Capsid-dependent lentiviral restrictions.

Host antiviral proteins inhibit primate lentiviruses and other retroviruses by targeting many features of the viral life cycle. The lentiviral capsid protein and the assembled viral core are known to be inhibited through multiple, directly acting antiviral proteins. Several phenotypes, including those known as Lv1 through Lv5, have been described as cell type-specific blocks to infection against some but not all primate lentiviruses. Here we review important features of known capsid-targeting blocks to infection together with several blocks to infection for which the genes responsible for the inhibition still remain to be identified. We outline the features of these blocks as well as how current methodologies are now well suited to find these antiviral genes and solve these long-standing mysteries in the HIV and retrovirology fields.

ATF5-regulated Mitochondrial Unfolded Protein Response Attenuates Neuronal Damage in Epileptic Rat by Reducing Endoplasmic Reticulum Stress Through Mitochondrial ROS.

Endoplasmic reticulum (ER) dysfunction caused by excessive ER stress is a crucial mechanism underlying seizures-induced neuronal injury. Studies have shown that mitochondrial reactive oxygen species (ROS) are closely related to ER stress, and our previous study showed that activating transcription factor 5 (ATF5)-regulated mitochondrial unfolded protein response (mtUPR) modulated mitochondrial ROS generation in a hippocampal neuronal culture model of seizures. However, the effects of ATF5-regulated mtUPR on ER stress and the underlying mechanisms remain uncertain in epilepsy. In this study, ATF5 upregulation by lentivirus infection attenuated seizures-induced neuronal damage and apoptosis in a rat model of pilocarpine-induced epilepsy, whereas ATF5 downregulation by lentivirus infection had the opposite effects. ATF5 upregulation potentiated mtUPR by increasing the expression of mitochondrial chaperone heat shock protein 60 (HSP60) and caseinolytic protease proteolytic subunit (ClpP) and reducing mitochondrial ROS generation in pilocarpine-induced seizures in rats. Additionally, upregulation of ATF5 reduced the expression of glucose-regulated protein 78 (GRP78), protein kinase RNA-like endoplasmic reticulum kinase (PERK), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP), suggesting suppression of ER stress; Moreover, ATF5 upregulation attenuated apoptosis-related proteins such as B-cell lymphoma-2 (BCL2) downregulation, BCL2-associated X (BAX) and cleaved-caspase-3 upregulation. However, ATF5 downregulation exerted the opposite effects. Furthermore, pretreatment with the mitochondria-targeted antioxidant mito-TEMPO attenuated the harmful effects of ATF5 downregulation on ER stress and neuronal apoptosis by reducing mitochondrial ROS generation. Overall, our study suggested that ATF5-regulated mtUPR exerted neuroprotective effects against pilocarpine-induced seizures in rats and the underlying mechanisms might involve mitochondrial ROS-mediated ER stress.

A negative feedback loop centered on SMAD3 expression in transforming growth factor ß1-induced corneal myofibroblast differentiation.

SMAD3 downregulation is documented in transforming growth factor ß1 (TGF-ß1)-induced corneal fibroblasts differentiation to myofibroblasts ("fibroTOmyoDiff") or corneal wound healing. However, the exact regulatory mechanism of TGF-ß1/SMAD3 pathway in this context remains unclear. Here, we investigated the role and related mechanism of SMAD3 down-regulation in TGF-ß1-induced human corneal fibroTOmyoDiff. By detecting expression changes of SMAD family during this process, we demonstrated that SMAD3 protein expression was dramatically decreased in the process and the decrease occurred mainly in SMAD3 gene transcription. Furthermore, SMAD3 overexpression using lentivirus infection and knockdown using sgRNA lentivirus infection or siRNAs revealed that SMAD3 overexpression enhanced TGF-ß1-induced corneal fibroTOmyoDiff and vice versa. In addition, specific siRNAs and inhibitors targeting particular signaling pathway were used to figure out the intracellular signaling pathway regulating SMAD3, and the result showed that the decease of SMAD3 induced by TGF-ß1 stimulation in human corneal fibroblasts (HCFs) was strikingly prevented by SMAD4 knockdown or p38 signaling inhibitor SB203580 treatment. Collectively, these results demonstrate that, in TGF-ß1 induced corneal fibroTOmyoDiff, down-regulation of SMAD3 expression regulated by SMAD4 and p38 signaling pathways forms a negative feedback loop of TGFß signaling to avoid excessive activation of the signaling, which suggest that SMAD3 may be a key target for corneal fibrosis treatment.

Effect of lentivirus-mediated growth and differentiation factor-5 transfection on differentiation of rabbit nucleus pulposus mesenchymal stem cells.

BACKGROUND: Intervertebral disc degeneration (IDD) is a natural progression of age-related processes. Associated with IDD, degenerative disc disease (DDD) is a pathologic condition implicated as a major cause of chronic lower back pain, which can have a severe impact on the quality of life of patients. As degeneration progression is associated with elevated levels of inflammatory cytokines, enhanced aggrecan and collagen degradation, and changes in the disc cell phenotype. The purpose of this study was to investigate the biological and cytological characteristics of rabbit nucleus pulposus mesenchymal stem cells (NPMSCs)-a key factor in IDD-and to determine the effect of the growth and differentiation factor-5 (GDF5) on the differentiation of rabbit NPMSCs transduced with a lentivirus vector. METHODS: An in vitro culture model of rabbit NPMSCs was established and NPMSCs were identified by flow cytometry (FCM) and quantitative real-time PCR (qRT-PCR). Subsequently, NPMSCs were randomly divided into three groups: a transfection group (the lentiviral vector carrying GDF5 gene used to transfect NPMSCs); a control virus group (the NPMSCs transfected with an ordinary lentiviral vector); and a normal group (the NPMSCs alone). FCM, qRT-PCR, and western blot (WB) were used to detect the changes in NPMSCs. RESULTS: The GDF5-transfected NPMSCs displayed an elongated shape, with decreased cell density, and significantly increased GDF5 positivity rate in the transfected group compared to the other two groups (P < 0.01). The mRNA levels of Krt8, Krt18, and Krt19 in the transfected group were significantly higher in comparison with the other two groups (P < 0.01), and the WB results were consistent with that of qRT-PCR. CONCLUSIONS: GDF5 could induce the differentiation of NPMSCs. The lentiviral vector carrying the GDF5 gene could be integrated into the chromosome genome of NPMSCs and promoted differentiation of NPMSCs into nucleus pulposus cells. Our findings advance the development of feasible and effective therapies for IDD.

Persistent lentivirus infection induces early myeloid suppressor cells expansion to subvert protective memory CD8 T cell response✰,✰✰.

BACKGROUND: Memory CD8+T cell responses play an essential role in protection against persistent infection. However, HIV-1 evades vaccine-induced memory CD8+T cell response by mechanisms that are not fully understood. METHODS: We analyzed the temporal dynamics of CD8+T cell recall activity and function during EcoHIV infection in a potent PD1-based vaccine immunized immunocompetent mice. FINDINGS: Upon intraperitoneal EcoHIV infection, high levels of HIV-1 GAG-specific CD8+T lymphocytes recall response reduced EcoHIV-infected cells significantly. However, this protective effect diminished quickly after seven days, followed by a rapid reduction of GAG-specific CD8+T cell number and activity, and viral persistence. Mechanistically, EcoHIV activated dendritic cells (DCs) and myeloid cells. Myeloid cells were infected and rapidly expanded, exhibiting elevated PD-L1/-L2 expression and T cell suppressive function before day 7, and were resistant to CD8+T cell-mediated apoptosis. Depletion of myeloid-derived suppressor cells (MDSCs) reduced EcoHIV infection and boosted T cell responses. INTERPRETATION: This study provides an overview of the temporal interplay of persistent virus, DCs, MDSCs and antigen-specific CD8+T cells during acute infection. We identify MDSCs as critical gatekeepers that restrain antiviral T cell memory responses, and highlight MDSCs as an important target for developing effective vaccines against chronic human infections. FUNDING: Hong Kong Research Grant Council (T11-709/18-N, HKU5/CRF/13G), General Research Fund (17122915 and 17114114), Hong Kong Health and Medical Research Fund (11100752, 14130582, 16150662), Grant RGC-ANR A-HKU709/14, the San-Ming Project of Medicine (SZSM201512029), University Development Fund of the University of Hong Kong and Li Ka Shing Faculty of Medicine Matching Fund to HKU AIDS Institute.

Structural Insights into APOBEC3-Mediated Lentiviral Restriction.

Mammals have developed clever adaptive and innate immune defense mechanisms to protect against invading bacterial and viral pathogens. Human innate immunity is continuously evolving to expand the repertoire of restriction factors and one such family of intrinsic restriction factors is the APOBEC3 (A3) family of cytidine deaminases. The coordinated expression of seven members of the A3 family of cytidine deaminases provides intrinsic immunity against numerous foreign infectious agents and protects the host from exogenous retroviruses and endogenous retroelements. Four members of the A3 proteins-A3G, A3F, A3H, and A3D-restrict HIV-1 in the absence of virion infectivity factor (Vif); their incorporation into progeny virions is a prerequisite for cytidine deaminase-dependent and -independent activities that inhibit viral replication in the host target cell. HIV-1 encodes Vif, an accessory protein that antagonizes A3 proteins by targeting them for polyubiquitination and subsequent proteasomal degradation in the virus producing cells. In this review, we summarize our current understanding of the role of human A3 proteins as barriers against HIV-1 infection, how Vif overcomes their antiviral activity, and highlight recent structural and functional insights into A3-mediated restriction of lentiviruses.

The expression of cytokines in the milk somatic cells, blood leukocytes and serum of goats infected with small ruminant lentivirus.

BACKGROUND: The present study aimed to determine the expression of cytokines, which is associated with the immunological response of dairy goats against small ruminant lentivirus (SRLV). The study was conducted on 26 dairy goats in their second to sixth lactation, which were divided by breed and parity into two groups: SRLV naturally infected (N = 13) and non-infected (N = 13) animals. All goats in the study were asymptomatic. The milk and blood samples, which served as studied material were taken on days 7, 30, 120 and 240 of the lactation. The gene and protein expression of several cytokines was studied using Real-Time PCR and ELISA methods. RESULTS: INF-ß and INF-γ expression was down-regulated in the milk somatic cells (MSC) of SRLV-infected goats. However, an increased concentration of INF-ß was observed in the MSC in SRLV-infected goats, while INF-γ expression was not observed in both SRLV-infected and non-infected animals The SRLV-infected goats also displayed decreased expression of IL-1α, IL-1ß, IL-6 and INF-γ genes in the blood leukocytes,with IL-1α, IL-1ß and IL-6 protein levels also being decreased in the sera. TNF-α was the only gene that demonstrated increased expression in both the MSC and the blood of infected animals; however, no such overexpression was observed at the protein level. CONCLUSIONS: SRLV probably influences the immune system of infected animals by deregulating of the expression of cytokines. Further, epigenetic studies may clarify the mechanisms by which SRLV regulates the gene and protein expression of the host.

Small ruminant lentivirus infection influences expression of acute phase proteins and cathelicidin genes in milk somatic cells and peripheral blood leukocytes of dairy goats.

The aim of the study was to analyze acute phase protein and cathelicidin gene responses to small ruminant lentivirus (SRLV) infection in goats. In uninfected goats, we found higher Cp and lower Fbγ mRNA levels in blood leucocytes (BL) than in milk somatic cells (MSC), as well as lower SAA, Hp, and CRP and higher Cp and AGP concentrations in blood serum than in milk. In SRLV-infected goats, we found higher Fbγ and MAP28 and lower Cp expression in MSC than in BL, and higher SAA, Hp, Fb, and MAP28 and lower AGP concentrations in milk than in blood serum. Higher SAA and Hp expressions in BL and Hp expression in MSC were found in SRLV-infected goats. In SRLV-infected goats, we observed a higher concentration of SAA in blood serum, while in milk, lower SAA, Cp, and MAP28 and higher MAP34 concentrations were observed. The expression profiles of the studied genes differed between BL/serum and MSC/milk. The elevated SAA concentration in blood serum was accompanied by a decreased concentration of SAA and Cp in the milk of infected goats. No differences in the expression of the other studied genes may mean that the SRLV has the ability to evade the immune system, continuing to replicate. The elevated concentration of SAA in blood serum may promote viral multiplication. This higher concentration of SAA in blood serum and simultaneous reduced concentration of SAA and Cp in milk may be additive indicators of this infection.

Robust Enhancement of Lentivirus Production by Promoter Activation.

Lentiviral vectors are a valuable tool to deliver exogenous genes for stable expression in cells. While much progress has been made in processing lentiviral vector-containing culture medium, it remains to be explored how the production of lentiviral vector from producer cells can be increased. We initially found that co-expression of the SPRY domain-containing SOCS box protein 1 (SPSB1) promotes the production of human immunodeficiency virus type 1 (HIV-1) and lentiviral vector with increased expression of the Gag and envelope proteins and activation of the HIV-1 LTR and CMV promoter. The presence of AP-1, NF-κB and CREB/ATF recognition sites in these promoters prompted us to utilize human T-lymphotropic virus type 1 (HTLV-1) Tax for lentiviral vector production because Tax activates all these transcription factors. Co-expression of a small amount of Tax markedly increased both the expression of viral structural proteins in producer cells and release of lentiviral vector particles, resulting in a more than 10-fold enhancement of transduction efficiency. Of note, the Tax protein was not detected in the lentiviral vector particles concentrated by ultracentrifugation, supporting the safety of this preparation. Collectively, these results indicate that promoter activation in producer cells represents a promising approach to preparing high-titer lentiviral vectors.

A CRISPR screen for factors regulating SAMHD1 degradation identifies IFITMs as potent inhibitors of lentiviral particle delivery.

The InterFeron Induced TransMembrane (IFITM) proteins are interferon stimulated genes that restrict many viruses, including HIV-1. SAMHD1 is another restriction factor blocking replication of HIV-1 and other viruses. Some lentiviruses evolved Vpx/Vpr proteins to degrade SAMHD1. However, this viral antagonism can be perturbed by host mechanisms: a recent study showed that in interferon (IFN) treated THP1 cells, Vpx is unable to degrade SAMHD1. In the present work, we designed an Interferon Stimulated Genes (ISGs)-targeted CRISPR knockout screen in order to identify ISGs regulating this phenotype. We found that IFITM proteins contribute to the IFNα-mediated protection of SAMHD1 by blocking VSV-G-mediated entry of the lentiviral particles delivering Vpx. Consistent with this, IFNα treatment and IFITM expression had no effect when the A-MLV envelope was used for pseudotyping. Using an assay measuring viral entry, we show that IFNα and IFITMs directly block the delivery of Vpx into cells by inhibiting VSV-G viral fusion. Strikingly, the VSV-G envelope was significantly more sensitive to this IFNα entry block and to IFITMs than HIV-1's natural envelope. This highlights important differences between VSV-G pseudotyped and wild-type HIV-1, in particular relative to the pathways they use for viral entry, suggesting that HIV-1 may have evolved to escape restriction factors blocking entry.

B23/nucleophosmin interacts with bovine immunodeficiency virus Rev protein and facilitates viral replication.

The bovine immunodeficiency virus (BIV) Rev shuttling protein contains nuclear/nucleolar localization signals and nuclear import/export mechanisms that are novel among lentivirus Rev proteins. Several viral proteins localize to the nucleolus, which may play a role in processes that are essential to the outcome of viral replication. Although BIV Rev localizes to the nucleoli of transfected/infected cells and colocalizes with one of its major proteins, nucleophosmin (NPM1, also known as B23), the role of the nucleolus and B23 in BIV replication remains to be determined. Here, we demonstrate for the first time that BIV Rev interacts with nucleolar phosphoprotein B23 in cells. Using small interfering RNA (siRNA) technology, we show that depletion of B23 expression inhibits virus production by BIV-infected cells, indicating that B23 plays an important role in BIV replication. The interaction between Rev and B23 may represent a potential new target for the development of antiviral drugs against lentiviruses.

The Impact of the CD9 Tetraspanin on Lentivirus Infectivity and Exosome Secretion.

Efficient transduction tools are a hallmark for both research and therapy development. Here, we introduce new insights into the generation of lentiviral vectors with improved performance by utilizing producer cells with increased production rates of extracellular vesicles through CD9 overexpression. Most human cells secrete small vesicles from their surface (microvesicles) or intraluminal endosome-derived membranes (exosomes). In particular, enhanced levels of the tetraspanin CD9 result in significantly increased numbers of extracellular vesicles with exosome-like features that were secreted from four different human cell lines. Intriguingly, exosomes and their biogenesis route display similarities to lentivirus and we examined the impact of CD9 expression on release and infectivity of recombinant lentiviral vectors. Although the titers of released viral particles were not increased upon production in high CD9 cells, we observed improved performance in terms of both speed and efficiency of lentiviral gene delivery into numerous human cell lines, including HEK293, HeLa, SH-SY5Y, as well as B and T lymphocytes. Here, we demonstrate that enhanced CD9 enables lentiviral transduction in the absence of any pseudotyping viral glycoprotein or fusogenic molecule. Our findings indicate an important role of CD9 for lentiviral vector and exosome biogenesis and point out a remarkable function of this tetraspanin in membrane fusion, viral infectivity, and exosome-mediated horizontal information transfer.

Non-canonical function of Tat in regulating host microtubule dynamics: Implications for the pathogenesis of lentiviral infections.

Lentiviruses are a class of genetically unique retroviruses that share similar features, despite their wide variety of host species. Transactivator of transcription (Tat) proteins of lentiviruses are critical for the regulation of viral transcription and replication. Recent studies demonstrate that in addition to mediating transactivation, Tat binds to the microtubule cytoskeleton of the host cell and interferes with microtubule dynamics, ultimately triggering apoptosis. This non-canonical function of Tat appears to be critical for the pathogenesis of lentiviral diseases, such as acquired immunodeficiency syndrome. Here, we compare the structure and activity of Tat proteins from three different types of lentiviruses, focusing on the roles of these proteins in the alteration of host microtubule dynamics and induction of apoptosis. We propose that further investigation of the Tat-microtubule interaction will provide important insight into the process of lentiviral pathogenesis and elucidate new avenues for the development of antiviral therapies.

Insulin Treatment Prevents Neuroinflammation and Neuronal Injury with Restored Neurobehavioral Function in Models of HIV/AIDS Neurodegeneration.

HIV-1 infection of the brain causes the neurodegenerative syndrome HIV-associated neurocognitive disorders (HAND), for which there is no specific treatment. Herein, we investigated the actions of insulin using ex vivo and in vivo models of HAND. Increased neuroinflammatory gene expression was observed in brains from patients with HIV/AIDS. The insulin receptor was detected on both neurons and glia, but its expression was unaffected by HIV-1 infection. Insulin treatment of HIV-infected primary human microglia suppressed supernatant HIV-1 p24 levels, reduced CXCL10 and IL-6 transcript levels, and induced peroxisome proliferator-activated receptor gamma (PPAR-γ) expression. Insulin treatment of primary human neurons prevented HIV-1 Vpr-mediated cell process retraction and death. In feline immunodeficiency virus (FIV) infected cats, daily intranasal insulin treatment (20.0 IU/200 µl for 6 weeks) reduced CXCL10, IL-6, and FIV RNA detection in brain, although PPAR-γ in glia was increased compared with PBS-treated FIV+ control animals. These molecular changes were accompanied by diminished glial activation in cerebral cortex and white matter of insulin-treated FIV+ animals, with associated preservation of cortical neurons. Neuronal counts in parietal cortex, striatum, and hippocampus were higher in the FIV+/insulin-treated group compared with the FIV+/PBS-treated group. Moreover, intranasal insulin treatment improved neurobehavioral performance, including both memory and motor functions, in FIV+ animals. Therefore, insulin exerted ex vivo and in vivo antiviral, anti-inflammatory, and neuroprotective effects in models of HAND, representing a new therapeutic option for patients with inflammatory or infectious neurodegenerative disorders including HAND. SIGNIFICANCE STATEMENT: HIV-associated neurocognitive disorders (HAND) represent a spectrum disorder of neurocognitive dysfunctions resulting from HIV-1 infection. Although the exact mechanisms causing HAND are unknown, productive HIV-1 infection in the brain with associated neuroinflammation is a potential pathogenic mechanism resulting in neuronal damage and death. We report that, in HIV-infected microglia cultures, insulin treatment led to reduced viral replication and inflammatory gene expression. In addition, intranasal insulin treatment of experimentally feline immunodeficiency virus-infected animals resulted in improved motor and memory performances. We show that insulin restored expression of the nuclear receptor peroxisome proliferator-activated receptor gamma (PPAR-γ), which is suppressed by HIV-1 replication. Our findings indicate a unique function for insulin in improving neurological outcomes in lentiviral infections, implicating insulin as a therapeutic intervention for HAND.

Structural Basis of Clade-specific Engagement of SAMHD1 (Sterile α Motif and Histidine/Aspartate-containing Protein 1) Restriction Factors by Lentiviral Viral Protein X (Vpx) Virulence Factors.

Sterile α motif (SAM) and histidine/aspartate (HD)-containing protein 1 (SAMHD1) restricts human/simian immunodeficiency virus infection in certain cell types and is counteracted by the virulence factor Vpx. Current evidence indicates that Vpx recruits SAMHD1 to the Cullin4-Ring Finger E3 ubiquitin ligase (CRL4) by facilitating an interaction between SAMHD1 and the substrate receptor DDB1- and Cullin4-associated factor 1 (DCAF1), thereby targeting SAMHD1 for proteasome-dependent down-regulation. Host-pathogen co-evolution and positive selection at the interfaces of host-pathogen complexes are associated with sequence divergence and varying functional consequences. Two alternative interaction interfaces are used by SAMHD1 and Vpx: the SAMHD1 N-terminal tail and the adjacent SAM domain or the C-terminal tail proceeding the HD domain are targeted by different Vpx variants in a unique fashion. In contrast, the C-terminal WD40 domain of DCAF1 interfaces similarly with the two above complexes. Comprehensive biochemical and structural biology approaches permitted us to delineate details of clade-specific recognition of SAMHD1 by lentiviral Vpx proteins. We show that not only the SAM domain but also the N-terminal tail engages in the DCAF1-Vpx interaction. Furthermore, we show that changing the single Ser-52 in human SAMHD1 to Phe, the residue found in SAMHD1 of Red-capped monkey and Mandrill, allows it to be recognized by Vpx proteins of simian viruses infecting those primate species, which normally does not target wild type human SAMHD1 for degradation.

Effects of lentiviral infection of mesenchymal stem cells on the expression of octamer transcription factor 4.

The present study aimed to investigate the effects of lentiviral infection of human umbilical cord mesenchymal stem cells (hUCMSCs) on the expression of octamer transcription factor 4 (Oct4). hUCMSCs were infected with lentivirus carrying the green fluorescent protein gene (GFP) at different multiplicities of infection (MOI), and the optimal MOI was determined by flow cytometry; the proliferation of non­infected and GFP-carrying lentivirus­infected hUCMSCs was evaluated by the MTT assay; and the expression of the Oct4 gene was measured by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunofluorescence staining in hUCMSCs cultured in vitro for eight weeks. Positive GFP staining of hUCMSCs was estimated at >75% at 48 h following infection with the GFP-carrying lentivirus (MOI=20); no effect on hUCMSC proliferation was detected by the MTT assay following the infection; immunofluorescence analysis detected positive Oct4 expression in the cell nuclei at two and eight weeks of culture, while the relative expression of Oct4 assessed by qRT-PCR was 0.9075±0.0124. The GFP gene carried by the lentivirus was successfully expressed in hUCMSCs and had no significant effect on Oct4 expression, which lays a solid foundation for future studies investigating gene functions via the use of exogenous markers.

Intestinal epithelial barrier disruption through altered mucosal microRNA expression in human immunodeficiency virus and simian immunodeficiency virus infections.

UNLABELLED: Epithelial barrier dysfunction during human immunodeficiency virus (HIV) infection has largely been attributed to the rapid and severe depletion of CD4(+) T cells in the gastrointestinal (GI) tract. Although it is known that changes in mucosal gene expression contribute to intestinal enteropathy, the role of small noncoding RNAs, specifically microRNA (miRNA), has not been investigated. Using the simian immunodeficiency virus (SIV)-infected nonhuman primate model of HIV pathogenesis, we investigated the effect of viral infection on miRNA expression in intestinal mucosa. SIV infection led to a striking decrease in the expression of mucosal miRNA compared to that in uninfected controls. This decrease coincided with an increase in 5'-3'-exoribonuclease 2 protein and alterations in DICER1 and Argonaute 2 expression. Targets of depleted miRNA belonged to molecular pathways involved in epithelial proliferation, differentiation, and immune response. Decreased expression of several miRNA involved in maintaining epithelial homeostasis in the gut was localized to the proliferative crypt region of the intestinal epithelium. Our findings suggest that SIV-induced decreased expression of miRNA involved in epithelial homeostasis, disrupted expression of miRNA biogenesis machinery, and increased expression of XRN2 are involved in the development of epithelial barrier dysfunction and gastroenteropathy. IMPORTANCE: MicroRNA (miRNA) regulate the development and function of intestinal epithelial cells, and many viruses disrupt normal host miRNA expression. In this study, we demonstrate that SIV and HIV disrupt expression of miRNA in the small intestine during infection. The depletion of several key miRNA is localized to the proliferative crypt region of the gut epithelium. These miRNA are known to control expression of genes involved in inflammation, cell death, and epithelial maturation. Our data indicate that this disruption might be caused by altered expression of miRNA biogenesis machinery during infection. These findings suggest that the disruption of miRNA in the small intestine likely plays a role in intestinal enteropathy during HIV infection.

Feline immunodeficiency virus envelope glycoproteins antagonize tetherin through a distinctive mechanism that requires virion incorporation.

UNLABELLED: BST2/tetherin inhibits the release of enveloped viruses from cells. Primate lentiviruses have evolved specific antagonists (Vpu, Nef, and Env). Here we characterized tetherin proteins of species representing both branches of the order Carnivora. Comparison of tiger and cat (Feliformia) to dog and ferret (Caniformia) genes demonstrated that the tiger and cat share a start codon mutation that truncated most of the tetherin cytoplasmic tail early in the Feliformia lineage (19 of 27 amino acids, including the dual tyrosine motif). Alpha interferon (IFN-α) induced tetherin and blocked feline immunodeficiency virus (FIV) replication in lymphoid and nonlymphoid feline cells. Budding of bald FIV and HIV particles was blocked by carnivore tetherins. However, infectious FIV particles were resistant, and spreading FIV replication was uninhibited. Antagonism mapped to the envelope glycoprotein (Env), which rescued FIV from carnivore tetherin restriction when expressed in trans but, in contrast to known antagonists, did not rescue noncognate particles. Also unlike the primate lentiviral antagonists, but similar to the Ebola virus glycoprotein, FIV Env did not reduce intracellular or cell surface tetherin levels. Furthermore, FIV-enveloped FIV particles actually required tetherin for optimal release from cells. The results show that FIV Envs mediate a distinctive tetherin evasion. Well adapted to a phylogenetically ancient tetherin tail truncation in the Felidae, it requires functional virion incorporation of Env, and it shields the budding particle without downregulating plasma membrane tetherin. Moreover, FIV has evolved dependence on this protein: particles containing FIV Env need tetherin for optimal release from the cell, while Env(-) particles do not. IMPORTANCE: HIV-1 antagonizes the restriction factor tetherin with the accessory protein Vpu, while HIV-2 and the filovirus Ebola use their envelope (Env) glycoproteins for this purpose. It turns out that the FIV tetherin antagonist is also its Env protein, but the mechanism is distinctive. Unlike other tetherin antagonists, FIV Env cannot act in trans to rescue vpu-deficient HIV-1. It must be incorporated specifically into FIV virions to be active. Also unlike other retroviral antagonists, but similar to Ebola virus Env, it does not act by downregulating or degrading tetherin. FIV Env might exclude tetherin locally or direct assembly to tetherin-negative membrane domains. Other distinctive features are apparent, including evidence that this virus evolved an equilibrium in which tetherin is both restriction factor and cofactor, as FIV requires tetherin for optimal particle release.