HIV-1 Nef Changes the Proteome of T Cells Extracellular Vesicles Depleting IFITMs and Other Antiviral Factors.

Extracellular vesicles (EVs) are biomolecule carriers for intercellular communication in health and disease. Nef is a HIV virulence factor that is released from cells within EVs and is present in plasma EVs of HIV-1 infected individuals. We performed a quantitative proteomic analysis to fully characterize the Nef-induced changes in protein composition of T cell-derived EVs and identify novel host targets of HIV. Several proteins with well-described roles in infection or not previously associated with HIV pathogenesis were specifically modulated by Nef in EVs. Among the downregulated proteins are the interferon-induced transmembrane 1, 2, and 3 (IFITM1-3) proteins, broad-spectrum antiviral factors known to be cell-to-cell transferable by EVs. We demonstrate that Nef depletes IFITM1-3 from EVs by excluding these proteins from the plasma membrane and lipid rafts, which are sites of EVs biogenesis in T cells. Our data establish Nef as a modulator of EVs' global protein content and as an HIV factor that antagonizes IFITMs.

IFITM proteins: Understanding their diverse roles in viral infection, cancer, and immunity.

Interferon-induced transmembrane proteins (IFITMs) are broad spectrum antiviral factors that inhibit the entry of a wide range of clinically important pathogens including influenza A virus, HIV-1, and Dengue virus. IFITMs are thought to act primarily by antagonizing virus-cell membrane fusion in this regard. However, recent work on these proteins has uncovered novel post-entry viral restriction mechanisms. IFITMs are also increasingly thought to have a role regulating immune responses, including innate antiviral and inflammatory responses as well as adaptive T-cell and B-cell responses. Further, IFITMs may have pathological activities in cancer, wherein IFITM expression can be a marker of therapeutically resistant and aggressive disease courses. In this review, we summarize the respective literatures concerning these apparently diverse functions with a view to identifying common themes and potentially yielding a more unified understanding of IFITM biology.

Influenza Virus Host Restriction Factors: The ISGs and Non-ISGs.

Influenza virus has been one of the most prevalent and researched viruses globally. Consequently, there is ample information available about influenza virus lifecycle and pathogenesis. However, there is plenty yet to be known about the determinants of influenza virus pathogenesis and disease severity. Influenza virus exploits host factors to promote each step of its lifecycle. In turn, the host deploys antiviral or restriction factors that inhibit or restrict the influenza virus lifecycle at each of those steps. Two broad categories of host restriction factors can exist in virus-infected cells: (1) encoded by the interferon-stimulated genes (ISGs) and (2) encoded by the constitutively expressed genes that are not stimulated by interferons (non-ISGs). There are hundreds of ISGs known, and many, e.g., Mx, IFITMs, and TRIMs, have been characterized to restrict influenza virus infection at different stages of its lifecycle by (1) blocking viral entry or progeny release, (2) sequestering or degrading viral components and interfering with viral synthesis and assembly, or (3) bolstering host innate defenses. Also, many non-ISGs, e.g., cyclophilins, ncRNAs, and HDACs, have been identified and characterized to restrict influenza virus infection at different lifecycle stages by similar mechanisms. This review provides an overview of those ISGs and non-ISGs and how the influenza virus escapes the restriction imposed by them and aims to improve our understanding of the host restriction mechanisms of the influenza virus.

The Antiviral Activity of Interferon-Induced Transmembrane Proteins and Virus Evasion Strategies.

Interferons (IFNs) are antiviral cytokines that defend against viral infections by inducing the expression of interferon-stimulated genes (ISGs). Interferon-inducible transmembrane proteins (IFITMs) 1, 2, and 3 are crucial ISG products and members of the CD225 protein family. Compelling evidence shows that IFITMs restrict the infection of many unrelated viruses by inhibiting the virus-cell membrane fusion at the virus entry step via the modulation of lipid composition and membrane properties. Meanwhile, viruses can evade IFITMs' restrictions by either directly interacting with IFITMs via viral glycoproteins or by altering the native entry pathway. At the same time, cumulative evidence suggests context-dependent and multifaceted roles of IFITMs in modulating virus infections and cell signaling. Here, we review the diverse antiviral mechanisms of IFITMs, the viral antagonizing strategies, and the regulation of IFITM activity in host cells. The mechanisms behind the antiviral activity of IFITMs could aid the development of broad-spectrum antivirals and enhance preparedness for future pandemics.

Inhibition of Arenavirus Entry and Replication by the Cell-Intrinsic Restriction Factor ZMPSTE24 Is Enhanced by IFITM Antiviral Activity.

In the absence of effective vaccines and treatments, annual outbreaks of severe human haemorrhagic fever caused by arenaviruses, such as Lassa virus, continue to pose a significant human health threat. Understanding the balance of cellular factors that inhibit or promote arenavirus infection may have important implications for the development of effective antiviral strategies. Here, we identified the cell-intrinsic zinc transmembrane metalloprotease, ZMPSTE24, as a restriction factor against arenaviruses. Notably, CRISPR-Cas9-mediated knockout of ZMPSTE24 in human alveolar epithelial A549 cells increased arenavirus glycoprotein-mediated viral entry in pseudoparticle assays and live virus infection models. As a barrier to viral entry and replication, ZMPSTE24 may act as a downstream effector of interferon-induced transmembrane protein (IFITM) antiviral function; though through a yet poorly understood mechanism. Overexpression of IFITM1, IFITM2, and IFITM3 proteins did not restrict the entry of pseudoparticles carrying arenavirus envelope glycoproteins and live virus infection. Furthermore, gain-of-function studies revealed that IFITMs augment the antiviral activity of ZMPSTE24 against arenaviruses, suggesting a cooperative effect of viral restriction. We show that ZMPSTE24 and IFITMs affect the kinetics of cellular endocytosis, suggesting that perturbation of membrane structure and stability is likely the mechanism of ZMPSTE24-mediated restriction and cooperative ZMPSTE24-IFITM antiviral activity. Collectively, our findings define the role of ZMPSTE24 host restriction activity in the early stages of arenavirus infection. Moreover, we provide insight into the importance of cellular membrane integrity for productive fusion of arenaviruses and highlight a novel avenue for therapeutic development.

Pregnancy complications and Interferon-induced transmembrane proteins (IFITM): balancing antiviral immunity and placental development.

Pregnancy complications occur frequently and are particularly prevalent during the first trimester. They are caused by a multitude of factors, including karyotypic, genetic or environmental conditions, congenital infections and inflammation. The molecular mechanisms leading to placental complications under inflammatory conditions remain unclear. In this review, we discuss how uncontrolled inflammation, triggered by viral infections or other diseases can lead to placental complications. We first highlight the importance of syncytins, ancestral retroviral genes co-opted by mammals including humans, millions of years ago for the process of placenta formation. We then focus on recent advances elucidating how interferon-induced transmembrane (IFITM) proteins, antiviral proteins rendering cells refractory to viral infections, interfere with placental development.

Certaines grossesses s'accompagnent de complications et sont dites pathologiques, elles sont particulièrement prévalentes lors du premier trimestre. Celles-ci peuvent être causées par une multitude de facteurs, comme les anormalités caryotypiques, des facteurs génétiques et environnementaux, des infections congénitales et une sur-inflammation. Dans cette revue, nous examinons comment une inflammation incontrôlée, déclenchée par des infections virales ou d'autres maladies inflammatoires, peut entraîner des complications placentaires. Dans un premier temps, nous mettrons en évidence l'importance des syncytines, protéines d'enveloppe rétrovirales capturées par les mammifères, dont l'homme, dans la formation du placenta. Dans un deuxième temps, nous nous concentrons sur la manière dont des protéines cellulaires appelées « IFITM ¼ (interferon-induced transmembrane proteins), qui sont des protéines antivirales rendant les cellules réfractaires aux infections virales, interfèrent avec un mécanisme clé du développement placentaire.

Interferon-Induced Transmembrane Proteins Inhibit Infection by the Kaposi's Sarcoma-Associated Herpesvirus and the Related Rhesus Monkey Rhadinovirus in a Cell-Specific Manner.

The interferon-induced transmembrane proteins (IFITMs) are broad-spectrum antiviral proteins that inhibit the entry of enveloped viruses. We analyzed the effect of IFITMs on the gamma-2 herpesviruses Kaposi's sarcoma-associated herpesvirus (KSHV) and the closely related rhesus monkey rhadinovirus (RRV). We used CRISPR/Cas9-mediated gene knockout to generate A549 cells, human foreskin fibroblasts (HFF), and human umbilical vein endothelial cells (HUVEC) with combined IFITM1/2/3 knockout and identified IFITMs as cell-dependent inhibitors of KSHV and RRV infection in A549 cells and HFF but not HUVEC. IFITM overexpression revealed IFITM1 as the relevant IFITM that inhibits KSHV and RRV infection. Fluorescent KSHV particles did not pronouncedly colocalize with IFITM-positive compartments. However, we found that KSHV and RRV glycoprotein-mediated cell-cell fusion is enhanced upon IFITM1/2/3 knockout. Taken together, we identified IFITM1 as a cell-dependent restriction factor of KSHV and RRV that acts at the level of membrane fusion. Of note, our results indicate that recombinant IFITM overexpression may lead to results that are not representative for the situation at endogenous levels. Strikingly, we observed that the endotheliotropic KSHV circumvents IFITM-mediated restriction in HUVEC despite high IFITM expression, while influenza A virus (IAV) glycoprotein-driven entry into HUVEC is potently restricted by IFITMs even in the absence of interferon. Mechanistically, we found that KSHV colocalizes less with IFITM1 and IFITM2 in HUVEC than in A549 cells immediately after attachment, potentially contributing to the observed difference in restriction. IMPORTANCE IFITM proteins are the first line of defense against infection by many pathogens and may also have therapeutic importance, as they, among other effectors, mediate the antiviral effect of interferons. Neither their function against herpesviruses nor their mechanism of action is well understood. We report here that in some cells but not in, for example, primary umbilical vein endothelial cells, IFITM1 restricts KSHV and RRV and that, mechanistically, this is likely effected by reducing the fusogenicity of the cell membrane. Further, we demonstrate potent inhibition of IAV glycoprotein-driven infection of cells of extrapulmonary origin by high constitutive IFITM expression.

S-palmitoylation of swine interferon-inducible transmembrane protein is essential for its anti-JEV activity.

Japanese encephalitis virus (JEV) is an infectious pathogen spreading in a wide range of vertebrate species. Pigs are amplifying hosts of JEV and thought to be maintained in nature predominantly by avian-mosquito cycles. In the innate immune system, interferon-inducible transmembrane protein (IFITM) is a small transmembrane protein family and has been identified as the first line of defense against a broad range of RNA virus invasion. In this paper, we found that swine IFITM (sIFITM) could restrict the replication of both JEV vaccine strain and wild strain NJ-2008. The cysteine S-palmitoylation modification of sIFITM plays important roles in their anti-JEV effects and intracellular distributions. Our findings show the anti-JEV activities of swine interferon-inducible transmembrane proteins and broaden the antiviral spectrum of IFITM protein family. The preliminary exploration of S-palmitoylation modification of sIFITM may contribute to understanding of the antiviral molecular mechanism of sIFITM.

Porcine IFITM1 is a host restriction factor that inhibits pseudorabies virus infection.

Interferon-inducible transmembrane proteins (IFITMs) restrict infection by several viruses, such as influenza A virus, West Nile virus and dengue virus. It has not been determined whether porcine IFITMs (pIFITMs) inhibit infection by pseudorabies virus (PRV), an enveloped, double-stranded DNA virus, which is the etiological agent of Aujeszky's disease in pigs. Here, we report that PRV infection elicited pIFITM1 expression in PK15 porcine kidney epithelial cells and 3D4/21 alveolar macrophages. pIFITM2 and pIFITM3 expression was only elevated in PK15 cells during PRV infection. Depletion of pIFITM1 using RNA interference, either in PK15 or in 3D4/21 cells, enhanced PRV infection while overexpression of pIFITM1 had the opposite effect. Knockdown of pIFITM2 and pIFITM3 did not influence PRV infection, suggesting that pIFITM2 and pIFITM3 are independent of PRV infection. PRV-induced pIFITM1 expression was dependent on the cGAS/STING/TBK1/IRF3 innate immune pathway and interferon-alpha receptor-1, suggesting that pIFITM1 is up-regulated by the type I interferon signaling pathway. The anti-PRV role of pIFITM1 was inhibited upon PRV entry. Our data demonstrate that pIFITM1 is a host restriction factor that inhibits PRV entry that may shed light on a strategy for prevention of PRV infection.

NP and NS1 proteins of H5N1 virus significantly upregulated IFITM1, IFITM2, and IFITM3 in A549 cells.

BACKGROUND: Avian influence virus H5N1 causes serious public health concern with significant morbidity and mortality from poultry to humans. Interferon-induced transmembrane (IFITM) proteins usually protect cells from many virus infections by viral entry and replication. OBJECTIVES: The purpose of this study was to investigate whether H5N1 viral proteins involved in regulation IFITM1, IFITM2, and IFITM3 following H5N1 infection. METHODS: NS1, M1, NP, PB2, HA and NA genes of H5N1 virus were generated by PCR and cloned into pcDNA3.1/myc-His (+) A vector for genes over-expression experiments. Gene expression levels was performed using Real-time PCR. RESULTS: Research displayed that NS1, M1, NP, and PB2 proteins of H5N1 virus increased IFITM1, IFITM2, and IFITM3 expression in A549 cells, only IFITM1 was upregulated by M1 in HEK293T cells. However, our study did not find that HA and NA of H5N1 virus affected IFITM genes family or interferon genes expression. CONCLUSION: Taken together, our data suggested that IFITM1, IFITM2, and IFITM3 might be directly upregulated via NS1, M1, NP, and PB2 proteins during H5N1 avian influenza virus infection. This study provided new insights into the influence of NS1 and NP proteins on regulation of IFITM1, IFITM2, and IFITM3 expression following H5N1 infection.

The expression profile of IFITM family gene in rats.

The interferon-inducible transmembrane proteins (IFITMs) are a family of small transmembrane proteins belonging to the interferon (IFN)-stimulated gene (ISG) superfamily and strongly induced by IFNs. In this paper, we studied the expression profile of IFITMs in 32 organ tissues. The IFITM mRNA expression profile showed that IFITM1, IFITM2 and IFITM3 were expressed in each tissue, especially, in spermatophore, spermaduct, testicle and epididymis. The expression of IFITM1, IFITM2 and IFITM3 showed a trend from high to low. Except for IFITM3 and IFITM6, the others IFITMs were highly expressed in the bone marrow, and the expression level of them was higher in the tibia than that in other parts of the long bones. In liver, the relative expression of IFITM1 and IFITM3 was higher than that of other members. The expression level of IFITM5 was the highest in bone marrow, successively in pancreas, and it was low in skin, smooth muscle and fat. Interestingly, the expression profile of IFITM2 and IFITM7 in tissues was similar to IFITM5. The expression of IFITM2, IFITM5 and IFITM10 were higher in smooth muscle than that in skeletal muscle. IFITM2, IFITM5, IFITM7 and IFITM10 were both highly expressed in esophagus and trachea. In addition, the expression of IFITM6 in eyes was high, and also in pancreas, gallbladder and bone. In the present study, we systematically analyzed the mRNA expression profile of IFITMs in 32 organ tissues, providing the foundation for the study of the function of the IFITMs.