Distinct genetic clusters in HIV-1 CRF01_AE-infected patients induced variable degrees of CD4+ T-cell loss.

CRF01_AE strains have been shown to form multiple transmission clusters in China, and some clusters have disparate pathogenicity in Chinese men who have sex with men. This study focused on other CRF01_AE clusters prevalent in heterosexual populations. The CD4+ T-cell counts from both cross-section data in National HIV Molecular Epidemiology Survey and seropositive cohort data were used to evaluate the pathogenicity of the CRF01_AE clusters and other HIV-1 sub-types. Their mechanisms of pathogenicity were evaluated by co-receptor tropisms, predicted by genotyping and confirmed with virus isolate phenotyping, as well as inflammation parameters. Our research elucidated that individuals infected with CRF01_AE clusters 1 and 2 exhibited significantly lower baseline CD4+ T-cell counts and greater CD4+ T-cell loss in cohort follow-up, compared with other HIV-1 sub-types and CRF01_AE clusters. The increased pathogenesis of cluster 1 or 2 was associated with higher CXCR4 tropisms, higher inflammation/immune activation, and increased pyroptosis. The protein structure modeling analysis revealed that the envelope V3 loop of clusters 1 and 2 viruses is favorable for CXCR4 co-receptor usage. Imbedded with the most mutating reverse transcriptase, HIV-1 is one of the most variable viruses. CRF01_AE clusters 1 and 2 have been found to have evolved into more virulent strains in regions with predominant heterosexual infections. The virulent strains increased the pressure for early diagnosis and treatment in HIV patients. To save more lives, HIV-1 surveillance systems should be upgraded from serology and genotyping to phenotyping, which could support precision interventions for those infected by virulent viruses. IMPORTANCE: Retroviruses swiftly adapt, employing error-prone enzymes for genetic and phenotypic evolution, optimizing survival strategies, and enhancing virulence levels. HIV-1 CRF01_AE has persistently undergone adaptive selection, and cluster 1 and 2 infections display lower counts and fast loss of CD4+ T cells than other HIV-1 sub-types and CRF01_AE clusters. Its mechanisms are associated with increased CXCR4 tropism due to an envelope structure change favoring a tropism shift from CCR5 to CXCR4, thereby shaping viral phenotype features and impacting pathogenicity. This underscores the significance of consistently monitoring HIV-1 genetic evolution and phenotypic transfer to see whether selection bias across risk groups alters the delicate balance of transmissible versus toxic trade-offs, since virulent strains such as CRF01_AE clusters 1 and 2 could seriously compromise the efficacy of antiviral treatment. Only through such early warning and diagnostic services can precise antiviral treatments be administered to those infected with more virulent HIV-1 strains.

Reforestation of Cunninghamia lanceolata changes the relative abundances of important prokaryotic families in soil.

Over the past decades, many forests have been converted to monoculture plantations, which might affect the soil microbial communities that are responsible for governing the soil biogeochemical processes. Understanding how reforestation efforts alter soil prokaryotic microbial communities will therefore inform forest management. In this study, the prokaryotic communities were comparatively investigated in a secondary Chinese fir forest (original) and a reforested Chinese fir plantation (reforested from a secondary Chinese fir forest) in Southern China. The results showed that reforestation changed the structure of the prokaryotic community: the relative abundances of important prokaryotic families in soil. This might be caused by the altered soil pH and organic matter content after reforestation. Soil profile layer depth was an important factor as the upper layers had a higher diversity of prokaryotes than the lower ones (p < 0.05). The composition of the prokaryotic community presented a seasonality characteristic. In addition, the results showed that the dominant phylum was Acidobacteria (58.86%) with Koribacteraceae (15.38%) as the dominant family in the secondary Chinese fir forest and the reforested plantation. Furthermore, soil organic matter, total N, hydrolyzable N, and NH4+-N were positively correlated with prokaryotic diversity (p < 0.05). Also, organic matter and NO3--N were positively correlated to prokaryotic abundance (p < 0.05). This study demonstrated that re-forest transformation altered soil properties, which lead to the changes in microbial composition. The changes in microbial community might in turn influence biogeochemical processes and the environmental variables. The study could contribute to forest management and policy-making.

PREB inhibits the replication of prototype foamy virus by affecting its transcription.

BACKGROUND: Foamy viruses (FVs) are unique nonpathogenic retroviruses, which remain latent in the host for a long time. Therefore, they may be safe, effective gene transfer vectors. In this study, were assessed FV-host cell interactions and the molecular mechanisms underlying FV latent infection. METHODS: We used the prototype FV (PFV) to infect HT1080 cells and a PFV indicator cell line (PFVL) to measure virus titers. After 48 h of infection, the culture supernatant (i.e., cell-free PFV particles) and transfected cells (i.e., cell-associated PFV particles) were harvested and incubated with PFVL. After another 48 h, the luciferase activity was used to measure virus titers. RESULTS: Through transcriptomics sequencing, we found that PREB mRNA expression was significantly upregulated. Moreover, PREB overexpression reduced PFV replication, whereas endogenous PREB knockdown increased PFV replication. PREB interacted with the Tas DNA-binding and transcriptional activation domains and interfered with its binding to the PFV long terminal repeat and internal promoter, preventing the recruitment of transcription factors and thereby inhibiting the transactivation function of Tas. PREB C-terminal 329-418 aa played a major role in inhibiting PFV replication; PREB also inhibited bovine FV replication. Therefore, PREB has a broad-spectrum inhibitory effect on FV replication. CONCLUSIONS: Our results demonstrated that PREB inhibits PFV replication by impeding its transcription.

Effects of Chemokine Ligand 2 on Budding of Bovine Foamy Virus.

The endosomal sorting complex required for transport (ESCRT) machinery is essential for the budding of retroviruses such as human immunodeficiency virus (HIV) and bovine foamy virus (BFV), which rely on their late domain to recruit ESCRT complexes to facilitate budding. However, the impact of intracellular host proteins on BFV budding remains poorly understood. In this study, we aimed to investigate the impact of CCL2 on BFV budding and interactions with key host proteins. Our results indicate that CCL2 promotes BFV budding in an ALG-2-interacting protein X (Alix)-dependent manner by enhancing the interaction between Alix and BFV Gag (BGag). Notably, we found a link between Alix, BGag and CCL2, with Alix mediating the interaction between the latter two. Furthermore, we observed that natural host bovine CCL2 also has a facilitating role in the budding process of BFV, similar to human CCL2. Taken together, these results demonstrate that CCL2 promotes BFV budding by enhancing the Alix-BGag association.

Comparative Analysis and Structure Identification of Oxidative Metabolites and Hydrogenation Metabolite Enantiomers for 2-Fluorodeschloroketamine.

In this study, we used solid-phase extraction with liquid chromatography-ion trap time-of-flight mass spectrometry (LC-IT-TOF-MS) to analyze 2-fluorodeschloroketamine (2-FDCK) metabolites in human urine. The complete set of oxidative metabolites was identified, with 17 compounds divided into four groups. Furthermore, we examined the hydroxy substitution site after oxidative metabolism with theoretical calculation and 2-FDCK nuclear magnetic resonance (NMR) data. We clarified the correlation of the oxidative metabolic sites with the electron cloud density in the structure. Additionally, two enantiomers of dihydro-2-fluorodeschloroketamine (dihydro-2-FDCK) were determined by using a laboratory-made dihydro-2-FDCK hydrochloride reference substance. Their configurations were determined via NMR spectrometry data prediction of the ACD Labs-Structure Elucidator Suite software and theoretical calculation. Moreover, the stereoselectivity of the related enzymes in hydrogenation metabolism in vivo was clarified. These findings provide an important reference for analyzing other oxidative metabolites, laying the foundation for future analysis, prediction, elucidation and identification of the latest ketamine-type new psychoactive substance metabolites.

Antiviral role of IFITM3 in prototype foamy virus infection.

BACKGROUND: Foamy viruses (FVs) are retroviruses with unique replication strategies that cause lifelong latent infections in their hosts. FVs can also produce foam-like cytopathic effects in vitro. However, the effect of host cytokines on FV replication requires further investigation. Although interferon induced transmembrane (IFITMs) proteins have become the focus of antiviral immune response research due to their broad-spectrum antiviral ability, it remains unclear whether IFITMs can affect FV replication. METHOD: In this study, the PFV virus titer was characterized by measuring luciferase activity after co-incubation of PFVL cell lines with the cell culture supernatants (cell-free PFV) or the cells transfected with pcPFV plasmid/infected with PFV (cell-associated PFV). The foam-like cytopathic effects of PFV infected cells was observed to reflect the virus replication. The total RNA of PFV infected cells was extracted, and the viral genome was quantified by Quantitative reverse transcription PCR to detect the PFV entry into target cells. RESULTS: In the present study, we demonstrated that IFITM1-3 overexpression inhibited prototype foamy virus (PFV) replication. In addition, an IFITM3 knockdown by small interfering RNA increased PFV replication. We further demonstrated that IFITM3 inhibited PFV entry into host cells. Moreover, IFITM3 also reduced the number of PFV envelope proteins, which was related to IFITM3 promoted envelope degradation through the lysosomal pathway. CONCLUSIONS: Taken together, these results demonstrate that IFITM3 inhibits PFV replication by inhibiting PFV entry into target cells and reducing the number of PFV envelope.

TRAF3IP3 Is Cleaved by EV71 3C Protease and Exhibits Antiviral Activity.

Enterovirus 71 (EV71) is one of the major pathogens of hand, foot, and mouth disease, which poses a major risk to public health and infant safety. 3C protease (3Cpro), a non-structural protein of EV71, promotes viral protein maturation by cleaving polyprotein precursors and facilitates viral immune escape by cleaving host proteins. In this study, we screened for human proteins that could interact with EV71 3Cpro using a yeast two-hybrid assay. Immune-associated protein TRAF3 Interacting Protein 3 (TRAF3IP3) was selected for further study. The results of co-immunoprecipitation and immunofluorescence demonstrated the interaction between TRAF3IP3 and EV71 3Cpro. A cleavage band was detected, indicating that both transfected 3Cpro and EV71 infection could cleave TRAF3IP3. 87Q-88G was identified as the only 3Cpro cleavage site in TRAF3IP3. In Jurkat and rhabdomyosarcoma (RD) cells, TRAF3IP3 inhibited EV71 replication, and 3Cpro cleavage partially resisted TRAF3IP3-induced inhibition. Additionally, the nuclear localization signal (NLS) and nuclear export signal (NES) of TRAF3IP3 were identified. The NES contributed to TRAF3IP3 alteration of 3Cpro localization and inhibition of EV71 replication. Together, these results indicate that TRAF3IP3 inhibits EV71 replication and 3Cpro resists such inhibition via proteolytic cleavage, providing a new example of virus-host interaction.

SGK1, a Serine/Threonine Kinase, Inhibits Prototype Foamy Virus Replication.

Foamy viruses (FVs) are complex retroviruses belonging to the Spumaretrovirinae subfamily of the Retroviridae family. In contrast to human immunodeficiency virus (HIV), another member of the Retroviridae family, FVs are nonpathogenic in their natural hosts or in experimentally infected animals. Prototype foamy virus (PFV) is the only foamy virus that can infect humans through cross-species transmission and does not show any pathogenicity after infection. Consequently, PFV is considered a safe and efficient gene transfer vector. Understanding the host proteins involved in the replication of PFV and the mechanism of interaction between the host and the virus might lead to studies to improve the efficiency of gene transfer. To date, only a few host factors have been identified that affect PFV replication. In the present study, we report that PFV infection enhances the promoter activity of SGK1 (encoding serum/glucocorticoid regulated kinase 1) via the Tas protein signaling pathway, and then upregulates the mRNA and protein levels of SGK1. Overexpression of SGK1 reduced PFV replication, whereas its depletion using small interfering RNA increased PFV replication. SGK1 inhibits PFV replication by impairing the function of the PFV Tas activation domain in a kinase-independent manner and reducing the stability of the Gag protein in a kinase-dependent manner. In addition, both human and bovine SGK1 proteins inhibit the replication of bovine foamy virus (BFV) and PFV. These findings not only improved our understanding of the function of SGK1 and its relationship with foamy viruses, but also contributed to determining the antiviral mechanism of the host. IMPORTANCE Foamy viruses can integrate into the host chromosome and are nonpathogenic in natural hosts or in experimentally infected animals. Therefore, foamy viruses are considered to be safe and efficient gene transfer vectors. Persistent infection of foamy viruses is partly caused by the restrictive effect of host factors on the virus. However, only a few cellular proteins are known to influence the replication of foamy viruses. In this study, we report that SGK1 inhibits the replication of prototype foamy virus by affecting the function of the transcription activator, Tas, and reducing the stability of the structural protein, Gag. These results will increase our understanding of the interaction between the virus and host factors, deepening our perception of host antiviral defenses and the function of SGK1, and could improve the gene transfer efficiency of foamy viruses.

Characterization of Bovine Foamy Virus Gag Late Assembly Domain Motifs and Their Role in Recruiting ESCRT for Budding.

A large number of retroviruses, such as human immunodeficiency virus (HIV) and prototype foamy virus (PFV), recruit the endosomal sorting complex required for transport (ESCRT) through the late domain (L domain) on the Gag structural protein for virus budding. However, little is known about the molecular mechanism of bovine foamy virus (BFV) budding. In the present study, we report that BFV recruits ESCRT for budding through the L domain of Gag. Specifically, knockdown of VPS4 (encoding vacuolar protein sorting 4), ALIX (encoding ALG-2-interacting protein X), and TSG101 (encoding tumor susceptibility 101) indicated that BFV uses ESCRT for budding. Mutational analysis of BFV Gag (BGag) showed that, in contrast to the classical L domain motifs, BGag contains two motifs, P56LPI and Y103GPL, with L domain functions. In addition, the two L domains are necessary for the cytoplasmic localization of BGag, which is important for effective budding. Furthermore, we demonstrated that the functional site of Alix is V498 in the V domain and the functional site of Tsg101 is N69 in the UBC-like domain for BFV budding. Taken together, these results demonstrate that BFV recruits ESCRT for budding through the PLPI and YGPL L domain motifs in BGag.

Enterovirus 71 Activates GADD34 via Precursor 3CD to Promote IRES-Mediated Viral Translation.

Enterovirus 71 (EV71) is the major pathogen of hand, foot, and mouth disease. In severe cases, it can cause life-threatening neurological complications, such as aseptic meningitis and polio-like paralysis. There are no specific antiviral treatments for EV71 infections. In a previous study, the host protein growth arrest and DNA damage-inducible protein 34 (GADD34) expression was upregulated during EV71 infection determined by ribosome profiling and RNA-sequencing. Here, we investigated the interactions of host protein GADD34 and EV71 during infections. Rhabdomyosarcoma (RD) cells were infected with EV71 resulting in a significant increase in expression of GADD34 mRNA and protein. Through screening of EV71 protein we determined that the non-structural precursor protein 3CD is responsible for upregulating GADD34. EV71 3CD increased the RNA and protein levels of GADD34, while the 3CD mutant Y441S could not. 3CD upregulated GADD34 translation via the upstream open reading frame (uORF) of GADD34 5'untranslated regions (UTR). EV71 replication was attenuated by the knockdown of GADD34. The function of GADD34 to dephosphorylate eIF2α was unrelated to the upregulation of EV71 replication, but the PEST 1, 2, and 3 regions of GADD34 were required. GADD34 promoted the EV71 internal ribosome entry site (IRES) activity through the PEST repeats and affected several other viruses. Finally, GADD34 amino acids 563 to 565 interacted with 3CD, assisting GADD34 to target the EV71 IRES. Our research reveals a new mechanism by which GADD34 promotes viral IRES and how the EV71 non-structural precursor protein 3CD regulates host protein expression to support viral replication. IMPORTANCE Identification of host factors involved in viral replication is an important approach in discovering viral pathogenic mechanisms and identifying potential therapeutic targets. Previously, we screened host proteins that were upregulated by EV71 infection. Here, we report the interaction between the upregulated host protein GADD34 and EV71. EV71 non-structural precursor protein 3CD activates the RNA and protein expression of GADD34. Our study reveals that 3CD regulates the uORF of the 5'-UTR to increase GADD34 translation, providing a new explanation for how viral proteins regulate host protein expression. GADD34 is important for EV71 replication, and the key functional domains of GADD34 that promote EV71 are PEST 1, 2, and 3 regions. We report that GADD34 promotes viral IRES for the first time and this process is independent of its eIF2α phosphatase activity.

3C protease of enterovirus 71 cleaves promyelocytic leukemia protein and impairs PML-NBs production.

BACKGROUND: Enterovirus 71 (EV71) usually infects infants causing hand-foot-mouth disease (HFMD), even fatal neurological disease like aseptic meningitis. Effective drug for preventing and treating EV71 infection is unavailable currently. EV71 3C mediated the cleavage of many proteins and played an important role in viral inhibiting host innate immunity. Promyelocytic leukemia (PML) protein, the primary organizer of PML nuclear bodies (PML-NBs), can be induced by interferon and is involved in antiviral activity. PML inhibits EV71 replication, and EV71 infection reduces PML expression, but the molecular mechanism is unclear. METHODS: The cleavage of PMLIII and IV was confirmed by co-transfection of EV71 3C protease and PML. The detailed cleavage sites were evaluated further by constructing the Q to A mutant of PML. PML knockout cells were infected with EV71 to identify the effect of cleavage on EV71 replication. Immunofluorescence analysis to examine the interference of EV71 3C on the formation of PML-NBs. RESULTS: EV71 3C directly cleaved PMLIII and IV. Furthermore, 3C cleaved PMLIV at the sites of Q430-A431 and Q444-S445 through its protease activity. Overexpression of PMLIV Q430A/Q444A variant exhibited stronger antiviral potential than the wild type. PMLIV Q430A/Q444A formed normal nuclear bodies that were not affected by 3C, suggesting that 3C may impair PML-NBs production via PMLIV cleavage and counter its antiviral activities. PML, especially PMLIV, which sequesters viral proteins in PML-NBs and inhibits viral production, is a novel target of EV71 3C cleavage. CONCLUSIONS: EV71 3C cleaves PMLIV at Q430-A431 and Q444-S445. Cleavage reduces the antiviral function of PML and decomposes the formation of PML-NBs, which is conducive to virus replication.

Effect of Different Nuclear Localization Signals on the Subcellular Localization and Anti-HIV-1 Function of the MxB Protein.

Interferon exerts its antiviral activity by stimulating the expression of antiviral proteins. These interferon stimulate genes (ISGs) often target a group of viruses with unique molecular mechanisms. One such ISG is myxovirus resistance B (MxB) that has been reported to inhibit human immunodeficiency virus type 1 (HIV-1) by targeting viral capsid and impairing nuclear import of viral DNA. The antiviral specificity of MxB is determined by its N-terminal 25 amino acids sequence which has the nuclear localization activity, therefore functions as a nuclear localization signal (NLS). In this study, we report that the bipartite NLS, but not the classic NLS, the PY-NLS, nor the arginine-rich NLS, when used to replace the N-terminal sequence of MxB, drastically suppress HIV-1 gene expression and virus production, thus creates a new anti-HIV-1 mechanism. MxB preserves its anti-HIV-1 activity when its N-terminal sequence is replaced by the arginine-rich NLS. Interestingly, the arginine-rich NLS allows MxB to inhibit HIV-1 CA mutants that are otherwise resistant to wild type MxB, which suggests sequence specific targeting of viral capsid. Together, these data implicate that it is not the nuclear import function itself, but rather the sequence and the mechanism of action of the NLS which define the antiviral property of MxB.

Identification of the internal ribosome entry sites in the 5'­untranslated region of the c­fos gene.

The Fos proto­oncogene, activator protein­1 (AP­1) transcription factor subunit (c­fos) gene, a member of the immediate early gene family, encodes c­Fos, which is a subunit of the AP­1 transcription factor. The present study aimed to investigate the mechanism by which the translation efficiency of c­fos mRNA is upregulated when cellular protein synthesis is shut off. The result of western blotting revealed that the protein expression levels of c­Fos were increased in rhabdomyosarcoma cells infected with enterovirus 71 (EV71) compared with uninfected cells. PCR was used to get the c­fos 5'­untranslated region (UTR). The luciferase assay of a bicistronic vector containing the c­fos 5'UTR revealed that the c­fos 5'UTR contains an internal ribosome entry site (IRES) sequence and a 175 nucleotide sequence (between 31 and 205 nt) that is essential for IRES activity. Analysis of potential IRES trans­acting factors revealed that poly(C)­binding protein 2 (PCBP2) negatively regulated the activity of the c­fos IRES, whereas the La autoantigen (La) positively regulated its activity. The results of RNA­protein immunoprecipitation demonstrated that both PCBP2 and La bound to the c­fos 5'UTR. Furthermore, the IRES activity of in vitro­transcribed c­fos mRNA was upregulated during EV71 infection. The present study suggested a mechanism for the effect of viral infection on host genes, and provided a novel target for gene translation regulation.

Human Schlafen 11 exploits codon preference discrimination to attenuate viral protein synthesis of prototype foamy virus (PFV).

Recently, the Schlafen (SLFN) proteins have been identified as a novel interferon-stimulated family with antiviral properties. In this study, we reported that SLFN11 inhibited prototype foamy virus (PFV) replication. Over-expression of human SLFN11 reduced viral production, while knockdown of SLFN11 enhanced viral infectivity. In addition, SLFN11 from cattle and African green monkey also suppressed PFV production. Both the ATPase activity and helicase activity of SLFN11 were required for its inhibitory function. Dephosphorylation activated the antiviral activity of SLFN11. More importantly, SLFN11 inhibited the expression of viral protein, which was rescued by viral gene codon optimization. Together, our results demonstrated that SLFN11 impaired PFV viral protein synthesis by exploiting the distinct codon usage between the virus and the host. These findings further broaden our understanding of the antiviral properties of the SLFN family and the molecular mechanism of PFV latent infection.

IFI44L expression is regulated by IRF-1 and HIV-1.

Interferon (IFN)-inducible 44 like (IFI44L) is an IFN-stimulated gene (ISG), which is located on the same chromosome as the known antiviral ISG IFI44. Expression of IFI44L is induced by IFN and HIV-1 infection. However, the mechanism by which IFN-I induces IFI44L production has not yet been determined. In this study, we analyzed transcriptional regulation of IFI44L via cloning of the IFI44L promoter. We found that IFI44L has two IFN-stimulated response elements (ISRE), which are necessary for the basal level of IFI44L transcription. IFN-I and IFN-II can activate the IFI44L promoter through one of the two ISREs. IFN regulatory factor (IRF)-1 can activate transcription of IFI44L by binding to one of the ISREs. Additionally, co-transfection of the IFI44L promoter with an HIV-1 infectious clone or HIV-1 infection activated IFI44L promoter transcription, but did not upregulate IFI44L expression via ISREs. These findings will help to understand the interaction between IFI44L and HIV-1, and aid in elucidation of the role of IFI44L in the antiviral innate immune response.

Palmitoylation of the Bovine Foamy Virus Envelope Glycoprotein Is Required for Viral Replication.

Membrane proteins of enveloped viruses have been reported to undergo palmitoylation, a post-translational modification often having a critical role in the function of these viral proteins and hence viral replication. In this study, we report that the foamy virus (FV) envelope (Env) glycoprotein is palmitoylated. Specifically, we found that bovine foamy virus (BFV) Env (BEnv) is palmitoylated at amino acid positions C58 and C59 by BDHHC3 and BDHHC20 in a DHHC motif-dependent manner. In addition, mutations C58S and C58/59S significantly decrease cell surface expression of BEnv, subviral particle (SVP) egress, and its membrane fusion activity, thus ultimately inhibiting BFV replication. The C59S mutation exerts a minor effect in this regard. Taken together, these data demonstrate that the function of BEnv in the context of BFV replication is under the regulation of palmitoylation.

HIV-1 resists MxB inhibition of viral Rev protein.

The interferon-inducible myxovirus resistance B (MxB) protein has been reported to inhibit HIV-1 and herpesviruses by blocking the nuclear import of viral DNA. Here, we report a new antiviral mechanism in which MxB restricts the nuclear import of HIV-1 regulatory protein Rev, and as a result, diminishes Rev-dependent expression of HIV-1 Gag protein. Specifically, MxB disrupts the interaction of Rev with the nuclear transport receptor, transportin 1 (TNPO1). Supporting this, the TNPO1-independent Rev variants become less restricted by MxB. In addition, HIV-1 can overcome this inhibition by MxB through increasing the expression of multiply spliced viral RNA and hence Rev protein. Therefore, MxB exerts its anti-HIV-1 function through interfering with the nuclear import of both viral DNA and viral Rev protein.

Prototype foamy virus downregulates RelB expression to facilitate viral replication.

Foamy viruses (FVs) are classified in the subfamily Spumaretrovirinae and bridge the gap between Orthoretrovirinae and Hepadnaviridae. FVs have strong cytopathic effects against cells cultured in vitro. However, they establish lifelong latent infections without evident pathology in the host. The roles of cellular factors in FV replication are poorly understood. To better understand this area, we determined the transcriptomes of HT1080 cells infected with prototype foamy virus (PFV) to measure the effect of PFV infection on the expression of cellular genes. We found that the level of RelB mRNA, a member of the nuclear factor-κB (NF-κB) protein family, was significantly decreased as a result of PFV infection, and this was further confirmed with real-time PCR. Interestingly, overexpression of RelB reduced PFV replication, whereas its depletion using small interfering RNA increased PFV replication. This inhibitory effect of RelB results from diminished transactivation of the viral long terminal repeat (LTR) promoter and an internal promoter (IP) by viral Tas protein. Together, these data demonstrate that PFV infection downregulates the viral inhibitory host factor RelB, which otherwise restricts viral gene expression.

Cyclic loading test study on a new cast-in-situ insulated sandwich concrete wall.

Insulated sandwich concrete panel (ISCP) is widely used because of its high thermal insulation efficiency and low construction cost. Aiming at improving traditional ISCP, a new cast-in-situ concrete wall structure made of ISCP is proposed, which is composed of thin-walled cold-formed steels, slant steel wire connectors, steel wire meshes, concrete layers, expanded polystyrene sheets and reinforced concrete embedded columns. In order to assess the hysteretic properties of the new insulated sandwich concrete wall and the influence of various parameters, low-frequency horizontal cyclic load tests were carried out on seven full-scale specimens of new type cast-in-situ insulated sandwich concrete wall. The specimens were compared and analyzed with respect to failure mode, bearing capacity, ductility, degradation characteristics and energy dissipation capacity. The results show that the final failure pattern of the specimen is two main diagonal cracks intersecting each other; the bearing capacity is greatly affected by concrete thickness and axial compression ratio, regardless of concrete strength. Brittle failure is typically observed when the steel wire spacing is large, while ductility is pronounced when the concrete layer thickness is small and the concrete strength is low; the smaller the thickness of concrete layer, the faster the stiffness degrades. The wall structure shows a better energy dissipation performance with a smaller steel wire spacing, lower concrete strength and smaller axial compression ratio.

Ribosome Profiling of Vaccinia Virus-Infected Cells.

Ribosome profiling is a method that determines genome-wide mRNA translation through measuring ribosome-protected mRNA fragments by deep sequencing. This method can be used to quantify gene expression at the translational level and precisely pinpoint ribosome loading onto mRNA with codon-level resolution. Genome-wide regulation of mRNA translation can also be determined if RNA-Sequencing (RNA-Seq) is carried out in parallel. Here, we describe a protocol for simultaneously performing ribosome profiling and RNA-Seq in cells infected with vaccinia virus.