bluuebunny/crossref_metadata_2025_split huggingface dataset:
HIV-1 Vpr- and Reverse Transcription-Induced Apoptosis in Resting Peripheral Blood CD4 T Cells and Protection by Common Gamma-Chain Cytokines
Authors: Trinité Benjamin , Chan Chi N. , Lee Caroline S. , Levy David N. 
Published: November 2015

ABSTRACTHIV-1 infection leads to the progressive depletion of the CD4 T cell compartment by various known and unknown mechanisms.In vivo, HIV-1 infects both activated and resting CD4 T cells, butin vitro, in the absence of any stimuli, resting CD4 T cells from peripheral blood are resistant to infection. This resistance is generally attributed to an intracellular environment that does not efficiently support processes such as reverse transcription (RT), resulting in abortive infection. Here, we show thatin vitroHIV-1 infection of resting CD4 T cells induces substantial cell death, leading to abortive infection.In vivo, however, various microenvironmental stimuli in lymphoid and mucosal tissues provide support for HIV-1 replication. For example, common gamma-chain cytokines (CGCC), such as interleukin-7 (IL-7), render resting CD4 T cells permissible to HIV-1 infection without inducing T cell activation. Here, we find that CGCC primarily allow productive infection by preventing HIV-1 triggering of apoptosis, as evidenced by early release of cytochromecand caspase 3/7 activation. Cell death is triggered both by products of reverse transcription and by virion-borne Vpr protein, and CGCC block both mechanisms. When HIV-1 RT efficiency was enhanced by SIVmac239 Vpx protein, cell death was still observed, indicating that the speed of reverse transcription and the efficiency of its completion contributed little to HIV-1-induced cell death in this system. These results show that a major restriction on HIV-1 infection in resting CD4 T cells resides in the capacity of these cells to survive the early steps of HIV-1 infection.IMPORTANCEA major consequence of HIV-1 infection is the destruction of CD4 T cells. Here, we show that delivery of virion-associated Vpr protein and the process of reverse transcription are each sufficient to trigger apoptosis of resting CD4 T cells isolated from peripheral blood. While these 2 mechanisms have been previously described in various cell types, we show for the first time their concerted effect in inducing resting CD4 T cell depletion. Importantly, we found that cytokines such as IL-7 and IL-4, which are particularly active in sites of HIV-1 replication, protect resting CD4 T cells from these cytopathic effects and, primarily through this protection, rather than through enhancement of specific replicative steps, they promote productive infection. This study provides important new insights for the understanding of the early steps of HIV-1 infection and T cell depletion.
Functional Characterization of the Serine-Rich Tract of Varicella-Zoster Virus IE62
Authors: Kim Seong K. , Shakya Akhalesh K. , Kim Seongman , O'Callaghan Dennis J. 
Published: November 2015

ABSTRACT

The immediate early 62 protein (IE62) of varicella-zoster virus (VZV), a major viral
trans
-activator, initiates the virus life cycle and is a key component of pathogenesis. The IE62 possesses several domains essential for
trans
-activation, including an acidic
trans
-activation domain (TAD), a serine-rich tract (SRT), and binding domains for USF, TFIIB, and TATA box binding protein (TBP). Transient-transfection assays showed that the VZV IE62 lacking the SRT
trans
-activated the early VZV ORF61 promoter at only 16% of the level of the full-length IE62. When the SRT of IE62 was replaced with the SRT of equine herpesvirus 1 (EHV-1) IEP, its
trans
-activation activity was completely restored. Herpes simplex virus 1 (HSV-1) ICP4 that lacks a TAD very weakly (1.5-fold)
trans
-activated the ORF61 promoter. An IE62 TAD-ICP4 chimeric protein exhibited
trans
-activation ability (10.2-fold), indicating that the IE62 TAD functions with the SRT of HSV-1 ICP4 to
trans
-activate viral promoters. When the serine and acidic residues of the SRT were replaced with Ala, Leu, and Gly,
trans
-activation activities of the modified IE62 proteins IE62-SRTΔSe and IE62-SRTΔAc were reduced to 46% and 29% of wild-type activity, respectively. Bimolecular complementation assays showed that the TAD of IE62, EHV-1 IEP, and HSV-1 VP16 interacted with Mediator 25 in human melanoma MeWo cells. The SRT of IE62 interacted with the nucleolar-ribosomal protein EAP, which resulted in the formation of globular structures within the nucleus. These results suggest that the SRT plays an important role in VZV viral gene expression and replication.

IMPORTANCE
The immediate early 62 protein (IE62) of varicella-zoster virus (VZV) is a major viral
trans
-activator and is essential for viral growth. Our data show that the serine-rich tract (SRT) of VZV IE62, which is well conserved within the alphaherpesviruses, is needed for
trans
-activation mediated by the acidic
trans
-activation domain (TAD). The TADs of IE62, EHV-1 IEP, and HSV-1 VP16 interacted with cellular Mediator 25 in bimolecular complementation assays. The interaction of the IE62 SRT with nucleolar-ribosomal protein EAP resulted in the formation of globular structures within the nucleus. Understanding the mechanisms by which the TAD and SRT of IE62 contribute to the function of this essential regulatory protein is important in understanding the gene program of this human pathogen.
Identification of an HIV-1 Mutation in Spacer Peptide 1 That Stabilizes the Immature CA-SP1 Lattice
Authors: Fontana Juan , Keller Paul W. , Urano Emiko , Ablan Sherimay D. , Steven Alasdair C. , Freed Eric O. 
Published: November 2015

ABSTRACT
Upon release of HIV-1 particles from the infected cell, the viral protease cleaves the Gag polyprotein at specific sites, triggering maturation. During this process, which is essential for infectivity, the capsid protein (CA) reassembles into a conical core. Maturation inhibitors (MIs) block HIV-1 maturation by interfering with protease-mediated CA-spacer peptide 1 (CA-SP1) processing, concomitantly stabilizing the immature CA-SP1 lattice; virions from MI-treated cells retain an immature-like CA-SP1 lattice, whereas mutational abolition of cleavage at the CA-SP1 site results in virions in which the CA-SP1 lattice converts to a mature-like form. We previously reported that propagation of HIV-1 in the presence of MI PF-46396 selected for assembly-defective, compound-dependent mutants with amino acid substitutions in the major homology region (MHR) of CA. Propagation of these mutants in the absence of PF-46396 resulted in the acquisition of second-site compensatory mutations. These included a Thr-to-Ile substitution at SP1 residue 8 (T8I), which results in impaired CA-SP1 processing. Thus, the T8I mutation phenocopies PF-46396 treatment in terms of its ability to rescue the replication defect imposed by the MHR mutations and to impede CA-SP1 processing. Here, we use cryo-electron tomography to show that, like MIs, the T8I mutation stabilizes the immature-like CA-SP1 lattice. These results have important implications for the mechanism of action of HIV-1 MIs; they also suggest that T8I may provide a valuable tool for structural definition of the CA-SP1 boundary region, which has thus far been refractory to high-resolution analysis, apparently because of conformational flexibility in this region of Gag.

IMPORTANCE
HIV-1 maturation involves dissection of the Gag polyprotein by the viral protease and assembly of a conical capsid enclosing the viral ribonucleoprotein. Maturation inhibitors (MIs) prevent the final cleavage step at the site between the capsid protein (CA) and spacer peptide 1 (SP1), apparently by binding at this site and denying the protease access. Additionally, MIs stabilize the immature-like CA-SP1 lattice, preventing release of CA into the soluble pool. We previously found that T8I, a mutation in SP1, rescues a PF-46396-dependent CA mutant and blocks CA-SP1 cleavage. In this study, we imaged T8I virions by cryo-electron tomography and showed that T8I mutants, like MI-treated virions, contain an immature CA-SP1 lattice. These results lay the groundwork needed to understand the structure of the CA-SP1 interface region and further illuminate the mechanism of action of MIs.
Structural Studies of Chikungunya Virus-Like Particles Complexed with Human Antibodies: Neutralization and Cell-to-Cell Transmission
Authors: Porta Jason , Mangala Prasad Vidya , Wang Cheng-I , Akahata Wataru , Ng Lisa F. P. , Rossmann Michael G. 
Published: November 2015

ABSTRACT
Chikungunya virus is a positive-stranded RNA alphavirus. Structures of chikungunya virus-like particles in complex with strongly neutralizing antibody Fab fragments (8B10 and 5F10) were determined using cryo-electron microscopy and X-ray crystallography. By fitting the crystallographically determined structures of these Fab fragments into the cryo-electron density maps, we show that Fab fragments of antibody 8B10 extend radially from the viral surface and block receptor binding on the E2 glycoprotein. In contrast, Fab fragments of antibody 5F10 bind the tip of the E2 B domain and lie tangentially on the viral surface. Fab 5F10 fixes the B domain rigidly to the surface of the virus, blocking exposure of the fusion loop on glycoprotein E1 and therefore preventing the virus from becoming fusogenic. Although Fab 5F10 can neutralize the wild-type virus, it can also bind to a mutant virus without inhibiting fusion or attachment. Although the mutant virus is no longer able to propagate by extracellular budding, it can, however, enter the next cell by traveling through junctional complexes without being intercepted by a neutralizing antibody to the wild-type virus, thus clarifying how cell-to-cell transmission can occur.

IMPORTANCE
Alphaviral infections are transmitted mainly by mosquitoes. Chikungunya virus (CHIKV), which belongs to the
Alphavirus
genus, has a wide distribution in the Old World that has expanded in recent years into the Americas. There are currently no vaccines or drugs against alphaviral infections. Therefore, a better understanding of CHIKV and its associated neutralizing antibodies will aid in the development of effective treatments.
Structural Insights into HIV-1 Vif-APOBEC3F Interaction
Authors: Nakashima Masaaki , Ode Hirotaka , Kawamura Takashi , Kitamura Shingo , Naganawa Yuriko , Awazu Hiroaki , Tsuzuki Shinya , Matsuoka Kazuhiro , Nemoto Michiko , Hachiya Atsuko , Sugiura Wataru , Yokomaku Yoshiyuki , Watanabe Nobuhisa , Iwatani Yasumasa 
Published: November 2015

ABSTRACT
The HIV-1 Vif protein inactivates the cellular antiviral cytidine deaminase APOBEC3F (A3F) in virus-infected cells by specifically targeting it for proteasomal degradation. Several studies identified Vif sequence motifs involved in A3F interaction, whereas a Vif-binding A3F interface was proposed based on our analysis of highly similar APOBEC3C (A3C). However, the structural mechanism of specific Vif-A3F recognition is still poorly understood. Here we report structural features of interaction interfaces for both HIV-1 Vif and A3F molecules. Alanine-scanning analysis of Vif revealed that six residues located within the conserved Vif F1-, F2-, and F3-box motifs are essential for both A3C and A3F degradation, and an additional four residues are uniquely required for A3F degradation. Modeling of the Vif structure on an HIV-1 Vif crystal structure revealed that three discontinuous flexible loops of Vif F1-, F2-, and F3-box motifs sterically cluster to form a flexible A3F interaction interface, which represents hydrophobic and positively charged surfaces. We found that the basic Vif interface patch (R17, E171, and R173) involved in the interactions with A3C and A3F differs. Furthermore, our crystal structure determination and extensive mutational analysis of the A3F C-terminal domain demonstrated that the A3F interface includes a unique acidic stretch (L291, A292, R293, and E324) crucial for Vif interaction, suggesting additional electrostatic complementarity to the Vif interface compared with the A3C interface. Taken together, these findings provide structural insights into the A3F-Vif interaction mechanism, which will provide an important basis for development of novel anti-HIV-1 drugs using cellular cytidine deaminases.

IMPORTANCE
HIV-1 Vif targets cellular antiviral APOBEC3F (A3F) enzyme for degradation. However, the details on the structural mechanism for specific A3F recognition remain unclear. This study reports structural features of interaction interfaces for both HIV-1 Vif and A3F molecules. Three discontinuous sequence motifs of Vif, F1, F2, and F3 boxes, assemble to form an A3F interaction interface. In addition, we determined a crystal structure of the wild-type A3F C-terminal domain responsible for the Vif interaction. These results demonstrated that both electrostatic and hydrophobic interactions are the key force driving Vif-A3F binding and that the Vif-A3F interfaces are larger than the Vif-A3C interfaces. These findings will allow us to determine the configurations of the Vif-A3F complex and to construct a structural model of the complex, which will provide an important basis for inhibitor development.
Differential Recognition of Influenza A Viruses by M158–66Epitope-Specific CD8+T Cells Is Determined by Extraepitopic Amino Acid Residues
Authors: van de Sandt Carolien E. , Kreijtz Joost H. C. M. , Geelhoed-Mieras Martina M. , Nieuwkoop Nella J. , Spronken Monique I. , van de Vijver David A. M. C. , Fouchier Ron A. M. , Osterhaus Albert D. M. E. , Rimmelzwaan Guus F. 
Published: November 2015

ABSTRACTNatural influenza A virus infections elicit both virus-specific antibody and CD4+and CD8+T cell responses. Influenza A virus-specific CD8+cytotoxic T lymphocytes (CTLs) contribute to clearance of influenza virus infections. Viral CTL epitopes can display variation, allowing influenza A viruses to evade recognition by epitope-specific CTLs. Due to functional constraints, some epitopes, like the immunodominant HLA-A*0201-restricted matrix protein 1 (M158–66) epitope, are highly conserved between influenza A viruses regardless of their subtype or host species of origin. We hypothesized that human influenza A viruses evade recognition of this epitope by impairing antigen processing and presentation by extraepitopic amino acid substitutions. Activation of specific T cells was used as an indication of antigen presentation. Here, we show that the M158–66epitope in the M1 protein derived from human influenza A virus was poorly recognized compared to the M1 protein derived from avian influenza A virus. Furthermore, we demonstrate that naturally occurring variations at extraepitopic amino acid residues affect CD8+T cell recognition of the M158–66epitope. These data indicate that human influenza A viruses can impair recognition by M158–66-specific CTLs while retaining the conserved amino acid sequence of the epitope, which may represent a yet-unknown immune evasion strategy for influenza A viruses. This difference in recognition may have implications for the viral replication kinetics in HLA-A*0201 individuals and spread of influenza A viruses in the human population. The findings may aid the rational design of universal influenza vaccines that aim at the induction of cross-reactive virus-specific CTL responses.IMPORTANCEInfluenza viruses are an important cause of acute respiratory tract infections. Natural influenza A virus infections elicit both humoral and cellular immunity. CD8+cytotoxic T lymphocytes (CTLs) are directed predominantly against conserved internal proteins and confer cross-protection, even against influenza A viruses of various subtypes. In some CTL epitopes, mutations occur that allow influenza A viruses to evade recognition by CTLs. However, the immunodominant HLA-A*0201-restricted M158–66epitope does not tolerate mutations without loss of viral fitness. Here, we describe naturally occurring variations in amino acid residues outside the M158–66epitope that influence the recognition of the epitope. These results provide novel insights into the epidemiology of influenza A viruses and their pathogenicity and may aid rational design of vaccines that aim at the induction of CTL responses.
Correlation between Virus Replication and Antibody Responses in Macaques following Infection with Pandemic Influenza A Virus
Authors: Koopman Gerrit , Mooij Petra , Dekking Liesbeth , Mortier Daniëlla , Nieuwenhuis Ivonne G. , van Heteren Melanie , Kuipers Harmjan , Remarque Edmond J. , Radošević Katarina , Bogers Willy M. J. M. 
Published: November 2015

ABSTRACT

Influenza virus infection of nonhuman primates is a well-established animal model for studying pathogenesis and for evaluating prophylactic and therapeutic intervention strategies. However, usually a standard dose is used for the infection, and there is no information on the relation between challenge dose and virus replication or the induction of immune responses. Such information is also very scarce for humans and largely confined to evaluation of attenuated virus strains. Here, we have compared the effect of a commonly used dose (4 × 10
6
50% tissue culture infective doses) versus a 100-fold-higher dose, administered by intrabronchial installation, to two groups of 6 cynomolgus macaques. Animals infected with the high virus dose showed more fever and had higher peak levels of gamma interferon in the blood. However, virus replication in the trachea was not significantly different between the groups, although in 2 out of 6 animals from the high-dose group it was present at higher levels and for a longer duration. The virus-specific antibody response was not significantly different between the groups. However, antibody enzyme-linked immunosorbent assay, virus neutralization, and hemagglutination inhibition antibody titers correlated with cumulative virus production in the trachea. In conclusion, using influenza virus infection in cynomolgus macaques as a model, we demonstrated a relationship between the level of virus production upon infection and induction of functional antibody responses against the virus.

IMPORTANCE
There is only very limited information on the effect of virus inoculation dose on the level of virus production and the induction of adaptive immune responses in humans or nonhuman primates. We found only a marginal and variable effect of virus dose on virus production in the trachea but a significant effect on body temperature. The induction of functional antibody responses, including virus neutralization titer, hemagglutination inhibition titer, and antibody-dependent cell-mediated cytotoxicity, correlated with the level of virus replication measured in the trachea. The study reveals a relationship between virus production and functional antibody formation, which could be relevant in defining appropriate criteria for new influenza virus vaccine candidates.
Spore Germination
Authors: Moir Anne , Cooper Gareth 
Published: November 2015

ABSTRACT

Despite being resistant to a variety of environmental insults, the bacterial endospore can sense the presence of small molecules and respond by germinating, losing the specialized structures of the dormant spore, and resuming active metabolism, before outgrowing into vegetative cells. Our current level of understanding of the spore germination process in bacilli and clostridia is reviewed, with particular emphasis on the germinant receptors characterized in
Bacillus subtilis
,
Bacillus cereus
, and
Bacillus anthracis
. The recent evidence for a local clustering of receptors in a “germinosome” would begin to explain how signals from different receptors could be integrated. The SpoVA proteins, involved in the uptake of Ca
2+
-dipicolinic acid into the forespore during sporulation, are also responsible for its release during germination. Lytic enzymes SleB and CwlJ, found in bacilli and some clostridia, hydrolyze the spore cortex: other clostridia use SleC for this purpose. With genome sequencing has come the appreciation that there is considerable diversity in the setting for the germination machinery between bacilli and clostridia.
Exposure to secondhand smoke from neighbours and respiratory symptoms in never-smoking adolescents in Hong Kong: a cross-sectional study
Authors: Leung Lok Tung , Ho Sai Yin , Wang Man Ping , Lo Wing Sze , Lam Tai Hing 
Published: November 2015

ObjectivesTo investigate secondhand smoke (SHS) exposure at home from neighbours in Hong Kong adolescents and its association with respiratory symptoms in never-smokers.DesignA cross-sectional study.Setting79 randomly selected secondary schools in Hong Kong.Participants61 810 secondary 1 (USA grade 7) to 7 students, in which 50 762 never-smokers were identified and included in the analysis of the association between SHS exposure at home from neighbours and respiratory symptoms.Main outcome measuresSmoking status, family smoking status, SHS exposure at home from inside the home and from neighbours in the past 7 days, respiratory symptoms and sociodemographic characteristics were reported. Adjusted ORs (AORs) of respiratory symptoms for SHS exposure from the 2 sources in never-smokers were calculated using logistic regression.ResultsIn all students, 33.2% were exposed to SHS at home, including 16.2% from inside the home only, 10.0% from neighbours only and 7.0% from both. The prevalence of SHS exposure from neighbours was 17.1%, including 13.5% for 1–4 days/week and 3.6% for 5–7 days/week. In never-smokers (n=50 762), respiratory symptoms were significantly associated with SHS exposure from neighbours with AORs (95% CI) of 1.29 (1.20 to 1.39) for any exposure (p<0.001), 1.21 (1.12 to 1.31) for 1–4 days/week (p<0.001) and 1.63 (1.44 to 1.86) for 5–7 days/week (p<0.001) (P for trend <0.001). Compared with no SHS exposure at home from any source, the AORs were 1.16 (1.07 to 1.25) for SHS from inside the home only (p<0.001), 1.20 (1.11 to 1.31) from neighbours only (p<0.001), and 1.74 (1.56 to 1.94) from both (p<0.001).ConclusionsSHS exposure at home from neighbours was prevalent in Hong Kong adolescents, and was associated with respiratory symptoms in never-smokers. SHS exposure at home may be underestimated by ignoring the neighbouring source. Smoke-free housing policy is needed to protect children and adolescents from harms of SHS.