bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Association of Recombination Events in the Founding and Emergence of Subgenogroup Evolutionary Lineages of Human Enterovirus 71
Authors: McWilliam Leitch E. C. , Cabrerizo M. , Cardosa J. , Harvala H. , Ivanova O. E. , Koike S. , Kroes A. C. M. , Lukashev A. , Perera D. , Roivainen M. , Susi P. , Trallero G. , Evans D. J. , Simmonds P. 
Published: December 2011

ABSTRACT

Enterovirus 71 (EV71) is responsible for frequent large-scale outbreaks of hand, foot, and mouth disease worldwide and represent a major etiological agent of severe, sometimes fatal neurological disease. EV71 variants have been classified into three genogroups (GgA, GgB, and GgC), and the latter two are further subdivided into subgenogroups B1 to B5 and C1 to C5. To investigate the dual roles of recombination and evolution in the epidemiology and transmission of EV71 worldwide, we performed a large-scale genetic analysis of isolates (
n
= 308) collected from 19 countries worldwide over a 40-year period. A series of recombination events occurred over this period, which have been identified through incongruities in sequence grouping between the VP1 and 3Dpol regions. Eleven 3Dpol clades were identified, each specific to EV71 and associated with specific subgenogroups but interspersed phylogenetically with clades of coxsackievirus A16 and other EV species A serotypes. The likelihood of recombination increased with VP1 sequence divergence; mean half-lives for EV71 recombinant forms (RFs) of 6 and 9 years for GgB and GgC overlapped with those observed for the EV-B serotypes, echovirus 9 (E9), E30, and E11, respectively (1.3 to 9.8 years). Furthermore, within genogroups, sporadic recombination events occurred, such as the linkage of two B4 variants to RF-W instead of RF-A and of two C4 variants to RF-H. Intriguingly, recombination events occurred as a founding event of most subgenogroups immediately preceding their lineage expansion and global emergence. The possibility that recombination contributed to their subsequent spread through improved fitness requires further biological and immunological characterization.
DNA-Binding Activity of Adeno-Associated Virus Rep Is Required for Inverted Terminal Repeat-Dependent Complex Formation with Herpes Simplex Virus ICP8
Authors: Alex Martin , Weger Stefan , Mietzsch Mario , Slanina Heiko , Cathomen Toni , Heilbronn Regine 
Published: December 2011

ABSTRACT

Herpes simplex virus (HSV) helper functions for (AAV) replication comprise HSV ICP8 and helicase-primase UL5/UL52/UL8. Here we show that N-terminal amino acids of AAV Rep78 that contact the Rep-binding site within the AAV inverted terminal repeat (ITR) are required for ternary-complex formation with infected-cell protein 8 (ICP8) on AAV single-strand DNA (ssDNA)
in vitro
and for colocalization in nuclear replication domains
in vivo
. Our data suggest that HSV-dependent AAV replication is initiated by Rep contacting the AAV ITR and by cooperative binding of ICP8 on AAV ssDNA.
Stimulation of BK Virus DNA Replication by NFI Family Transcription Factors
Authors: Liang Bo , Tikhanovich Irina , Nasheuer Heinz Peter , Folk William R. 
Published: December 2011

ABSTRACTBK polyomavirus (BKV) establishes persistent, low-level, and asymptomatic infections in most humans and causes polyomavirus-associated nephropathy (PVAN) and other pathologies in some individuals. The activation of BKV replication following kidney transplantation, leading to viruria, viremia, and, ultimately, PVAN, is associated with immune suppression as well as inflammation and stress from ischemia-reperfusion injury of the allograft, but the stimuli and molecular mechanisms leading to these pathologies are not well defined. The replication of BKV DNA in cell cultures is regulated by the viral noncoding control region (NCCR) comprising the core origin and flanking sequences, to which BKV T antigen (Tag), cellular proteins, and small regulatory RNAs bind. Six nuclear factor I (NFI) binding sites occur in sequences flanking the late side of the core origin (the enhancer) of the archetype virus, and their mutation, either individually orin toto, reduces BKV DNA replication when placed in competition with templates containing intact BKV NCCRs. NFI family members interacted with the helicase domain of BKV Tag in pulldown assays, suggesting that NFI helps recruit Tag to the viral core origin and may modulate its function. However, Tag may not be the sole target of the replication-modulatory activities of NFI: the NFIC/CTF1 isotype stimulates BKV template replicationin vitroat low concentrations of DNA polymerase-α primase (Pol-primase), and the p58 subunit of Pol-primase associates with NFIC/CTF1, suggesting that NFI also recruits Pol-primase to the NCCR. These results suggest that NFI proteins (and the signaling pathways that target them) activate BKV replication and contribute to the consequent pathologies caused by acute infection.
Coordinate Deletion of N-Glycans from the Heptad Repeats of the Fusion F Protein of Newcastle Disease Virus Yields a Hyperfusogenic Virus with Increased Replication, Virulence, and Immunogenicity
Authors: Samal Sweety , Khattar Sunil K. , Kumar Sachin , Collins Peter L. , Samal Siba K. 
Published: December 2011

ABSTRACT

The role of N-linked glycosylation of the Newcastle disease virus (NDV) fusion (F) protein in viral replication and pathogenesis was examined by eliminating potential acceptor sites using a reverse genetics system for the moderately pathogenic strain Beaudette C (BC). The NDV-BC F protein contains six potential acceptor sites for N-linked glycosylation at residues 85, 191, 366, 447, 471, and 541 (sites Ng1 to Ng6, respectively). The sites at Ng2 and Ng5 are present in heptad repeat (HR) domains HR1 and HR2, respectively, and thus might affect fusion. Each N-glycosylation site was eliminated individually by replacing asparagine (N) with glutamine (Q), and a double mutant (Ng2 + 5) involving the two HR domains was also made. Each mutant was successfully recovered by reverse genetics except for the one involving Ng6, which is present in the cytoplasmic domain. All of the F proteins expressed by the recovered mutant viruses were efficiently cleaved and transported to the infected-cell surface. None of the individual mutations affected viral fusogenicity, but the double mutation at Ng2 and Ng5 in HR1 and HR2 increased fusogenicity >12-fold. The single mutations at sites Ng1, Ng2, and Ng5 resulted in modestly reduced multicycle growth
in vitro
. These three single mutations were also the most attenuating in eggs and 1-day-old chicks and were associated with decreased replication and spread in 2-week-old chickens. In contrast, the combination of the mutations at Ng2 and Ng5 yielded a virus that, compared to the BC parent, replicated >100-fold more efficiently
in vitro
, was more virulent in eggs and chicks, replicated more efficiently in chickens with enhanced tropism for the brain and gut, and elicited stronger humoral cell responses. These results illustrate the effects of N-glycosylation of the F protein on NDV pathobiology and suggest that the N-glycans in HR1 and HR2 coordinately downregulate viral fusion and virulence.
A Leucine Zipper Motif of a Tegument Protein Triggers Final Envelopment of Human Cytomegalovirus
Authors: Meissner Christina Sylvia , Suffner Sascha , Schauflinger Martin , von Einem Jens , Bogner Elke 
Published: December 2011

ABSTRACT

The product of the human cytomegalovirus (HCMV) UL71 gene is conserved throughout the herpesvirus family. During HCMV infection, protein pUL71 is required for efficient virion egress and is involved in the final steps of secondary envelopment leading to infectious viral particles. We found strong indications for oligomerization of pUL71 under native conditions when recombinant pUL71 was negatively stained and analyzed by electron microscopy. Oligomerization of pUL71 during infection was further verified by native and reducing polyacrylamide gel electrophoresis (PAGE). By
in silico
analyses of the pUL71 sequence, we noticed a basic leucine zipper (bZIP)-like domain, which might serve as an oligomerization domain. We demonstrated the requirement of the bZIP-like domain for pUL71 oligomerization by coimmunoprecipitation and bimolecular fluorescence complementation using a panel of pUL71 mutants. These studies revealed that the mutation of two leucine residues is sufficient to abrogate oligomerization but that intracellular localization of pUL71 was unaffected. To investigate the relevance of the bZIP domain in the viral context, recombinant viruses carrying mutations identical to those in the panel of pUL71 mutants were generated. bZIP-defective viral mutants showed impaired viral growth, a small-plaque phenotype, and an ultrastructural phenotype similar to that of the previously described UL71 stop mutant virus. The majority of virus particles within the viral assembly compartment exhibited various stages of incomplete envelopment, which is consistent with the growth defect for the bZIP mutants. From these data we conclude that the bZIP-like domain is required for oligomerization of pUL71, which seems to be essential for correct envelopment of HCMV.
Adenovirus E4orf3 Targets Transcriptional Intermediary Factor 1γ for Proteasome-Dependent Degradation during Infection
Authors: Forrester Natalie A. , Patel Rakesh N. , Speiseder Thomas , Groitl Peter , Sedgwick Garry G. , Shimwell Neil J. , Seed Robert I. , Catnaigh Pól Ó , McCabe Christopher J. , Stewart Grant S. , Dobner Thomas , Grand Roger J. A. , Martin Ashley , Turnell Andrew S. 
Published: December 2011

ABSTRACT
The ability of adenovirus early region proteins, E1B-55K and E4orf6, to usurp control of cellular ubiquitin ligases and target proteins for proteasome-dependent degradation during infection is well established. Here we show that the E4 gene product, E4orf3 can, independently of E1B-55K and E4orf6, target the transcriptional corepressor transcriptional intermediary factor 1γ (TIF1γ) for proteasome-mediated degradation during infection. Initial mass spectrometric studies identified TIF1 family members—TIF1α, TIF1β, and TIF1γ—as E1B-55K-binding proteins in both transformed and infected cells, but analyses revealed that, akin to TIF1α, TIF1γ is reorganized in an E4orf3-dependent manner to promyelocytic leukemia protein-containing nuclear tracks during infection. The use of a number of different adenovirus early region mutants identified the specific and sole requirement for E4orf3 in mediating TIF1γ degradation. Further analyses revealed that TIF1γ is targeted for degradation by a number of divergent human adenoviruses, suggesting that the ability of E4orf3 to regulate TIF1γ expression is evolutionarily conserved. We also determined that E4orf3 does not utilize the Cullin-based ubiquitin ligases, CRL2 and CRL5, or the TIF1α ubiquitin ligase in order to promote TIF1γ degradation. Further studies suggested that TIF1γ possesses antiviral activity and limits adenovirus early and late gene product expression during infection. Indeed, TIF1γ knockdown accelerates the adenovirus-mediated degradation of MRE11, while TIF1γ overexpression delays the adenovirus-mediated degradation of MRE11. Taken together, these studies have identified novel adenovirus targets and have established a new role for the E4orf3 protein during infection.
Characterization of the R263K Mutation in HIV-1 Integrase That Confers Low-Level Resistance to the Second-Generation Integrase Strand Transfer Inhibitor Dolutegravir
Authors: Quashie Peter K. , Mesplède Thibault , Han Ying-Shan , Oliveira Maureen , Singhroy Diane N. , Fujiwara Tamio , Underwood Mark R. , Wainberg Mark A. 
Published: December 2011

ABSTRACT

Integrase (IN) strand transfer inhibitors (INSTIs) have been developed to inhibit the ability of HIV-1 integrase to irreversibly link the reverse-transcribed viral DNA to the host genome. INSTIs have proven their high efficiency in inhibiting viral replication
in vitro
and in patients. However, first-generation INSTIs have only a modest genetic barrier to resistance, allowing the virus to escape these powerful drugs through several resistance pathways. Second-generation INSTIs, such as dolutegravir (DTG, S/GSK1349572), have been reported to have a higher resistance barrier, and no novel drug resistance mutation has yet been described for this drug. Therefore, we performed
in vitro
selection experiments with DTG using viruses of subtypes B, C, and A/G and showed that the most common mutation to emerge was R263K. Further analysis by site-directed mutagenesis showed that R263K does confer low-level resistance to DTG and decreased integration in cell culture without altering reverse transcription. Biochemical cell-free assays performed with purified IN enzyme containing R263K confirmed the absence of major resistance against DTG and showed a slight decrease in 3′ processing and strand transfer activities compared to the wild type. Structural modeling suggested and
in vitro
IN-DNA binding assays show that the R263K mutation affects IN-DNA interactions.
Contribution of Intra- and Interhost Dynamics to Norovirus Evolution
Authors: Bull Rowena A. , Eden John-Sebastian , Luciani Fabio , McElroy Kerensa , Rawlinson William D. , White Peter A. 
Published: December 2011

ABSTRACT
Norovirus (NoV) is an emerging RNA virus that has been associated with global epidemics of gastroenteritis. Each global epidemic arises with the emergence of novel antigenic variants. While the majority of NoV infections are mild and self-limiting, in the young, elderly, and immunocompromised, severe and prolonged illness can result. As yet, there is no vaccine or therapeutic treatment to prevent or control infection. In order to design effective control strategies, it is important to understand the mechanisms and source of the new antigenic variants. In this study, we used next-generation sequencing (NGS) technology to investigate genetic diversification in three contexts: the impact of a NoV transmission event on viral diversity and the contribution to diversity of intrahost evolution over both a short period of time (10 days), in accordance with a typical acute NoV infection, and a prolonged period of time (288 days), as observed for NoV chronic infections of immunocompromised individuals. Investigations of the transmission event revealed that minor variants at frequencies as low as 0.01% were successfully transmitted, indicating that transmission is an important source of diversity at the interhost level of NoV evolution. Our results also suggest that chronically infected immunocompromised subjects represent a potential reservoir for the emergence of new viral variants. In contrast, in a typical acute NoV infection, the viral population was highly homogenous and relatively stable. These results indicate that the evolution of NoV occurs through multiple mechanisms.
The Amino Acid Linker between the Endonuclease and Helicase Domains of Adeno-Associated Virus Type 5 Rep Plays a Critical Role in DNA-Dependent Oligomerization
Authors: Maggin Jenna E. , James Jeffrey A. , Chappie Joshua S. , Dyda Fred , Hickman Alison Burgess 
Published: December 2011

ABSTRACT
The adeno-associated virus (AAV) genome encodes four Rep proteins, all of which contain an SF3 helicase domain. The larger Rep proteins, Rep78 and Rep68, are required for viral replication, whereas Rep40 and Rep52 are needed to package AAV genomes into preformed capsids; these smaller proteins are missing the site-specific DNA-binding and endonuclease domain found in Rep68/78. Other viral SF3 helicases, such as the simian virus 40 large T antigen and the papillomavirus E1 protein, are active as hexameric assemblies. However, Rep40 and Rep52 have not been observed to form stable oligomers on their own or with DNA, suggesting that important determinants of helicase multimerization lie outside the helicase domain. Here, we report that when the 23-residue linker that connects the endonuclease and helicase domains is appended to the adeno-associated virus type 5 (AAV5) helicase domain, the resulting protein forms discrete complexes on DNA consistent with single or double hexamers. The formation of these complexes does not require the Rep binding site sequence, nor is it nucleotide dependent. These complexes have stimulated ATPase and helicase activities relative to the helicase domain alone, indicating that they are catalytically relevant, a result supported by negative-stain electron microscopy images of hexameric rings. Similarly, the addition of the linker region to the AAV5 Rep endonuclease domain also confers on it the ability to bind and multimerize on nonspecific double-stranded DNA. We conclude that the linker is likely a key contributor to Rep68/78 DNA-dependent oligomerization and may play an important role in mediating Rep68/78's conversion from site-specific DNA binding to nonspecific DNA unwinding.
Vif Proteins of Human and Simian Immunodeficiency Viruses Require Cellular CBFβ To Degrade APOBEC3 Restriction Factors
Authors: Hultquist Judd F. , Binka Mawuena , LaRue Rebecca S. , Simon Viviana , Harris Reuben S. 
Published: December 2011

ABSTRACT
HIV-1 requires the cellular transcription factor CBFβ to stabilize its accessory protein Vif and promote APOBEC3G degradation. Here, we demonstrate that both isoforms of CBFβ allow for increased steady-state levels of Vif, enhanced APOBEC3G degradation, and increased viral infectivity. This conserved functional interaction enhances the steady-state levels of Vif proteins from multiple HIV-1 subtypes and is required for the degradation of all human and rhesus Vif-sensitive APOBEC3 proteins by their respective lentiviral Vif proteins.
Does the death rate of Hong Kong Chinese change during the lunar ghost month?: Table 1
Authors: Panesar Nirmal S , Graham Colin A 
Published: December 2011

BackgroundMost cultures believe in ghosts and for the Chinese, the seventh lunar month—the ghost month—causes particular concern. ‘The gates of hell’ are open for the first 14 days of the month which allows the restless ghosts of people who were hungry when they died to haunt the living. In this study, it was hypothesised that if the notion that ghosts are out to harm the living could affect the Chinese, this may be reflected in death statistics.MethodsBecause the Chinese believe death is more likely during the ghost month, male and female deaths from all causes and from four common causes of death in the first and second fortnights of the seventh lunar months of 1995–2000 were compared in Hong Kong Chinese. Deaths in two consecutive fortnights 30 days before each year's seventh lunar month were used as controls. Death data were compared using the binomial test with a null hypothesis probability of 0.5 between the first and second fortnights.ResultsThere was no difference in male deaths between the first and second fortnights of the control and seventh lunar months. While there were no significant differences in female deaths during the control month periods, fewer women died overall in the first fortnight of the seventh lunar month (p=0.026).ConclusionTo protect their family, the Chinese women postpone death until after the hungry ghosts have been fed and hopefully banished forever.
Potentially avoidable emergency department attendance: interview study of patients' reasons for attendance
Authors: Agarwal S , Banerjee J , Baker R , Conroy S , Hsu R , Rashid A , Camosso-Stefinovic J , Sinfield P , Habiba M 
Published: December 2011

ObjectivesTo explore the reasons for attendance at the emergency department (ED) by patients who could have been managed in an alternative service and the rate of acute admissions to one acute hospital.DesignInterview study.SettingOne acute hospital (University Hospitals of Leicester) in the East Midlands.Participants23 patients and/or their carers.MethodsA purposive sample of patients attending the ED and the linked urgent care centre was identified and recruited. Patients in the sample were approached by a clinician and a researcher and invited to take part in an interview. Patients of different ethnicities and from different age groups, arriving at the ED via different referral routes (self-referral, emergency ambulance, GP referral, out-of-hours services) and attending at different times of the day and night were included. The interviews were recorded and transcribed with the individuals' permission and analysed using the framework analysis approach.ResultsPatients' anxiety or concern about the presenting problem, the range of services available to the ED and the perceived efficacy of these services, patients' perceptions of access to alternative services including general practice and lack of alternative pathways were factors that influenced the decision to use the ED.ConclusionsAccess to general practice, anxiety about the presenting problem, awareness and perceptions of the efficacy of the services available in the ED and lack of alternative pathways are important predictors of attendance rates.
CD8+ T Cells Specific for Tumor Antigens Can Be Rendered Dysfunctional by the Tumor Microenvironment through Upregulation of the Inhibitory Receptors BTLA and PD-1
Authors: Fourcade Julien , Sun Zhaojun , Pagliano Ornella , Guillaume Philippe , Luescher Immanuel F. , Sander Cindy , Kirkwood John M. , Olive Daniel , Kuchroo Vijay , Zarour Hassane M. 
Published: December 2011

Abstract
Cytotoxic T cells that are present in tumors and capable of recognizing tumor epitopes are nevertheless generally impotent in eliciting tumor rejection. Thus, identifying the immune escape mechanisms responsible for inducing tumor-specific CD8+ T-cell dysfunction may reveal effective strategies for immune therapy. The inhibitory receptors PD-1 and Tim-3 are known to negatively regulate CD8+ T-cell responses directed against the well-characterized tumor antigen NY-ESO-1. Here, we report that the upregulation of the inhibitory molecule BTLA also plays a critical role in restricting NY-ESO-1–specific CD8+ T-cell expansion and function in melanoma. BTLA-expressing PD-1+Tim-3− CD8+ T cells represented the largest subset of NY-ESO-1–specific CD8+ T cells in patients with melanoma. These cells were partially dysfunctional, producing less IFN-γ than BTLA− T cells but more IFN-γ, TNF, and interleukin-2 than the highly dysfunctional subset expressing all three receptors. Expression of BTLA did not increase with higher T-cell dysfunction or upon cognate antigen stimulation, as it does with PD-1, suggesting that BTLA upregulation occurs independently of functional exhaustion driven by high antigen load. Added with PD-1 and Tim-3 blockades, BTLA blockade enhanced the expansion, proliferation, and cytokine production of NY-ESO-1–specific CD8+ T cells. Collectively, our findings indicate that targeting BTLA along with the PD-1 and Tim-3 pathways is critical to reverse an important mechanism of immune escape in patients with advanced melanoma. Cancer Res; 72(4); 887–96. ©2011 AACR.