interferon-induced transmembrane proteins (IFITMs) are up-regulated as part of a cell's defense against viral challenges. These small proteins inhibit entry of diverse viruses, such as influenza A (IAV), West Nile and dengue virus. The mechanism by which IFITMs block viral entry is not understood. Recent reports suggest that IFITMs inhibits viral hemifusion (merger of proximal leaflets of the viral and cellular membranes), presumably by disrupting cholesterol trafficking and causing aberrant cholesterol accumulation in late endosomes. Here we employed time-resolved single IAV imaging to identify fusion step(s) affected by IFITM3 protein ectopically expressed in lung epithelial cells. These experiments revealed that, contrary to previous reports, lipid mixing between IAV and endosomes was, in fact, promoted upon IFITM3 expression. In contrast, virus-cell fusion assays monitoring the release of the viral content showed marked inhibition of fusion pore formation by IFITM3. This effect was not due to excessive cholesterol accumulation in endosomes or to reduced endosome acidity in IFITM3-expressing cells. Furthermore, conditions that induced the accumulation of cholesterol in late endosomes/lysosomes did not restrict IAV fusion. To conclude, IFITM3 blocks IAV fusion by disfavoring the formation of fusion pores, and this phenomenon is independent of cholesterol levels. We propose that IFITM3 can block viral fusion at a post-hemifusion stage either directly, by inserting into and stabilizing the cytosolic leaflet of endosomal membranes, or indirectly, by altering the lipid composition and thus disfavoring the formation of fusion pores. This work was partially supported by the NIH R01 GM054787 (to GBM) and 1R01AI091786 (to ALB) grants.
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Interferon Inducible Transmembrane Protein 3 (IFITM3) Restricts Reovirus Cell Entry
The Journal of biological chemistry, 2013Co-Authors: Amanda A. Anafu, Abraham L Brass, Christopher R Chin, Christopher H. Bowen, Geoffrey H. HolmAbstract:Reoviruses are double-stranded RNA viruses that infect the mammalian respiratory and gastrointestinal tract. Reovirus infection elicits production of type I interferons (IFNs), which trigger antiviral pathways through the induction of interferon-stimulated genes (ISGs). Although hundreds of ISGs have been identified, the functions of many of these genes are unknown. The interferon-inducible transmembrane (IFITM) proteins are one class of ISGs that restrict the cell entry of some enveloped viruses, including influenza A virus. One family member, IFITM3, localizes to late endosomes, where reoviruses undergo proteolytic disassembly; therefore, we sought to determine whether IFITM3 also restricts reovirus entry. IFITM3-expressing cell lines were less susceptible to infection by reovirus, as they exhibited significantly lower percentages of infected cells in comparison to control cells. Reovirus replication was also significantly reduced in IFITM3-expressing cells. Additionally, cells expressing an shRNA targeting IFITM3 exhibited a smaller decrease in infection after IFN treatment than the control cells, indicating that endogenous IFITM3 restricts reovirus infection. However, IFITM3 did not restrict entry of reovirus infectious subvirion particles (ISVPs), which do not require endosomal proteolysis, indicating that restriction occurs in the endocytic pathway. Proteolysis of outer capsid protein μ1 was delayed in IFITM3-expressing cells in comparison to control cells, suggesting that IFITM3 modulates the function of late endosomal compartments either by reducing the activity of endosomal proteases or delaying the proteolytic processing of virions. These data provide the first evidence that IFITM3 restricts infection by a nonenveloped virus and suggest that IFITM3 targets an increasing number of viruses through a shared requirement for endosomes during cell entry.
Christopher R Chin - One of the best experts on this subject based on the ideXlab platform.
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IFITM3 restricts influenza a virus entry by blocking the formation of fusion pores following virus endosome hemifusion
PLOS Pathogens, 2014Co-Authors: Tanay M. Desai, George Savidis, Abraham L Brass, Christopher R Chin, Mariana Marin, Gregory B. MelikyanAbstract:Interferon-induced transmembrane proteins (IFITMs) inhibit infection of diverse enveloped viruses, including the influenza A virus (IAV) which is thought to enter from late endosomes. Recent evidence suggests that IFITMs block virus hemifusion (lipid mixing in the absence of viral content release) by altering the properties of cell membranes. Consistent with this mechanism, excess cholesterol in late endosomes of IFITM-expressing cells has been reported to inhibit IAV entry. Here, we examined IAV restriction by IFITM3 protein using direct virus-cell fusion assay and single virus imaging in live cells. IFITM3 over-expression did not inhibit lipid mixing, but abrogated the release of viral content into the cytoplasm. Although late endosomes of IFITM3-expressing cells accumulated cholesterol, other interventions leading to aberrantly high levels of this lipid did not inhibit virus fusion. These results imply that excess cholesterol in late endosomes is not the mechanism by which IFITM3 inhibits the transition from hemifusion to full fusion. The IFITM3's ability to block fusion pore formation at a post-hemifusion stage shows that this protein stabilizes the cytoplasmic leaflet of endosomal membranes without adversely affecting the lumenal leaflet. We propose that IFITM3 interferes with pore formation either directly, through partitioning into the cytoplasmic leaflet of a hemifusion intermediate, or indirectly, by modulating the lipid/protein composition of this leaflet. Alternatively, IFITM3 may redirect IAV fusion to a non-productive pathway, perhaps by promoting fusion with intralumenal vesicles within multivesicular bodies/late endosomes.
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The Viral Restriction Factor IFITM3 Promotes Hemifusion but Blocks Full Fusion of Influenza Virus
Biophysical Journal, 2014Co-Authors: Tanay M. Desai, George Savidis, Abraham L Brass, Christopher R Chin, Mariana Marin, Gregory B. MelikyanAbstract:Interferon-induced transmembrane proteins (IFITMs) are up-regulated as part of a cell's defense against viral challenges. These small proteins inhibit entry of diverse viruses, such as influenza A (IAV), West Nile and dengue virus. The mechanism by which IFITMs block viral entry is not understood. Recent reports suggest that IFITMs inhibits viral hemifusion (merger of proximal leaflets of the viral and cellular membranes), presumably by disrupting cholesterol trafficking and causing aberrant cholesterol accumulation in late endosomes. Here we employed time-resolved single IAV imaging to identify fusion step(s) affected by IFITM3 protein ectopically expressed in lung epithelial cells. These experiments revealed that, contrary to previous reports, lipid mixing between IAV and endosomes was, in fact, promoted upon IFITM3 expression. In contrast, virus-cell fusion assays monitoring the release of the viral content showed marked inhibition of fusion pore formation by IFITM3. This effect was not due to excessive cholesterol accumulation in endosomes or to reduced endosome acidity in IFITM3-expressing cells. Furthermore, conditions that induced the accumulation of cholesterol in late endosomes/lysosomes did not restrict IAV fusion. To conclude, IFITM3 blocks IAV fusion by disfavoring the formation of fusion pores, and this phenomenon is independent of cholesterol levels. We propose that IFITM3 can block viral fusion at a post-hemifusion stage either directly, by inserting into and stabilizing the cytosolic leaflet of endosomal membranes, or indirectly, by altering the lipid composition and thus disfavoring the formation of fusion pores. This work was partially supported by the NIH R01 GM054787 (to GBM) and 1R01AI091786 (to ALB) grants.
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amphotericin b increases influenza a virus infection by preventing IFITM3 mediated restriction
Cell Reports, 2013Co-Authors: Tsaiyu Lin, George Savidis, Jill M. Perreira, Christopher R Chin, Aaron R Everitt, Simon Clare, Aaron M Aker, Sinu P John, David SarlahAbstract:The IFITMs inhibit influenza A virus (IAV) replication in vitro and in vivo. Here, we establish that the antimycotic heptaen, amphotericin B (AmphoB), prevents IFITM3-mediated restriction of IAV, thereby increasing viral replication. Consistent with its neutralization of IFITM3, a clinical preparation of AmphoB, AmBisome, reduces the majority of interferon’s protective effect against IAV in vitro. Mechanistic studies reveal that IFITM1 decreases host-membrane fluidity, suggesting both a possible mechanism for IFITM-mediated restriction and its negation by AmphoB. Notably, we reveal that mice treated with AmBisome succumbed to a normally mild IAV infection, similar to animals deficient in IFITM3. Therefore, patients receiving antifungal therapy with clinical preparations of AmphoB may be functionally immunocompromised and thus more vulnerable to influenza, as well as other IFITM3-restricted viral infections.
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the cd225 domain of IFITM3 is required for both ifitm protein association and inhibition of influenza a virus and dengue virus replication
Journal of Virology, 2013Co-Authors: Sinu P John, George Savidis, Jill M. Perreira, Sarah E Smith, Christopher R Chin, Aaron M Aker, Eric M Feeley, Andrew E H Elia, Aaron R EverittAbstract:The interferon-induced transmembrane protein 3 (IFITM3) gene is an interferon-stimulated gene that inhibits the replication of multiple pathogenic viruses in vitro and in vivo. IFITM3 is a member of a large protein superfamily, whose members share a functionally undefined area of high amino acid conservation, the CD225 domain. We performed mutational analyses of IFITM3 and identified multiple residues within the CD225 domain, consisting of the first intramembrane domain (intramembrane domain 1 [IM1]) and a conserved intracellular loop (CIL), that are required for restriction of both influenza A virus (IAV) and dengue virus (DENV) infection in vitro. Two phenylalanines within IM1 (F75 and F78) also mediate a physical association between IFITM proteins, and the loss of this interaction decreases IFITM3-mediated restriction. By extension, similar IM1-mediated associations may contribute to the functions of additional members of the CD225 domain family. IFITM3's distal N-terminal domain is also needed for full antiviral activity, including a tyrosine (Y20), whose alteration results in mislocalization of a portion of IFITM3 to the cell periphery and surface. Comparative analyses demonstrate that similar molecular determinants are needed for IFITM3's restriction of both IAV and DENV. However, a portion of the CIL including Y99 and R87 is preferentially needed for inhibition of the orthomyxovirus. Several IFITM3 proteins engineered with rare single-nucleotide polymorphisms demonstrated reduced expression or mislocalization, and these events were associated with enhanced viral replication in vitro, suggesting that possessing such alleles may impact an individual's risk for viral infection. On the basis of this and other data, we propose a model for IFITM3-mediated restriction.
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Interferon Inducible Transmembrane Protein 3 (IFITM3) Restricts Reovirus Cell Entry
The Journal of biological chemistry, 2013Co-Authors: Amanda A. Anafu, Abraham L Brass, Christopher R Chin, Christopher H. Bowen, Geoffrey H. HolmAbstract:Reoviruses are double-stranded RNA viruses that infect the mammalian respiratory and gastrointestinal tract. Reovirus infection elicits production of type I interferons (IFNs), which trigger antiviral pathways through the induction of interferon-stimulated genes (ISGs). Although hundreds of ISGs have been identified, the functions of many of these genes are unknown. The interferon-inducible transmembrane (IFITM) proteins are one class of ISGs that restrict the cell entry of some enveloped viruses, including influenza A virus. One family member, IFITM3, localizes to late endosomes, where reoviruses undergo proteolytic disassembly; therefore, we sought to determine whether IFITM3 also restricts reovirus entry. IFITM3-expressing cell lines were less susceptible to infection by reovirus, as they exhibited significantly lower percentages of infected cells in comparison to control cells. Reovirus replication was also significantly reduced in IFITM3-expressing cells. Additionally, cells expressing an shRNA targeting IFITM3 exhibited a smaller decrease in infection after IFN treatment than the control cells, indicating that endogenous IFITM3 restricts reovirus infection. However, IFITM3 did not restrict entry of reovirus infectious subvirion particles (ISVPs), which do not require endosomal proteolysis, indicating that restriction occurs in the endocytic pathway. Proteolysis of outer capsid protein μ1 was delayed in IFITM3-expressing cells in comparison to control cells, suggesting that IFITM3 modulates the function of late endosomal compartments either by reducing the activity of endosomal proteases or delaying the proteolytic processing of virions. These data provide the first evidence that IFITM3 restricts infection by a nonenveloped virus and suggest that IFITM3 targets an increasing number of viruses through a shared requirement for endosomes during cell entry.
Alex A. Compton - One of the best experts on this subject based on the ideXlab platform.
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Homology-guided identification of a conserved motif linking the antiviral functions of IFITM3 to its oligomeric state
eLife, 2020Co-Authors: Kazi Rahman, Saliha Majdoul, Charles A. Coomer, Selena Y Ding, Sergi Padilla-parra, Alex A. ComptonAbstract:The interferon-inducible transmembrane (IFITM) proteins belong to the Dispanin/CD225 family and inhibit diverse virus infections. IFITM3 reduces membrane fusion between cells and virions through a poorly characterized mechanism. Mutation of proline-rich transmembrane protein 2 (PRRT2), a regulator of neurotransmitter release, at glycine-305 was previously linked to paroxysmal neurological disorders in humans. Here, we show that glycine-305 and the homologous site in IFITM3, glycine-95, drive protein oligomerization from within a GxxxG motif. Mutation of glycine-95 (and to a lesser extent, glycine-91) disrupted IFITM3 oligomerization and reduced its antiviral activity against Influenza A virus. An oligomerization-defective variant was used to reveal that IFITM3 promotes membrane rigidity in a glycine-95-dependent and amphipathic helix-dependent manner. Furthermore, a compound which counteracts virus inhibition by IFITM3, Amphotericin B, prevented the IFITM3-mediated rigidification of membranes. Overall, these data suggest that IFITM3 oligomers inhibit virus-cell fusion by promoting membrane rigidity.
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Homology-guided identification of a conserved motif linking the antiviral functions of IFITM3 to its oligomeric state
2020Co-Authors: Kazi Rahman, Saliha Majdoul, Charles A. Coomer, Selena Y Ding, Sergi Padilla-parra, Alex A. ComptonAbstract:Abstract The interferon-inducible transmembrane (IFITM) proteins belong to the Dispanin/CD225 family and inhibit diverse virus infections. IFITM3 reduces membrane fusion between cells and virions through a poorly characterized mechanism. We identified a GxxxG motif in many CD225 proteins, including IFITM3 and proline rich transmembrane protein 2 (PRRT2). Mutation of PRRT2, a regulator of neurotransmitter release, at glycine-305 was previously linked to paroxysmal neurological disorders in humans. Here, we show that glycine-305 and the homologous site in IFITM3, glycine-95, drive protein oligomerization from within a GxxxG motif. Mutation of glycine-95 in IFITM3 disrupted its oligomerization and reduced its antiviral activity against Influenza A and HIV-1. The oligomerization-defective variant was used to reveal that IFITM3 promotes membrane rigidity in a glycine-95-dependent manner. Furthermore, a compound which counteracts virus inhibition by IFITM3, amphotericin B, prevented the IFITM3-mediated rigidification of membranes. Overall, these data suggest that IFITM3 oligomers inhibit virus-cell fusion by promoting membrane rigidity.
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IFITM3 Reduces Retroviral Envelope Abundance and Function and Is Counteracted by glycoGag.
mBio, 2020Co-Authors: Yadvinder S. Ahi, Diborah Yimer, Guoli Shi, Saliha Majdoul, Kazi Rahman, Alan Rein, Alex A. ComptonAbstract:Interferon-induced transmembrane (IFITM) proteins are encoded by many vertebrate species and exhibit antiviral activities against a wide range of viruses. IFITM3, when present in virus-producing cells, reduces the fusion potential of HIV-1 virions, but the mechanism is poorly understood. To define the breadth and mechanistic basis for the antiviral activity of IFITM3, we took advantage of a murine leukemia virus (MLV)-based pseudotyping system. By carefully controlling amounts of IFITM3 and envelope protein (Env) in virus-producing cells, we found that IFITM3 potently inhibits MLV infectivity when Env levels are limiting. Loss of infectivity was associated with defective proteolytic processing of Env and lysosomal degradation of the Env precursor. Ecotropic and xenotropic variants of MLV Env, as well as HIV-1 Env and vesicular stomatitis virus glycoprotein (VSV-G), are sensitive to IFITM3, whereas Ebola glycoprotein is resistant, suggesting that IFITM3 selectively inactivates certain viral glycoproteins. Furthermore, endogenous IFITM3 in human and murine cells negatively regulates MLV Env abundance. However, we found that the negative impact of IFITM3 on virion infectivity is greater than its impact on decreasing Env incorporation, suggesting that IFITM3 may impair Env function, as well as reduce the amount of Env in virions. Finally, we demonstrate that loss of virion infectivity mediated by IFITM3 is reversed by the expression of glycoGag, a murine retrovirus accessory protein previously shown to antagonize the antiviral activity of SERINC proteins. Overall, we show that IFITM3 impairs virion infectivity by regulating Env quantity and function but that enhanced Env expression and glycoGag confer viral resistance to IFITM3.IMPORTANCE The viral envelope glycoprotein, known as "Env" in Retroviridae, is found on the virion surface and facilitates virus entry into cells by mediating cell attachment and fusion. Env is a major structural component of retroviruses and is targeted by all arms of the immune response, including adaptive and innate immunity. Less is known about how cell-intrinsic immunity prevents retrovirus replication at the level of individual cells. Here, we show that cellular IFITM3 and IFITM2 inhibit the fusion potential of retroviral virions by inhibiting Env protein via a two-pronged mechanism. IFITM proteins inhibit Env abundance in cells and also impair its function when levels are low. The posttranslational block of retroviral Env function by IFITM proteins is likely to impede both exogenous and endogenous retrovirus replication. In support of a relevant role for IFITM3 in retrovirus control, the retroviral accessory protein glycoGag counteracts IFITM3 function to promote virus infectivity.
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mTOR inhibitors lower an intrinsic barrier to virus infection mediated by IFITM3.
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Guoli Shi, Stosh Ozog, Bruce E. Torbett, Alex A. ComptonAbstract:Rapamycin and its derivatives are specific inhibitors of mammalian target of rapamycin (mTOR) kinase and, as a result, are well-established immunosuppressants and antitumorigenic agents. Additionally, this class of drug promotes gene delivery by facilitating lentiviral vector entry into cells, revealing its potential to improve gene therapy efforts. However, the precise mechanism was unknown. Here, we report that mTOR inhibitor treatment results in down-regulation of the IFN-induced transmembrane (IFITM) proteins. IFITM proteins, especially IFITM3, are potent inhibitors of virus-cell fusion and are broadly active against a range of pathogenic viruses. We found that the effect of rapamycin treatment on lentiviral transduction is diminished upon IFITM silencing or knockout in primary and transformed cells, and the extent of transduction enhancement depends on basal expression of IFITM proteins, with a major contribution from IFITM3. The effect of rapamycin treatment on IFITM3 manifests at the level of protein, but not mRNA, and is selective, as many other endosome-associated transmembrane proteins are unaffected. Rapamycin-mediated degradation of IFITM3 requires endosomal trafficking, ubiquitination, endosomal sorting complex required for transport (ESCRT) machinery, and lysosomal acidification. Since IFITM proteins exhibit broad antiviral activity, we show that mTOR inhibition also promotes infection by another IFITM-sensitive virus, Influenza A virus, but not infection by Sendai virus, which is IFITM-resistant. Our results identify the molecular basis by which mTOR inhibitors enhance virus entry into cells and reveal a previously unrecognized immunosuppressive feature of these clinically important drugs. In addition, this study uncovers a functional convergence between the mTOR pathway and IFITM proteins at endolysosomal membranes.
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More than meets the I: the diverse antiviral and cellular functions of interferon-induced transmembrane proteins
Retrovirology, 2017Co-Authors: Guoli Shi, Olivier Schwartz, Alex A. ComptonAbstract:The first responders of human antiviral immunity are components of the intrinsic immune response that reside within each and every one of our cells. This cell-autonomous arsenal consists of nucleic acid sensors and antiviral effectors strategically placed by evolution to detect and restrict invading viruses. While some factors are present at baseline to allow for constant surveillance of the cell interior, others are upregulated by cytokines (such as interferons) that signal a viral infection underway in neighboring cells. In this review, we highlight the multiple roles played by the interferon-induced transmembrane (IFITM) proteins during viral infection, with focuses on IFITM3 and HIV-1. Moreover, we discuss the cellular pathways in which IFITM proteins are intertwined and the various functions they have been ascribed outside the context of infection. While appreciated as broadly-acting, potent restriction factors that prevent virus infection and pathogenesis in cell culture and in vivo, questions remain regarding their precise mode of action and importance in certain viral contexts. Continued efforts to study IFITM protein function will further cement their status as critical host determinants of virus susceptibility and prioritize them in the development of new antiviral therapies.
Chen Liang - One of the best experts on this subject based on the ideXlab platform.
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Interferon inducible transmembrane protein 3 (IFITM3) inhibits influenza virus replication and inflammation by interacting with ABHD16A
2020Co-Authors: Chen Liang, Zhu Limei, Chen JunAbstract:IFITM family proteins have broad-spectrum antiviral capabilities. Preliminary studies in our laboratory have preliminarily proved that IFITMs have the effect of inhibiting influenza viruses. In order to further study its mechanism and role in the occurrence and development of influenza, relevant studies have been carried out. Fluorescence quantitative PCR detection, yeast two-hybrid test and optical confocal microscopy were used to investigate the effect of hIFITM3 on influenza virus replication, the interaction with hABHD16A and the expression of inflammation-related factors. In HEK293 cells, overexpression of hIFITM3 protein significantly inhibited the replication of influenza virus at 24h, 48h, and 72h; yeast two-hybrid experiment proved that IFITM3 interacts with ABHD16A; laser confocal microscopy observations showed that IFITM3 and ABHD16A co-localized in Cell membrane area; the expression level of inflammation-related factors in cells overexpressing hIFITM3 or hABHD16A was detected by fluorescence quantitative PCR, and the results showed that the mRNA levels of IL-1β, IL-6, IL-10, TNF-a and COX2 were significantly increased . But when IFITM3/ABHD16A was co-expressed, the mRNA expression levels of these cytokines were significantly reduced except for COX2. When influenza virus infected cells co-expressing IFITM3/ABHD16A, the expression level of inflammatory factors decreased compared with the control group, indicating that IFITM3 can play an important role in regulating inflammation balance. This study confirmed that hIFITM3 has an effect of inhibiting influenza virus replication. Furthermore, it was found that hIFITM3 interacts with hABHD16A, following which it can better inhibit the replication of influenza virus and the inflammatory response caused by the disease process.
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The V3 Loop of HIV-1 Env Determines Viral Susceptibility to IFITM3 Impairment of Viral Infectivity.
Journal of virology, 2017Co-Authors: Yimeng Wang, Shilei Ding, Qinghua Pan, Shanlu Liu, Andres Finzi, Zhen Wang, Chen LiangAbstract:Interferon-inducible transmembrane proteins (IFITMs) inhibit a broad spectrum of viruses, including HIV-1. IFITM proteins deter HIV-1 entry when expressed in target cells and also impair HIV-1 infectivity when expressed in virus producer cells. However, little is known about how viruses resist IFITM inhibition. In this study, we have investigated the susceptibilities of different primary isolates of HIV-1 to the inhibition of viral infectivity by IFITMs. Our results demonstrate that the infectivity of different HIV-1 primary isolates, including transmitted founder viruses, is diminished by IFITM3 to various levels, with strain AD8-1 exhibiting strong resistance. Further mutagenesis studies revealed that HIV-1 Env, and the V3 loop sequence in particular, determines the extent of inhibition of viral infectivity by IFITM3. IFITM3-sensitive Env proteins are also more susceptible to neutralization by soluble CD4 or the 17b antibody than are IFITM3-resistant Env proteins. Together, data from our study suggest that the propensity of HIV-1 Env to sample CD4-bound-like conformations modulates viral sensitivity to IFITM3 inhibition.IMPORTANCE Results of our study have revealed the key features of the HIV-1 envelope protein that are associated with viral resistance to the IFITM3 protein. IFITM proteins are important effectors in interferon-mediated antiviral defense. A variety of viruses are inhibited by IFITMs at the virus entry step. Although it is known that envelope proteins of several different viruses resist IFITM inhibition, the detailed mechanisms are not fully understood. Taking advantage of the fact that envelope proteins of different HIV-1 strains exhibit different degrees of resistance to IFITM3 and that these HIV-1 envelope proteins share the same domain structure and similar sequences, we performed mutagenesis studies and determined the key role of the V3 loop in this viral resistance phenotype. We were also able to associate viral resistance to IFITM3 inhibition with the susceptibility of HIV-1 to inhibition by soluble CD4 and the 17b antibody that recognizes CD4-binding-induced epitopes.
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Natural mutations in IFITM3 modulate post‐translational regulation and toggle antiviral specificity
EMBO Reports, 2016Co-Authors: Alex A. Compton, Chen Liang, Jacob S. Yount, Nicolas Roy, Françoise Porrot, Anne Billet, Nicoletta Casartelli, Olivier SchwartzAbstract:The interferon-induced transmembrane (IFITM) proteins protect host cells from diverse virus infections. IFITM proteins also incorporate into HIV-1 virions and inhibit virus fusion and cell-to-cell spread, with IFITM3 showing the greatest potency. Here, we report that amino-terminal mutants of IFITM3 preventing ubiquitination and endocytosis are more abundantly incorporated into virions and exhibit enhanced inhibition of HIV-1 fusion. An analysis of primate genomes revealed that IFITM3 is the most ancient antiviral family member of the IFITM locus and has undergone a repeated duplication in independent host lineages. Some IFITM3 genes in nonhuman primates, including those that arose following gene duplication, carry amino-terminal mutations that modify protein localization and function. This suggests that "runaway" IFITM3 variants could be selected for altered antiviral activity. Furthermore, we show that adaptations in IFITM3 result in a trade-off in antiviral specificity, as variants exhibiting enhanced activity against HIV-1 poorly restrict influenza A virus. Overall, we provide the first experimental evidence that diversification of IFITM3 genes may boost the antiviral coverage of host cells and provide selective functional advantages.
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natural mutations in IFITM3 modulate post translational regulation and toggle antiviral specificity
EMBO Reports, 2016Co-Authors: Alex A. Compton, Chen Liang, Olivier Schwartz, Jacob S. Yount, Françoise Porrot, Anne Billet, Nicoletta CasartelliAbstract:The interferon-induced transmembrane (IFITM) proteins protect host cells from diverse virus infections. IFITM proteins also incorporate into HIV-1 virions and inhibit virus fusion and cell-to-cell spread, with IFITM3 showing the greatest potency. Here, we report that amino-terminal mutants of IFITM3 preventing ubiquitination and endocytosis are more abundantly incorporated into virions and exhibit enhanced inhibition of HIV-1 fusion. An analysis of primate genomes revealed that IFITM3 is the most ancient antiviral family member of the IFITM locus and has undergone a repeated duplication in independent host lineages. Some IFITM3 genes in nonhuman primates, including those that arose following gene duplication, carry amino-terminal mutations that modify protein localization and function. This suggests that "runaway" IFITM3 variants could be selected for altered antiviral activity. Furthermore, we show that adaptations in IFITM3 result in a trade-off in antiviral specificity, as variants exhibiting enhanced activity against HIV-1 poorly restrict influenza A virus. Overall, we provide the first experimental evidence that diversification of IFITM3 genes may boost the antiviral coverage of host cells and provide selective functional advantages.
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the c terminal sequence of ifitm1 regulates its anti hiv 1 activity
PLOS ONE, 2015Co-Authors: Rui Jia, Shilei Ding, Qinghua Pan, Shanlu Liu, Wentao Qiao, Chen LiangAbstract:The interferon-inducible transmembrane (IFITM) proteins inhibit a wide range of viruses. We previously reported the inhibition of human immunodeficiency virus type 1 (HIV-1) strain BH10 by human IFITM1, 2 and 3. It is unknown whether other HIV-1 strains are similarly inhibited by IFITMs and whether there exists viral countermeasure to overcome IFITM inhibition. We report here that the HIV-1 NL4-3 strain (HIV-1NL4-3) is not restricted by IFITM1 and its viral envelope glycoprotein is partly responsible for this insensitivity. However, HIV-1NL4-3 is profoundly inhibited by an IFITM1 mutant, known as Δ(117–125), which is deleted of 9 amino acids at the C-terminus. In contrast to the wild type IFITM1, which does not affect HIV-1 entry, the Δ(117–125) mutant diminishes HIV-1NL4-3 entry by 3-fold. This inhibition correlates with the predominant localization of Δ(117–125) to the plasma membrane where HIV-1 entry occurs. In spite of strong conservation of IFITM1 among most species, mouse IFITM1 is 19 amino acids shorter at its C-terminus as compared to human IFITM1 and, like the human IFITM1 mutant Δ(117–125), mouse IFITM1 also inhibits HIV-1 entry. This is the first report illustrating the role of viral envelope protein in overcoming IFITM1 restriction. The results also demonstrate the importance of the C-terminal region of IFITM1 in modulating the antiviral function through controlling protein subcellular localization.
Michael Farzan - One of the best experts on this subject based on the ideXlab platform.
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The Interferon-Stimulated Gene IFITM3 Restricts Infection and Pathogenesis of Arthritogenic and Encephalitic Alphaviruses.
Journal of virology, 2016Co-Authors: Subhajit Poddar, Jennifer L. Hyde, Matthew J. Gorman, Michael Farzan, Michael S. DiamondAbstract:UNLABELLED Host cells respond to viral infections by producing type I interferon (IFN), which induces the expression of hundreds of interferon-stimulated genes (ISGs). Although ISGs mediate a protective state against many pathogens, the antiviral functions of the majority of these genes have not been identified. IFITM3 is a small transmembrane ISG that restricts a broad range of viruses, including orthomyxoviruses, flaviviruses, filoviruses, and coronaviruses. Here, we show that alphavirus infection is increased in IFITM3(-/-) and Ifitm locus deletion (Ifitm-del) fibroblasts and, reciprocally, reduced in fibroblasts transcomplemented with IFITM3. Mechanistic studies showed that IFITM3 did not affect viral binding or entry but inhibited pH-dependent fusion. In a murine model of chikungunya virus arthritis, IFITM3(-/-) mice sustained greater joint swelling in the ipsilateral ankle at days 3 and 7 postinfection, and this correlated with higher levels of proinflammatory cytokines and viral burden. Flow cytometric analysis suggested that IFITM3(-/-) macrophages from the spleen were infected at greater levels than observed in wild-type (WT) mice, results that were supported by experiments with IFITM3(-/-) bone marrow-derived macrophages. IFITM3(-/-) mice also were more susceptible than WT mice to lethal alphavirus infection with Venezuelan equine encephalitis virus, and this was associated with greater viral burden in multiple organs. Collectively, our data define an antiviral role for IFITM3 in restricting infection of multiple alphaviruses. IMPORTANCE The interferon-induced transmembrane protein 3 (IFITM3) inhibits infection of multiple families of viruses in cell culture. Compared to other viruses, much less is known about the antiviral effect of IFITM3 on alphaviruses. In this study, we characterized the antiviral activity of mouse IFITM3 against arthritogenic and encephalitic alphaviruses using cells and animals with a targeted gene deletion of IFITM3 as well as deficient cells transcomplemented with IFITM3. Based on extensive virological analysis, we demonstrate greater levels of alphavirus infection and disease pathogenesis when IFITM3 expression is absent. Our data establish an inhibitory role for IFITM3 in controlling infection of alphaviruses.
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The Interferon-Stimulated Gene IFITM3 Restricts West Nile Virus Infection and Pathogenesis
Journal of virology, 2016Co-Authors: Matthew J. Gorman, Subhajit Poddar, Michael Farzan, Michael S. DiamondAbstract:ABSTRACT The interferon-induced transmembrane protein (IFITM) family of proteins inhibit infection of several different enveloped viruses in cell culture by virtue of their ability to restrict entry and fusion from late endosomes. As few studies have evaluated the importance of IFITM3 in vivo in restricting viral pathogenesis, we investigated its significance as an antiviral gene against West Nile virus (WNV), an encephalitic flavivirus, in cells and mice. IFITM3 −/− mice were more vulnerable to lethal WNV infection, and this was associated with greater virus accumulation in peripheral organs and central nervous system tissues. As no difference in viral burden in the brain or spinal cord was observed after direct intracranial inoculation, IFITM3 likely functions as an antiviral protein in nonneuronal cells. Consistent with this, IFITM3 −/− fibroblasts but not dendritic cells resulted in higher yields of WNV in multistep growth analyses. Moreover, transcomplementation experiments showed that IFITM3 inhibited WNV infection independently of Ifitm1, Ifitm2, Ifitm5, and Ifitm6. Beyond a direct effect on viral infection in cells, analysis of the immune response in WNV-infected IFITM3 −/− mice showed decreases in the total number of B cells, CD4 + T cells, and antigen-specific CD8 + T cells. Finally, bone marrow chimera experiments demonstrated that IFITM3 functioned in both radioresistant and radiosensitive cells, as higher levels of WNV were observed in the brain only when IFITM3 was absent from both compartments. Our analyses suggest that IFITM3 restricts WNV pathogenesis likely through multiple mechanisms, including the direct control of infection in subsets of cells. IMPORTANCE As part of the mammalian host response to viral infections, hundreds of interferon-stimulated genes (ISGs) are induced. The inhibitory activity of individual ISGs varies depending on the specific cell type and viral pathogen. Among ISGs, the genes encoding interferon-induced transmembrane protein ( IFITM ) have been reported to inhibit multiple families of viruses in cell culture. However, few reports have evaluated the impact of IFITM genes on viral pathogenesis in vivo . In this study, we characterized the antiviral activity of IFITM3 against West Nile virus (WNV), an encephalitic flavivirus, using mice with a targeted gene deletion of IFITM3 . Based on extensive virological and immunological analyses, we determined that IFITM3 protects mice from WNV-induced mortality by restricting virus accumulation in peripheral organs and, subsequently, in central nervous system tissues. Our data suggest that IFITM3 restricts WNV pathogenesis by multiple mechanisms and functions in part by controlling infection in different cell types.
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the antiviral restriction factors ifitm1 2 and 3 do not inhibit infection of human papillomavirus cytomegalovirus and adenovirus
PLOS ONE, 2014Co-Authors: Cody J Warren, Ichueh Huang, Michael Farzan, Laura M Griffin, Alexander S Little, Dohun PyeonAbstract:Type I interferons (IFN-α and β) induce dynamic host defense mechanisms to inhibit viral infections. It has been recently recognized that the interferon-inducible transmembrane proteins (IFITM) 1, 2 and 3 can block entry of a broad spectrum of RNA viruses. However, no study to date has focused on the role of IFITM proteins in DNA virus restriction. Here, we demonstrate that IFN-α or -β treatment of keratinocytes substantially decreases human papillomavirus 16 (HPV16) infection while robustly inducing IFITM1, 2 and 3 expression. However, IFITM1, 2 and 3 overexpression did not inhibit HPV16 infection; rather, IFITM1 and IFITM3 modestly enhanced HPV16 infection in various cell types including primary keratinocytes. Moreover, IFITM1, 2 and 3 did not inhibit infection by two other DNA viruses, human cytomegalovirus (HCMV) and adenovirus type 5 (Ad5). Taken together, we reveal that the entry of several DNA viruses, including HPV, HCMV, and Ad5 is not affected by IFITM1, 2 and 3 expression. These results imply that HPV, and other DNA viruses, may bypass IFITM restriction during intracellular trafficking.
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interferon induced transmembrane protein 3 is a type ii transmembrane protein
Journal of Biological Chemistry, 2013Co-Authors: Charles C. Bailey, Ichueh Huang, Hema R Kondur, Michael FarzanAbstract:The interferon-induced transmembrane (IFITM) proteins are a family of small membrane proteins that inhibit the cellular entry of several genera of viruses. These proteins had been predicted to adopt a two-pass, type III transmembrane topology with an intracellular loop, two transmembrane helices (TM1 and TM2), and extracellular N and C termini. Recent work, however, supports an intramembrane topology for the helices with cytosolic orientation of both termini. Here we determined the topology of murine IFITM3. We found that the N terminus of IFITM3 could be stained by antibodies at the cell surface but that this conformation was cell type-dependent and represented a minority of the total plasma membrane pool. In contrast, the C terminus was readily accessible to antibodies at the cell surface and extracellular C termini comprised most or all of those present at the plasma membrane. The addition of a C-terminal KDEL endoplasmic reticulum retention motif to IFITM3 resulted in sequestration of IFITM3 in the ER, demonstrating an ER-luminal orientation of the C terminus. C-terminal, but not N-terminal, epitope tags were also degraded within lysosomes, consistent with their luminal orientation. Furthermore, epitope-tagged IFITM3 TM2 functioned as a signal anchor sequence when expressed in isolation. Collectively, our results demonstrate a type II transmembrane topology for IFITM3 and will provide insight into its interaction with potential targets and cofactors.
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IFITM3 Limits the Severity of Acute Influenza in Mice
PLoS pathogens, 2012Co-Authors: Charles C. Bailey, Ichueh Huang, Christina W. Kam, Michael FarzanAbstract:Interferon-induced transmembrane (IFITM) proteins are a family of viral restriction factors that inhibit the entry processes of several pathogenic viruses, including influenza A virus (IAV), in vitro. Here we report that IAV-infected knockout mice lacking the Ifitm locus on chromosome 7 exhibited accelerated disease progression, greater mortality, and higher pulmonary and systemic viral burdens as compared to wild type controls. We further observed that the phenotype of IFITM3-specific knockout mice was indistinguishable from that of mice lacking the entire Ifitm locus. IFITM3 was expressed by IAV target cells including alveolar type II pneumocytes and tracheal/bronchial respiratory epithelial cells. Robust IFITM3 expression was also observed in several tissues in the absence of infection. Among murine Ifitm promoters, only that of IFITM3 could be induced by type I and II interferons. IFITM3 could also be upregulated by the gp130 cytokines IL-6 and oncostatin M on cells expressing appropriate receptors, suggesting that multiple cytokine signals could contribute to IFITM3 expression in a cell or tissue-specific manner. Collectively, these findings establish a central role for IFITM3 in limiting acute influenza in vivo, and provide further insight into IFITM3 expression and regulation.
