bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantitation of HLA Proteins Incorporated by Human Immunodeficiency Virus Type 1 and Assessment of Neutralizing Activity of Anti-HLA Antibodies
Authors: Lakhashe Samir K. , Thakar Madhuri R. , Bharucha K. E. , Paranjape Ramesh S.
Published: October 2007

Human anti-human leukocyte antigen (HLA) antibodies were assessed for neutralizing activity against human immunodeficiency virus type 1 (HIV-1) carrying HLA alleles with matching specificity. Multiparous women carrying anti-HLA antibodies were identified. Plasma samples from those women were confirmed as having antibodies that specifically bound to HLA proteins expressed on the peripheral blood mononuclear cells (PBMCs) of their husbands. A primary HIV-1 isolate was cultured in the husband's PBMCs so that the virus carried matching HLA alleles. To determine the HIV-1-neutralizing activity of anti-HLA antibodies, the infectivity of the virus for GHOST cells (which express green fluorescent protein after HIV infection) was investigated in the presence of a plasma sample positive for the respective anti-HLA antibody. A neutralization assay was also performed using purified immunoglobulin G (IgG) from two plasma samples, and two plasma samples were investigated in the presence of complement. The prerequisite for anti-HLA antibody-mediated neutralization is incorporation of HLA proteins by HIV-1. Therefore, the extent of incorporation of HLA proteins by the primary HIV-1 isolate was estimated. The ratios of HLA class I protein to HIV-1 capsid (p24) protein cultured in the PBMCs of two healthy individuals were 0.017 and 0.054. These ratios suggested that the HIV-1 strain used in the assay incorporated more HLA proteins than gp160 trimers. Anti-HLA antibody-positive plasma was found to contain antibodies that specifically reacted to HIV-1 carrying cognate HLA alleles. However, incubation of HIV-1 with anti-HLA antibody- positive plasma or purified IgG did not show a reduction in viral infectivity. HIV-1-neutralizing activity was also not detected in the presence of complement. This study shows that HIV-1 primary isolates cultured in PBMCs contain significant amounts of HLA proteins. However, the binding of antibodies to those HLA proteins does not mediate a reduction in viral infectivity.
Distinct RIG-I and MDA5 Signaling by RNA Viruses in Innate Immunity
Authors: Loo Yueh-Ming , Fornek Jamie , Crochet Nanette , Bajwa Gagan , Perwitasari Olivia , Martinez-Sobrido Luis , Akira Shizuo , Gill Michelle A. , García-Sastre Adolfo , Katze Michael G. , Gale Michael
Published: October 2007

Alpha/beta interferon immune defenses are essential for resistance to viruses and can be triggered through the actions of the cytoplasmic helicases retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5). Signaling by each is initiated by the recognition of viral products such as RNA and occurs through downstream interaction with the IPS-1 adaptor protein. We directly compared the innate immune signaling requirements of representative viruses of the
Flaviviridae
,
Orthomyxoviridae
,
Paramyxoviridae
, and
Reoviridae
for RIG-I, MDA5, and interferon promoter-stimulating factor 1 (IPS-1). In cultured fibroblasts, IPS-1 was essential for innate immune signaling of downstream interferon regulatory factor 3 activation and interferon-stimulated gene expression, but the requirements for RIG-I and MDA5 were variable. Each was individually dispensable for signaling triggered by reovirus and dengue virus, whereas RIG-I was essential for signaling by influenza A virus, influenza B virus, and human respiratory syncytial virus. Functional genomics analyses identified cellular genes triggered during influenza A virus infection whose expression was strictly dependent on RIG-I and which are involved in processes of innate or adaptive immunity, apoptosis, cytokine signaling, and inflammation associated with the host response to contemporary and pandemic strains of influenza virus. These results define IPS-1-dependent signaling as an essential feature of host immunity to RNA virus infection. Our observations further demonstrate differential and redundant roles for RIG-I and MDA5 in pathogen recognition and innate immune signaling that may reflect unique and shared biologic properties of RNA viruses whose differential triggering and control of gene expression may impact pathogenesis and infection.
Comparison of Plasma Viremia and Antibody Responses in Macaques Inoculated with Envelope Variants of Single-Cycle Simian Immunodeficiency Virus Differing in Infectivity and Cellular Tropism
Authors: DeGottardi M. Quinn , Lew Sharon K. , Piatak Michael , Jia Bin , Feng Yang , Lee Sandra J. , Brenchley Jason M. , Douek Daniel C. , Kodama Toshiaki , Lifson Jeffrey D. , Evans David T.
Published: October 2007

Molecular differences in the envelope glycoproteins of human immunodeficiency virus type 1 and simian immunodeficiency virus (SIV) determine virus infectivity and cellular tropism. To examine how these properties contribute to productive infection in vivo, rhesus macaques were inoculated with strains of single-cycle SIV (scSIV) engineered to express three different envelope glycoproteins with full-length (TM
open
) or truncated (TM
stop
) cytoplasmic tails. The 239 envelope uses CCR5 for infection of memory CD4
+
T cells, the 316 envelope also uses CCR5 but has enhanced infectivity for primary macrophages, and the 155T3 envelope uses CXCR4 for infection of both naive and memory CD4
+
T cells. Separate groups of six rhesus macaques were inoculated intravenously with mixtures of TM
open
and TM
stop
scSIV
mac
239, scSIV
mac
316, and scSIV
mac
155T3. A multiplex real-time PCR assay specific for unique sequence tags engineered into each virus was then used to measure viral loads for each strain independently. Viral loads in plasma peaked on day 4 for each strain and were resolved below the threshold of detection within 4 to 10 weeks. Truncation of the envelope cytoplasmic tail significantly increased the peak of viremia for all three envelope variants and the titer of SIV-specific antibody responses. Although peak viremias were similar for both R5- and X4-tropic viruses, clearance of scSIV
mac
155T3 TM
stop
was significantly delayed relative to the other strains, possibly reflecting the infection of a CXCR4
+
cell population that is less susceptible to the cytopathic effects of virus infection. These studies reveal differences in the peaks and durations of a single round of productive infection that reflect envelope-specific differences in infectivity, chemokine receptor specificity, and cellular tropism.
Novel Cytotoxic T-Lymphocyte Escape Mutation by a Three-Amino-Acid Insertion in the Human Immunodeficiency Virus Type 1 p6
Pol
and p6
Gag
Late Domain Associated with Drug Resistance
Authors: Cao Jianhong , McNevin John , McSweyn Matthew , Liu Yi , Mullins James I. , McElrath M. Juliana
Published: October 2007

Cytolytic T lymphocytes (CTL) play a major role in controlling human immunodeficiency virus type 1 (HIV-1) infection. To evade immune pressure, HIV-1 is selected at targeted CTL epitopes, which may consequentially alter viral replication fitness. In our longitudinal investigations of the interplay between T-cell immunity and viral evolution following acute HIV-1 infection, we observed in a treatment-naïve patient the emergence of highly avid, gamma interferon-secreting, CD8
+
CTL recognizing an HLA-Cw*0102-restricted epitope, NSPTRREL (NL8). This epitope lies in the p6
Pol
protein, located in the transframe region of the Gag-Pol polyprotein. Over the course of infection, an unusual viral escape mutation arose within the p6
Pol
epitope through insertion of a 3-amino-acid repeat, NSPT(SPT)RREL, with a concomitant insertion in the p6
Gag
late domain, PTAPP(APP). Interestingly, this p6
Pol
insertion mutation is often selected in viruses with the emergence of antiretroviral drug resistance, while the p6
Gag
late-domain PTAPP motif binds Tsg101 to permit viral budding. These results are the first to demonstrate viral evasion of immune pressure by amino acid insertions. Moreover, this escape mutation represents a novel mechanism whereby HIV-1 can alter its sequence within both the Gag and Pol proteins with potential functional consequences for viral replication and budding.
Protective Immunity toPseudomonas aeruginosaInduced with a Capsid-Modified Adenovirus ExpressingP. aeruginosaOprF
Authors: Worgall Stefan , Krause Anja , Qiu JianPing , Joh Ju , Hackett Neil R. , Crystal Ronald G.
Published: October 2007

This study focuses on the development of a new clinical vaccine candidate (AdOprF.RGD.Epi8) againstPseudomonas aeruginosausing an E1−E3−adenovirus (Ad) vector expressing OprF (AdOprF.RGD.Epi8) and modifications of the Ad genome providing two capsid changes: (i) modification of the Ad hexon gene to incorporate an immune-dominant OprF epitope (Epi8) into loop 1 of the hexon, enabling repeat administration to boost the anti-OprF immune response, and (ii) modification of the fiber gene to incorporate an integrin-binding RGD sequence to enhance gene delivery to antigen-presenting cells. Western analysis confirmed that AdOprF.RGD.Epi8 expresses OprF, contains Epi8 in the hexon protein, and enhances gene transfer to dendritic cells compared to AdOprF, a comparable Ad vector expressing OprF with an unmodified capsid. Intramuscular immunization of C57BL/6 mice with AdOprF.RGD.Epi8 resulted in the generation of anti-OprF antibodies at comparable levels to those induced following immunization with AdOprF, but immunization with AdOprF.RGD.Epi8 was associated with increased CD4 and CD8 gamma interferon T-cell responses against OprF as well as increased survival against lethal pulmonary challenge with agar-encapsulatedP. aeruginosa. Importantly, repeat administration of AdOprF.RGD.Epi8 resulted in boosting of the humoral anti-OprF response as well as increased protection, whereas no boosting could be achieved with repeat administration of AdOprF. This suggests that the capsid-modified AdOprF.RGD.Epi8 vector is a more effective immunogen compared to a comparable wild-type Ad capsid, making it a good candidate for an anti-P. aeruginosavaccine.
Mouse Cytomegalovirus MicroRNAs Dominate the Cellular Small RNA Profile during Lytic Infection and Show Features of Posttranscriptional Regulation
Authors: Dölken Lars , Perot Jonathan , Cognat Valérie , Alioua Abdelmalek , John Matthias , Soutschek Jürgen , Ruzsics Zsolt , Koszinowski Ulrich , Voinnet Olivier , Pfeffer Sébastien
Published: October 2007

MicroRNAs (miRNAs) are small, noncoding RNA molecules that regulate gene expression at the posttranscriptional level. Originally identified in a variety of organisms ranging from plants to mammals, miRNAs have recently been identified in several viruses. Viral miRNAs may play a role in modulating both viral and host gene expression. Here, we report on the identification and characterization of 18 viral miRNAs from mouse fibroblasts lytically infected with the murine cytomegalovirus (MCMV). The MCMV miRNAs are expressed at early times of infection and are scattered in small clusters throughout the genome with up to four distinct miRNAs processed from a single transcript. No significant homologies to human CMV-encoded miRNAs were found. Remarkably, as soon as 24 h after infection, MCMV miRNAs constituted about 35% of the total miRNA pool, and at 72 h postinfection, this proportion was increased to more than 60%. However, despite the abundance of viral miRNAs during the early phase of infection, the expression of some MCMV miRNAs appeared to be regulated. Hence, for three miRNAs we observed polyuridylation of their 3′ end, coupled to subsequent degradation. Individual knockout mutants of two of the most abundant MCMV miRNAs, miR-m01-4 and miR-M44-1, or a double knockout mutant of miR-m21-1 and miR-M23-2, incurred no or only a very mild growth deficit in murine embryonic fibroblasts in vitro.
Structure of the
Acidianus
Filamentous Virus 3 and Comparative Genomics of Related Archaeal Lipothrixviruses
Authors: Vestergaard Gisle , Aramayo Ricardo , Basta Tamara , Häring Monika , Peng Xu , Brügger Kim , Chen Lanming , Rachel Reinhard , Boisset Nicolas , Garrett Roger A. , Prangishvili David
Published: October 2007

Four novel filamentous viruses with double-stranded DNA genomes, namely,
Acidianus
filamentous virus 3 (AFV3), AFV6, AFV7, and AFV8, have been characterized from the hyperthermophilic archaeal genus
Acidianus
, and they are assigned to the
Betalipothrixvirus
genus of the family
Lipothrixviridae
. The structures of the approximately 2-μm-long virions are similar, and one of them, AFV3, was studied in detail. It consists of a cylindrical envelope containing globular subunits arranged in a helical formation that is unique for any known double-stranded DNA virus. The envelope is 3.1 nm thick and encases an inner core with two parallel rows of protein subunits arranged like a zipper. Each end of the virion is tapered and carries three short filaments. Two major structural proteins were identified as being common to all betalipothrixviruses. The viral genomes were sequenced and analyzed, and they reveal a high level of conservation in both gene content and gene order over large regions, with this similarity extending partly to the earlier described betalipothrixvirus
Sulfolobus islandicus
filamentous virus. A few predicted gene products of each virus, in addition to the structural proteins, could be assigned specific functions, including a putative helicase involved in Holliday junction branch migration, a nuclease, a protein phosphatase, transcriptional regulators, and glycosyltransferases. The AFV7 genome appears to have undergone intergenomic recombination with a large section of an AFV2-like viral genome, apparently resulting in phenotypic changes, as revealed by the presence of AFV2-like termini in the AFV7 virions. Shared features of the genomes include (i) large inverted terminal repeats exhibiting conserved, regularly spaced direct repeats; (ii) a highly conserved operon encoding the two major structural proteins; (iii) multiple overlapping open reading frames, which may be indicative of gene recoding; (iv) putative 12-bp genetic elements; and (v) partial gene sequences corresponding closely to spacer sequences of chromosomal repeat clusters.
Serum-Derived Hepatitis C Virus Infection of Primary Human Hepatocytes Is Tetraspanin CD81 Dependent
Authors: Molina Sonia , Castet Valerie , Pichard-Garcia Lydiane , Wychowski Czeslaw , Meurs Eliane , Pascussi Jean-Marc , Sureau Camille , Fabre Jean-Michel , SaCunha Antonio , Larrey Dominique , Dubuisson Jean , Coste Joliette , McKeating Jane , Maurel Patrick , Fournier-Wirth Chantal
Published: October 2007

Hepatitis C virus-positive serum (HCVser, genotypes 1a to 3a) or HCV cell culture (JFH1/HCVcc) infection of primary normal human hepatocytes was assessed by measuring intracellular HCV RNA strands. Anti-CD81 antibodies and siRNA-CD81 silencing markedly inhibited (>90%) HCVser infection irrespective of HCV genotype, viral load, or liver donor, while hCD81-large intracellular loop (LEL) had no effect. However, JFH1/HCVcc infection of hepatocytes was modestly inhibited (40 to 60%) by both hCD81-LEL and anti-CD81 antibodies. In conclusion, CD81 is involved in HCVser infection of human hepatocytes, and comparative studies of HCVser versus JFH1/HCVcc infection of human hepatocytes and Huh-7.5 cells revealed that the cell-virion combination is determinant of the entry process.
Rational Design of a Multiepitope Vaccine Encoding T-Lymphocyte Epitopes for Treatment of Chronic Hepatitis B Virus Infections
Authors: Depla Erik , Van der Aa Annegret , Livingston Brian D. , Crimi Claire , Allosery Koen , De Brabandere Veronique , Krakover Jonathan , Murthy Sidharta , Huang Manley , Power Scott , Babé Lilia , Dahlberg Carol , McKinney Denise , Sette Alessandro , Southwood Scott , Philip Ramilla , Newman Mark J. , Meheus Lydie
Published: October 2007

Protein sequences from multiple hepatitis B virus (HBV) isolates were analyzed for the presence of amino acid motifs characteristic of cytotoxic T-lymphocyte (CTL) and helper T-lymphocyte (HTL) epitopes with the goal of identifying conserved epitopes suitable for use in a therapeutic vaccine. Specifically, sequences bearing HLA-A1, -A2, -A3, -A24, -B7, and -DR supertype binding motifs were identified, synthesized as peptides, and tested for binding to soluble HLA. The immunogenicity of peptides that bound with moderate to high affinity subsequently was assessed using HLA transgenic mice (CTL) and HLA cross-reacting H-2bxd(BALB/c × C57BL/6J) mice (HTL). Through this process, 30 CTL and 16 HTL epitopes were selected as a set that would be the most useful for vaccine design, based on epitope conservation among HBV sequences and HLA-based predicted population coverage in diverse ethnic groups. A plasmid DNA-based vaccine encoding the epitopes as a single gene product, with each epitope separated by spacer residues to enhance appropriate epitope processing, was designed. Immunogenicity testing in mice demonstrated the induction of multiple CTL and HTL responses. Furthermore, as a complementary approach, mass spectrometry allowed the identification of correctly processed and major histocompatibility complex-presented epitopes from human cells transfected with the DNA plasmid. A heterologous prime-boost immunization with the plasmid DNA and a recombinant MVA gave further enhancement of the immune responses. Thus, a multiepitope therapeutic vaccine candidate capable of stimulating those cellular immune responses thought to be essential for controlling and clearing HBV infection was successfully designed and evaluated in vitro and in HLA transgenic mice.
Lymphocytoid Choriomeningitis Virus Activates Plasmacytoid Dendritic Cells and Induces a Cytotoxic T-Cell Response via MyD88
Authors: Jung Andreas , Kato Hiroki , Kumagai Yutaro , Kumar Himanshu , Kawai Taro , Takeuchi Osamu , Akira Shizuo
Published: October 2007

Toll-like receptors (TLRs) and retinoic acid-inducible gene I-like helicases (RLHs) are two major machineries recognizing RNA virus infection of innate immune cells. Intracellular signaling for TLRs and RLHs is mediated by their cytoplasmic adaptors, i.e., MyD88 or TRIF and IPS-1, respectively. In the present study, we investigated the contributions of TLRs and RLHs to the cytotoxic T-lymphocyte (CTL) response by using lymphocytoid choriomeningitis virus (LCMV) as a model virus. The generation of virus-specific cytotoxic T lymphocytes was critically dependent on MyD88 but not on IPS-1. Type I interferons (IFNs) are known to be important for the development of the CTL response to LCMV infection. Serum levels of type I IFNs and proinflammatory cytokines were mainly dependent on the presence of MyD88, although IPS-1
−/−
mice showed a decrease in IFN-α levels but not in IFN-β and proinflammatory cytokine levels. Analysis of
Ifna6
+/GFP
reporter mice revealed that plasmacytoid dendritic cells (DCs) are the major source of IFN-α in LCMV infection. MyD88
−/−
mice were highly susceptible to LCMV infection in vivo. These results suggest that recognition of LCMV by plasmacytoid DCs via TLRs is responsible for the production of type I IFNs in vivo. Furthermore, the activation of a MyD88-dependent innate mechanism induces a CTL response, which eventually leads to virus elimination.
Structural Abnormalities in Neurons Are Sufficient To Explain the Clinical Disease and Fatal Outcome of Experimental Rabies in Yellow Fluorescent Protein-Expressing Transgenic Mice
Authors: Scott Courtney A. , Rossiter John P. , Andrew R. David , Jackson Alan C.
Published: October 2007

Under natural conditions and in some experimental models, rabies virus infection of the central nervous system causes relatively mild histopathological changes, without prominent evidence of neuronal death despite its lethality. In this study, the effects of rabies virus infection on the structure of neurons were investigated with experimentally infected transgenic mice expressing yellow fluorescent protein (YFP) in neuronal subpopulations. Six-week-old mice were inoculated in the hind-limb footpad with the CVS strain of fixed virus or were mock infected with vehicle (phosphate-buffered saline). Brain regions were subsequently examined by light, epifluorescent, and electron microscopy. In moribund CVS-infected mice, histopathological changes were minimal in paraffin-embedded tissue sections, although mild inflammatory changes were present. Terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling and caspase-3 immunostaining showed only a few apoptotic cells in the cerebral cortex and hippocampus. Silver staining demonstrated the preservation of cytoskeletal integrity in the cerebral cortex. However, fluorescence microscopy revealed marked beading and fragmentation of the dendrites and axons of layer V pyramidal neurons in the cerebral cortex, cerebellar mossy fibers, and axons in brainstem tracts. At an earlier time point, when mice displayed hind-limb paralysis, beading was observed in a few axons in the cerebellar commissure. Toluidine blue-stained resin-embedded sections from moribund YFP-expressing animals revealed vacuoles within the perikarya and proximal dendrites of pyramidal neurons in the cerebral cortex and hippocampus. These vacuoles corresponded with swollen mitochondria under electron microscopy. Vacuolation was also observed ultrastructurally in axons and in presynaptic nerve endings. We conclude that the observed structural changes are sufficient to explain the severe clinical disease with a fatal outcome in this experimental model of rabies.
Identification of Hexon-Specific CD4 and CD8 T-Cell Epitopes for Vaccine and Immunotherapy
Authors: Leen Ann M. , Christin Anne , Khalil Mariam , Weiss Heidi , Gee Adrian P. , Brenner Malcolm K. , Heslop Helen E. , Rooney Cliona M. , Bollard Catherine M.
Published: October 2007

Adenoviral infections in the immunocompromised host are associated with significant morbidity and mortality. Although the adoptive transfer of adenovirus-specific T cells may prevent and treat such infections, the T-cell immune response to the multiplicity of adenovirus serotypes and subspecies that infect humans has not been well characterized, impeding the development of such approaches. We have, therefore, analyzed the specificities of T-cell responses to the viral capsid hexon antigen, since this structure is highly conserved in human pathogens. We screened 25 human cytotoxic T-cell lines with adenovirus specificity to extensively characterize their responses to adenoviral hexon and to identify a panel of novel CD4+and CD8+T-cell epitopes. Using a peptide library spanning the entire sequence of the hexon protein, we confirmed the responsiveness of these cytotoxic T-cell lines to seven peptides described previously and also identified 33 new CD4- or CD8-restricted hexon epitopes. Importantly, the majority of these epitopes were shared among different adenovirus subspecies, suggesting that T cells with such specificities could recognize and be protective against multiple serotypes, simplifying the task of effective adoptive transfer or vaccine-based immunotherapy for treating infection by this virus.
Heptad Repeat-Derived Peptides Block Protease-Mediated Direct Entry from the Cell Surface of Severe Acute Respiratory Syndrome Coronavirus but Not Entry via the Endosomal Pathway
Authors: Ujike Makoto , Nishikawa Hiroki , Otaka Akira , Yamamoto Naoki , Yamamoto Norio , Matsuoka Masao , Kodama Eiichi , Fujii Nobutaka , Taguchi Fumihiro
Published: October 2007

The peptides derived from the heptad repeat (HRP) of severe acute respiratory syndrome coronavirus (SCoV) spike protein (sHRPs) are known to inhibit SCoV infection, yet their efficacies are fairly low. Recently our research showed that some proteases facilitated SCoV's direct entry from the cell surface, resulting in a more efficient infection than the previously known infection via endosomal entry. To compare the inhibitory effect of the sHRP in each pathway, we selected two sHRPs, which showed a strong inhibitory effect on the interaction of two heptad repeats in a rapid and virus-free in vitro assay system. We found that they efficiently inhibited SCoV infection of the protease-mediated cell surface pathway but had little effect on the endosomal pathway. This finding suggests that sHRPs may effectively prevent infection in the lungs, where SCoV infection could be enhanced by proteases produced in this organ. This is the first observation that HRP exhibits different effects on virus that takes the endosomal pathway and virus that enters directly from the cell surface.
Altered Pathogenesis of Porcine Respiratory Coronavirus in Pigs due to Immunosuppressive Effects of Dexamethasone: Implications for Corticosteroid Use in Treatment of Severe Acute Respiratory Syndrome Coronavirus
Authors: Jung Kwonil , Alekseev Konstantin P. , Zhang Xinsheng , Cheon Doo-Sung , Vlasova Anastasia N. , Saif Linda J.
Published: October 2007

The pathogenesis and optimal treatments for severe acute respiratory syndrome (SARS) are unclear, although corticosteroids were used to reduce lung and systemic inflammation. Because the pulmonary pathology of porcine respiratory coronavirus (PRCV) in pigs resembles SARS, we used PRCV as a model to clarify the effects of the corticosteroid dexamethasone (DEX) on coronavirus (CoV)-induced pneumonia. Conventional weaned pigs (n= 130) in one of four groups (PRCV/phosphate-buffered saline [PBS] [n= 41], PRCV/DEX [n= 41], mock/PBS [n= 23], and mock/DEX [n= 25]) were inoculated intranasally and intratracheally with the ISU-1 strain of PRCV (1 × 107PFU) or cell culture medium. DEX was administered (once daily, 2 mg/kg of body weight/day, intramuscularly) from postinoculation day (PID) 1 to 6. In PRCV/DEX pigs, significantly milder pneumonia, fewer PRCV-positive cells, and lower viral RNA titers were present in lungs early at PID 2; however, at PID 4, 10, and 21, severe bronchointerstitial pneumonia, significantly higher numbers of PRCV-positive cells, and higher viral RNA titers were observed compared to results for PRCV/PBS pigs. Significantly lower numbers of CD2+, CD3+, CD4+, and CD8+T cells were also observed in lungs of PRCV/DEX pigs than in those of PRCV/PBS pigs at PID 8 and 10, coincident with fewer gamma interferon (IFN-γ)-secreting cells in the tracheobronchial lymph nodes as determined by enzyme-linked immunospot assay. Our results confirm that DEX treatment alleviates PRCV pneumonia early (PID 2) in the infection but continued use through PID 6 exacerbates later stages of infection (PID 4, 10, and 21), possibly by decreasing cellular immune responses in the lungs (IFN-γ-secreting T cells), thereby creating an environment for more-extensive viral replication. These data have potential implications for corticosteroid use with SARS-CoV patients and suggest a precaution against prolonged use based on their unproven efficacy in humans, including possible detrimental secondary effects.
Antiretroviral Activity and Vif Sensitivity of Rhesus Macaque APOBEC3 Proteins
Authors: Virgen Cesar A. , Hatziioannou Theodora
Published: October 2007

The inability of human immunodeficiency virus type 1(HIV-1) to replicate in rhesus macaque cells is in part due to the failure of HIV-1 Vif to counteract the restriction factor APOBEC3G. However, in this study we demonstrate that several rhesus macaque APOBEC3 (rhAPOBEC3) proteins are capable of inhibiting HIV-1 infectivity. There was considerable variation in the ability of a panel of Vif proteins to induce degradation of rhAPOBEC3 proteins, and mutations within HIV-1 Vif that render it capable of degrading rhAPOBEC3G did not confer activity against other antiviral rhAPOBEC3 proteins. These findings suggest that multiple APOBEC3 proteins can contribute to primate lentivirus species tropism.
Binding of Human Papillomavirus Type 16 E6 to E6AP Is Not Required for Activation of hTERT
Authors: Sekaric Pedja , Cherry Jonathan J. , Androphy Elliot J.
Published: October 2007

The human papillomavirus (HPV) type 16 (HPV16) E6 protein stimulates transcription of the catalytic subunit of telomerase, hTERT, in epithelial cells. It has been reported that binding to the ubiquitin ligase E6AP is required for this E6 activity, with E6 directing E6AP to the hTERT promoter. We previously reported two E6AP binding-defective HPV16 E6 mutations that induced immortalization of human mammary epithelial cells. Because activation of hTERT is proposed to be necessary for epithelial cell immortalization, we sought to further characterize the relationship between E6/E6AP association and telomerase induction. We demonstrate that while these E6 mutants do not bind E6AP, they retain the capability to stimulate the expression of hTERT. Chromatin immunoprecipitation assays confirmed the presence of Myc, wild-type E6, and the E6AP binding-defective E6 mutants, but not E6AP itself, at the endogenous hTERT promoter. Interestingly, an immortalization-defective E6 mutant localized to the hTERT promoter but failed to increase transcription. We conclude that binding to E6AP is not necessary for E6 localization to or activation of the hTERT promoter and that another activity of E6 is involved in hTERT activation.
Crystal Structure of the NS3 Protease-Helicase from Dengue Virus
Authors: Luo Dahai , Xu Ting , Hunke Cornelia , Grüber Gerhard , Vasudevan Subhash G. , Lescar Julien
Published: October 2007

Several flaviviruses are important human pathogens, including dengue virus, a disease against which neither a vaccine nor specific antiviral therapies currently exist. During infection, the flavivirus RNA genome is translated into a polyprotein, which is cleaved into several components. Nonstructural protein 3 (NS3) carries out enzymatic reactions essential for viral replication, including proteolysis of the polyprotein through its serine protease N-terminal domain, with a segment of 40 residues from the NS2B protein acting as a cofactor. The ATPase/helicase domain is located at the C terminus of NS3. Atomic structures are available for these domains separately, but a molecular view of the full-length flavivirus NS3 polypeptide is still lacking. We report a crystallographic structure of a complete NS3 molecule fused to 18 residues of the NS2B cofactor at a resolution of 3.15 Å. The relative orientation between the protease and helicase domains is drastically different than the single-chain NS3-NS4A molecule from hepatitis C virus, which was caught in the act ofciscleavage at the NS3-NS4A junction. Here, the protease domain sits beneath the ATP binding site, giving the molecule an elongated shape. The domain arrangement found in the crystal structure fits nicely into an envelope determined ab initio using small-angle X-ray scattering experiments in solution, suggesting a stable molecular conformation. We propose that a basic patch located at the surface of the protease domain increases the affinity for nucleotides and could also participate in RNA binding, explaining the higher unwinding activity of the full-length enzyme compared to that of the isolated helicase domain.
The Human T-Cell Leukemia Virus Type 1 Tax Oncoprotein Requires the Ubiquitin-Conjugating Enzyme Ubc13 for NF-κB Activation
Authors: Shembade Noula , Harhaj Nicole S. , Yamamoto Masahiro , Akira Shizuo , Harhaj Edward W.
Published: October 2007

Ubiquitination of the human T-cell leukemia virus 1 Tax oncoprotein provides an important regulatory mechanism that promotes the Tax-mediated activation of NF-κB. However, the type of polyubiquitin chain linkages and the host factors that are required for Tax ubiquitination have not been identified. Here, we demonstrate that Tax polyubiquitin chains are composed predominantly of lysine 63-linked chains. Furthermore, the ubiquitination of Tax is critically dependent on the E2 ubiquitin-conjugating enzyme Ubc13. Tax interacts with Ubc13, and small interfering RNA-mediated knockdown of Ubc13 expression abrogates Tax ubiquitination and the activation of NF-κB. Mouse fibroblasts lacking Ubc13 exhibit impaired Tax activation of NF-κB despite normal tumor necrosis factor- and interleukin-1-mediated NF-κB activation. Finally, the interaction of Tax with NEMO is disrupted in the absence of Tax ubiquitination and Ubc13 expression, suggesting that Tax ubiquitination is critical for NEMO binding. Collectively, our results reveal that Ubc13 is essential for Tax ubiquitination, its interaction with NEMO, and Tax-mediated NF-κB activation.
Characterization of the Human Cytomegalovirus gH/gL/UL128-131 Complex That Mediates Entry into Epithelial and Endothelial Cells
Authors: Ryckman Brent J. , Rainish Barb L. , Chase Marie C. , Borton Jamie A. , Nelson Jay A. , Jarvis Michael A. , Johnson David C.
Published: October 2007

The entry of human cytomegalovirus (HCMV) into biologically relevant epithelial and endothelial cells involves endocytosis followed by low-pH-dependent fusion. This entry pathway is facilitated by the HCMV UL128, UL130, and UL131 proteins, which form one or more complexes with the virion envelope glycoprotein gH/gL. gH/gL/UL128-131 complexes appear to be distinct from the gH/gL/gO complex, which likely facilitates entry into fibroblasts. In order to better understand the assembly and protein-protein interactions of gH/gL/UL128-131 complexes, we generated HCMV mutants lacking UL128-131 proteins and nonreplicating adenovirus vectors expressing gH, gL, UL128, UL130, and UL131. Our results demonstrate that UL128, UL130, and UL131 can each independently assemble onto gH/gL scaffolds. However, the binding of individual UL128-131 proteins onto gH/gL can significantly affect the binding of other proteins; for example, UL128 increased the binding of both UL130 and UL131 to gH/gL. Direct interactions between gH/UL130, UL130/UL131, gL/UL128, and UL128/UL130 were also observed. The export of gH/gL complexes from the endoplasmic reticulum (ER) to the Golgi apparatus and cell surface was dramatically increased when all of UL128, UL130, and UL131 were coexpressed with gH/gL (with or without gO expression). Incorporation of gH/gL complexes into the virion envelope requires transport beyond the ER. Thus, we concluded that UL128, UL130, and UL131 must all bind simultaneously onto gH/gL for the production of complexes that can function in entry into epithelial and endothelial cells.
SUMMARY OF WORKING GROUP 1: ELECTRON SOURCES
Authors: FERRARIO MASSIMO , GATTI GIANCARLO
Published: October 2007

This report summarizes the presentations and discussions in the Working Group "Electron Sources" during the ICFA Advanced Accelerator Workshop on "The Physics and Applications of High-Brightness Electron Beams", held at the Ettore Majorana Foundation and Center for Scientific Culture, Erice, October 9-14, 2005. Latest developments towards a better understanding of high-brightness photo injectors physics and technology are reported.
DESIGN AND RF MEASUREMENTS OF AN X-BAND ACCELERATING STRUCTURE FOR THE SPARC PROJECT
Authors: Alesini D. , Bacci A. , Falone A. , Migliorati M. , Mostacci A. , Palpini F. , Palumbo L. , Spataro B.
Published: October 2007

The paper presents the design of an X-band accelerating section for linearizing the longitudinal phase space in the Frascati Linac Coherent Light Source (SPARC). The nine cells structure, operating on the π standing wave mode, is fed by a central coupler and has been designed to obtain a 42 MV/m accelerating gradient. The 2D profile has been obtained using the electromagnetic codes Superfish and Oscar2D while the coupler has been designed using HFSS. Bead-pull measurements made on a copper prototype have been performed and the results are illustrated and compared with the numerical predictions.
RF GUN PHOTO-EMISSION MODEL FOR METAL CATHODES INCLUDING TIME DEPENDENT EMISSION
Authors: SCHMERGE J. F. , CLENDENIN J. E. , DOWELL D. H. , GIERMAN S. M.
Published: October 2007

The quantum efficiency from a metal cathode is strongly dependent on the field at the cathode due to the Schottky effect. Since the field is time dependent the quantum efficiency is also time dependent. Thus the laser pulse shape used to generate electrons in a photocathode rf gun is not the same as the electron bunch shape. In addition since the thermal emittance and quantum efficiency are related, the thermal emittance is also time dependent.
SUPERCONDUCTING PHOTOCATHODES
Authors: SMEDLEY J. , RAO T. , WARREN J. , SEKUTOWICZ J. , LANGNER J. , STRZYŻEWSKI P. , LEFFERTS R. , LIPSKI A.
Published: October 2007

We present the results of our investigation of lead and niobium as suitable photocathode materials for superconducting RF injectors. Quantum efficiencies (QE) have been measured for a range of incident photon energies and a variety of cathode preparation methods, including various lead plating techniques on a niobium substrate. The effects of operating at ambient and cryogenic temperatures and different vacuum levels on the cathode QE have also been studied.
OBSERVATION OF COHERENT EDGE RADIATION EMITTED BY A 100 FEMTOSECOND COMPRESSED ELECTRON BEAM
Authors: ANDONIAN G. , DUNNING M. , HEMSING E. , ROSENZWEIG J. B. , COOK A. , MUROKH A. , REICHE S. , BABZIEN M. , BEN-ZVI I. , KUSCHE K. , YAKIMENKO V. , ALESINI D. , PALUMBO L. , VICARIO C.
Published: October 2007

A chicane compressor developed by UCLA for the production of ultra-short, 60 MeV electron beams at the Brookhaven National Laboratory Accelerator Test Facility has been commissioned, and initial beam physics experiments have been performed. These measurements have established the compression of electron beams to the 100 femtosecond (1 kA peak current) regime, via coherent transition radiation (CTR) based measurements. Investigations of coherent edge radiation (CER) include signatures that differentiate it from coherent synchrotron radiation (CSR), such as polarization and far-field angular distribution. Additionally, the radiation wavelength spectrum is determined from autocorrelation measurements. Radiation properties are compared to detailed start-to-end simulations derived from PARMELA and QUINDI (a Lienard-Wiechert code developed at UCLA). Plans for future experiments which further explore the observed wavelength spectra are presented.
PARMELA SIMULATIONS FOR PITZ: FIRST MACHINE STUDIES AND INTERPRETATION OF MEASUREMENTS
Authors: BOSCOLO M. , FERRARIO M. , CIALDI S. , KRASILNIKOV M. , OPPELT A.
Published: October 2007

First comparisons between measurements and simulations are discussed for the PITZ1.5 facility. In particular, dedicated PARMELA simulations performed using the realistic laser pulse hitting the cathode are compared to first emittance measurements performed with the SPARC e -meter and with the EMSY system. Measured energy spectra are compared to simulations, as well. It is discussed the overall good agreement between calculations and measurements.
THE LCLS SINGLE-SHOT RELATIVE BUNCH LENGTH MONITOR SYSTEM
Authors: DUNNING M. P. , TRAVISH G. , COHEN A. , FRIGOLA P. , REICHE S. , ROSENZWEIG J.
Published: October 2007

The Particle Beam Physics Lab (PBPL) at UCLA is developing a single-shot relative bunch length monitor system for the Linac Coherent Light Source (LCLS) X-ray Free Electron Laser at SLAC. To ensure high-quality lasing, feedback systems are needed. The bunch length monitor system will be a crucial part of the linac longitudinal feedback system, which is necessary to keep the final peak beam current stable. The system will consist of two single-channel detector arrangements, one after each bunch compressor, and will use the coherent synchrotron radiation (CSR) and coherent edge radiation (CER) from bunch compressor chicane dipole magnets to monitor the bunch length. Details of the system are presented, with an emphasis on the device for bunch compressor one (BC1), and the system is compared to alternatives. Plans for testing are presented as well.
BEAM SHAPING AND PERMANENT MAGNET QUADRUPOLE FOCUSING WITH APPLICATIONS TO THE PLASMA WAKEFIELD ACCELERATOR
Authors: ENGLAND R. J. , ROSENZWEIG J. B. , TRAVISH G. , DOYURAN A. , WILLIAMS O. , O'SHEA B. , ALESINI D.
Published: October 2007

It has recently been proposed to use a dispersionless translating section (dogleg) with sextupole correction magnets as a bunch compressor to create longitudinally shaped (linearly ramped) electron bunches. We discuss the experiment soon to be underway at the UCLA Neptune Linear Accelerator Laboratory to test this technique with the 300 pC, 13 MeV electron bunches produced by the Neptune S -Band photoinjector. The experiment will utilize a dipole-mode deflecting cavity, as a temporal diagnostic, and a final focus system of permanent magnet quadrupoles with field gradients of 110 T/m. We also discuss the potential scaling of this technique to bunches of high (i.e. >1nC) charge for the purpose of creating a suitable drive beam for the plasma wakefield accelerator, operating in the blowout regime.
COMMISSIONING OF THE SPARC MOVABLE EMITTANCE METER AND ITS FIRST OPERATION AT PITZ
Authors: Filippetto D. , Castellano M. , Di Pirro G. , Vicario C. , Catani L. , Chiadroni E. , Cianchi A. , Hartrott M. V. , Oppelt A. , Stephan F. , Krasilnikov M.
Published: October 2007

For the SPARC Project a novel diagnostic device, called "Emittance-meter", has been conceived and constructed to perform a detailed study of the emittance compensation process in the SPARC photo-injector and to optimize the RF-gun and the accelerator working point. It consists of a movable emittance measurement system, based on the 1D pepper-pot method, installed between two long bellows with the possibility to scan a region 1.2 m long downstream the RF-gun. The construction of the device was completed in the first part of this year and a series of laboratory tests, to evaluate its performances, were carried out in Spring 2005. At the beginning of the summer the complete system was moved to DESY at Zeuthen to be installed on the Photo Injector Test Facility PITZ. After the commissioning it will used for measurements of the PITZ electron beam in the framework of a collaboration between the SPARC and PITZ Projects aiming on studies and operations with photo injectors.
SYNCHRONIZATION BETWEEN THE LASER AND ELECTRON BEAM IN A PHOTOCATHODE RF GUN
Authors: SAKUMI A. , IIJIMA H. , UESAKA M. , YOSHII K. , UEDA T. , MUROYA Y. , FUKASAWA A. , KUMAGAI N. , TOMIZAWA H. , URAKAWA J.
Published: October 2007

The chemical reactions of hot or room temperature and/or critical water in a time-range of picosecond and sub-picoseconds have been carried out by the 18 MeV S -band linac and a Mg photocathode RF gun with the irradiation of third harmonic Ti : Sapphire laser, at Nuclear Engineering Research Laboratory (NERL), the University of Tokyo. Although this short bunch and 100 fs laser light are enough to perform the experiment of radiation chemistry in the time-range of sub-picoseconds, the total time-resolution become worse by the instability of synchronization between laser and radio frequency of linac. We found that the fluctuation of room temperature causes the instability, particularly the cycle of turning on/off of the air-conditioner. When we decrease the fraction of the room temperature within 0.1 degrees, the timing drift in an hour reaches 600 fs, closed to the timing jitter of 340 fs than previous results.
SUMMARY OF WORKING GROUP 3: NEW CHALLENGES IN THEORY AND MODELING
Authors: GIANNESSI LUCA
Published: October 2007

Working group three was devoted to contributions on theory and modeling of devices and new ideas, aiming at increasing the beam brightness and at generating radiation at short wavelengths. The contributions presented span from the beam dynamics in radiofrequency accelerators, emittance compensation schemes and optimization of the beam brightness at low energy, analysis of the emittance dilution processes during acceleration and compression, laser plasma sources and the generation of radiation in FELs and other radiation sources. Analysis specifically oriented to the modeling of the beam and radiation dynamics were also presented.
RESONANT MODES IN A 1.6 CELLS RF GUN
Authors: FERRARIO MASSIMO , RONSIVALLE CONCETTA
Published: October 2007

The SPARC photoinjector RF gun consists in the BNL/SLAC/UCLA 1.6 cell structure designed to resonate at 2856 MHz in the π mode. It will be demonstrated by a numerical modelization based on SUPERFISH code combined with the LC-circuit analysis that the two oscillating modes of the system usually indicated as 0-mode and π-mode (the operating mode) are in reality a π/3-mode-like and a π-mode-like. The consequences on the definition of the coupling coefficient and on the use of mode-separation based RF measurements are described.
SIMULATIONS OF COHERENT SYNCHROTRON RADIATION EFFECTS IN ELECTRON MACHINES
Authors: MIGLIORATI M. , SCHIAVI A. , DATTOLI G.
Published: October 2007

Coherent synchrotron radiation (CSR) generated by high intensity electron beams can be a source of undesirable effects limiting the performance of storage rings. The complexity of the physical mechanisms underlying the interplay between the electron beam and the CSR demands for reliable simulation codes. In the past, codes based on Lie algebraic techniques have been very efficient to treat transport problems in accelerators. The extension of these methods to the non linear case is ideally suited to treat wakefields - beam interaction. In this paper we report on the development of a numerical code, based on the solution of the Vlasov equation, which includes the non linear contribution due to wakefields. The proposed solution method exploits an algebraic technique that uses the exponential operators. We show that, in the case of CSR wakefields, the integration procedure is capable of reproducing the onset of an instability which leads to microbunching of the beam thus increasing the CSR at short wavelengths. In addition, considerations on the threshold of the instability for Gaussian bunches is also reported.
QFEL: A NUMERICAL CODE FOR MULTI-DIMENSIONAL SIMULATION OF FREE ELECTRON LASERS IN THE QUANTUM REGIME
Authors: SCHIAVI A. , PIOVELLA N. , ROBB G. R. M. , BONIFACIO R.
Published: October 2007

A new simulation tool has been developed for the simulation of the FEL equations in both the classical and quantum regimes to be applied to the investigation of proposed FEL models and to the exploration of parameter space for new experiments.
FIRST SIMULATIONS RESULTS ON LASER PULSE JITTER AND MICROBUNCHING INSTABILITY AT SPARXINO
Authors: BOSCOLO M. , FERRARIO M. , FUSCO V. , MIGLIORATI M. , PALUMBO L. , SPATARO B. , VACCAREZZA C. , GIANNESSI L. , QUATTROMINI M. , RONSIVALLE C. , SERAFINI L.
Published: October 2007

In the frame of the R&D activity foreseen for the first phase of the SPARX project, the SPARXINO test facility is meant to provide an ultra-high electron beam at 1.2-1.5 GeV, able to drive SASE and seeded FEL experiments in the 10-3 nm range. The proposed layout is analyzed regarding its sensitivity to laser pulse jitter effects and microbunching instability limitations. First simulation results are presented.
STUDY OF TRANSVERSE EFFECTS IN THE PRODUCTION OF X-RAYS WITH A FREE-ELECTRON LASER BASED ON AN OPTICAL UNDULATOR
Authors: BACCI A. , MAROLI C. , PETRILLO V. , SERAFNI L. , FERRARIO M.
Published: October 2007

The interaction between high-brilliance electron beams and counter-propagating laser pulses produces X rays via Thomson back-scattering. If the laser source is long and intense enough, the electrons of the beam can bunch and a regime of collective effects can establish. In this case of dominating collective effects, the FEL instability can develop and the system behaves like a free-electron laser based on an optical undulator. Coherent X-rays can be irradiated, with a bandwidth very much thinner than that of the corresponding incoherent emission. The emittance of the electron beam and the distribution of the laser energy are the principal quantities that limit the growth of the X-ray signal. In this work we analyse with a 3-D code the transverse effects in the emission produced by a relativistic electron beam when it is under the action of an optical laser pulse and the X-ray spectra obtained. The scalings typical of the optical wiggler, characterized by very short gain lengths and overall time durations of the process make possible considerable emission also with emittance of the order of 1mm mrad.