bluuebunny/crossref_metadata_2025_split huggingface dataset:
pH-Dependent Formation and Disintegration of the Influenza A Virus Protein Scaffold To Provide Tension for Membrane Fusion
Authors: Batishchev O. V. , Shilova L. A. , Kachala M. V. , Tashkin V. Y. , Sokolov V. S. , Fedorova N. V. , Baratova L. A. , Knyazev D. G. , Zimmerberg J. , Chizmadzhev Y. A. 
Published: October 2015

ABSTRACT
Influenza virus is taken up from a pH-neutral extracellular milieu into an endosome, whose contents then acidify, causing changes in the viral matrix protein (M1) that coats the inner monolayer of the viral lipid envelope. At a pH of ∼6, M1 interacts with the viral ribonucleoprotein (RNP) in a putative priming stage; at this stage, the interactions of the M1 scaffold coating the lipid envelope are intact. The M1 coat disintegrates as acidification continues to a pH of ∼5 to clear a physical path for the viral genome to transit from the viral interior to the cytoplasm. Here we investigated the physicochemical mechanism of M1's pH-dependent disintegration. In neutral media, the adsorption of M1 protein on the lipid bilayer was electrostatic in nature and reversible. The energy of the interaction of M1 molecules with each other in M1 dimers was about 10 times as weak as that of the interaction of M1 molecules with the lipid bilayer. Acidification drives conformational changes in M1 molecules due to changes in the M1 charge, leading to alterations in their electrostatic interactions. Dropping the pH from 7.1 to 6.0 did not disturb the M1 layer; dropping it lower partially desorbed M1 because of increased repulsion between M1 monomers still stuck to the membrane. Lipid vesicles coated with M1 demonstrated pH-dependent rupture of the vesicle membrane, presumably because of the tension generated by this repulsive force. Thus, the disruption of the vesicles coincident with M1 protein scaffold disintegration at pH 5 likely stretches the lipid membrane to the point of rupture, promoting fusion pore widening for RNP release.

IMPORTANCE
Influenza remains a top killer of human beings throughout the world, in part because of the influenza virus's rapid binding to cells and its uptake into compartments hidden from the immune system. To attack the influenza virus during this time of hiding, we need to understand the physical forces that allow the internalized virus to infect the cell. In particular, we need to know how the protective coat of protein inside the viral surface reacts to the changes in acid that come soon after internalization. We found that acid makes the molecules of the protein coat push each other while they are still stuck to the virus, so that they would like to rip the membrane apart. This ripping force is known to promote membrane fusion, the process by which infection actually occurs.
Crystal Structure of the Conserved Amino Terminus of the Extracellular Domain of Matrix Protein 2 of Influenza A Virus Gripped by an Antibody
Authors: Cho Ki Joon , Schepens Bert , Moonens Kristof , Deng Lei , Fiers Walter , Remaut Han , Saelens Xavier 
Published: October 2015

ABSTRACT
We report the crystal structure of the M2 ectodomain (M2e) in complex with a monoclonal antibody that binds the amino terminus of M2. M2e extends into the antibody binding site to form an N-terminal β-turn near the bottom of the paratope. This M2e folding differs significantly from that of M2e in complex with an antibody that binds another part of M2e. This suggests that M2e can adopt at least two conformations that can elicit protective antibodies.
Respiratory Syncytial Virus Attachment Glycoprotein Contribution to Infection Depends on the Specific Fusion Protein
Authors: Meng Jia , Hotard Anne L. , Currier Michael G. , Lee Sujin , Stobart Christopher C. , Moore Martin L. 
Published: October 2015

ABSTRACT

Human respiratory syncytial virus (RSV) is an important pathogen causing acute lower respiratory tract disease in children. The RSV attachment glycoprotein (G) is not required for infection, as G-null RSV replicates efficiently in several cell lines. Our laboratory previously reported that the viral fusion (F) protein is a determinant of strain-dependent pathogenesis. Here, we hypothesized that virus dependence on G is determined by the strain specificity of F. We generated recombinant viruses expressing G and F, or null for G, from the laboratory A2 strain (Katushka RSV-A2GA2F [kRSV-A2GA2F] and kRSV-GstopA2F) or the clinical isolate A2001/2-20 (kRSV-2-20G2-20F and kRSV-Gstop2-20F). We quantified the virus cell binding, entry kinetics, infectivity, and growth kinetics of these four recombinant viruses
in vitro
. RSV expressing the 2-20 G protein exhibited the greatest binding activity. Compared to the parental viruses expressing G and F, removal of 2-20 G had more deleterious effects on binding, entry, infectivity, and growth than removal of A2 G. Overall, RSV expressing 2-20 F had a high dependence on G for binding, entry, and infection.

IMPORTANCE
RSV is the leading cause of childhood acute respiratory disease requiring hospitalization. As with other paramyxoviruses, two major RSV surface viral glycoproteins, the G attachment protein and the F fusion protein, mediate virus binding and subsequent membrane fusion, respectively. Previous work on the RSV A2 prototypical strain demonstrated that the G protein is functionally dispensable for
in vitro
replication. This is in contrast to other paramyxoviruses that require attachment protein function as a prerequisite for fusion. We reevaluated this requirement for RSV using G and F proteins from clinical isolate 2-20. Compared to the laboratory A2 strain, the G protein from 2-20 had greater contributions to virus binding, entry, infectivity, and
in vitro
growth kinetics. Thus, the clinical isolate 2-20 F protein function depended more on its G protein, suggesting that RSV has a higher dependence on G than previously thought.
Single-Genome Sequencing of Hepatitis C Virus in Donor-Recipient Pairs Distinguishes Modes and Models of Virus Transmission and Early Diversification
Authors: Li Hui , Stoddard Mark B. , Wang Shuyi , Giorgi Elena E. , Blair Lily M. , Learn Gerald H. , Hahn Beatrice H. , Alter Harvey J. , Busch Michael P. , Fierer Daniel S. , Ribeiro Ruy M. , Perelson Alan S. , Bhattacharya Tanmoy , Shaw George M. 
Published: October 2015

ABSTRACT
Despite the recent development of highly effective anti-hepatitis C virus (HCV) drugs, the global burden of this pathogen remains immense. Control or eradication of HCV will likely require the broad application of antiviral drugs and development of an effective vaccine. A precise molecular identification of transmitted/founder (T/F) HCV genomes that lead to productive clinical infection could play a critical role in vaccine research, as it has for HIV-1. However, the replication schema of these two RNA viruses differ substantially, as do viral responses to innate and adaptive host defenses. These differences raise questions as to the certainty of T/F HCV genome inferences, particularly in cases where multiple closely related sequence lineages have been observed. To clarify these issues and distinguish between competing models of early HCV diversification, we examined seven cases of acute HCV infection in humans and chimpanzees, including three examples of virus transmission between linked donors and recipients. Using single-genome sequencing (SGS) of plasma vRNA, we found that inferred T/F sequences in recipients were identical to viral sequences in their respective donors. Early in infection, HCV genomes generally evolved according to a simple model of random evolution where the coalescent corresponded to the T/F sequence. Closely related sequence lineages could be explained by high multiplicity infection from a donor whose viral sequences had undergone a pretransmission bottleneck due to treatment, immune selection, or recent infection. These findings validate SGS, together with mathematical modeling and phylogenetic analysis, as a novel strategy to infer T/F HCV genome sequences.

IMPORTANCE
Despite the recent development of highly effective, interferon-sparing anti-hepatitis C virus (HCV) drugs, the global burden of this pathogen remains immense. Control or eradication of HCV will likely require the broad application of antiviral drugs and the development of an effective vaccine, which could be facilitated by a precise molecular identification of transmitted/founder (T/F) viral genomes and their progeny. We used single-genome sequencing to show that inferred HCV T/F sequences in recipients were identical to viral sequences in their respective donors and that viral genomes generally evolved early in infection according to a simple model of random sequence evolution. Altogether, the findings validate T/F genome inferences and illustrate how T/F sequence identification can illuminate studies of HCV transmission, immunopathogenesis, drug resistance development, and vaccine protection, including sieving effects on breakthrough virus strains.
Pan-ebolavirus and Pan-filovirus Mouse Monoclonal Antibodies: Protection against Ebola and Sudan Viruses
Authors: Holtsberg Frederick W. , Shulenin Sergey , Vu Hong , Howell Katie A. , Patel Sonal J. , Gunn Bronwyn , Karim Marcus , Lai Jonathan R. , Frei Julia C. , Nyakatura Elisabeth K. , Zeitlin Larry , Douglas Robin , Fusco Marnie L. , Froude Jeffrey W. , Saphire Erica Ollmann , Herbert Andrew S. , Wirchnianski Ariel S. , Lear-Rooney Calli M. , Alter Galit , Dye John M. , Glass Pamela J. , Warfield Kelly L. , Aman M. Javad 
Published: October 2015

ABSTRACT

The unprecedented 2014-2015 Ebola virus disease (EVD) outbreak in West Africa has highlighted the need for effective therapeutics against filoviruses. Monoclonal antibody (MAb) cocktails have shown great potential as EVD therapeutics; however, the existing protective MAbs are virus species specific. Here we report the development of pan-ebolavirus and pan-filovirus antibodies generated by repeated immunization of mice with filovirus glycoproteins engineered to drive the B cell responses toward conserved epitopes. Multiple pan-ebolavirus antibodies were identified that react to the Ebola, Sudan, Bundibugyo, and Reston viruses. A pan-filovirus antibody that was reactive to the receptor binding regions of all filovirus glycoproteins was also identified. Significant postexposure efficacy of several MAbs, including a novel antibody cocktail, was demonstrated. For the first time, we report cross-neutralization and
in vivo
protection against two highly divergent filovirus species, i.e., Ebola virus and Sudan virus, with a single antibody. Competition studies indicate that this antibody targets a previously unrecognized conserved neutralizing epitope that involves the glycan cap. Mechanistic studies indicated that, besides neutralization, innate immune cell effector functions may play a role in the antiviral activity of the antibodies. Our findings further suggest critical novel epitopes that can be utilized to design effective cocktails for broad protection against multiple filovirus species.

IMPORTANCE
Filoviruses represent a major public health threat in Africa and an emerging global concern. Largely driven by the U.S. biodefense funding programs and reinforced by the 2014 outbreaks, current immunotherapeutics are primarily focused on a single filovirus species called Ebola virus (EBOV) (formerly Zaire Ebola virus). However, other filoviruses including Sudan, Bundibugyo, and Marburg viruses have caused human outbreaks with mortality rates as high as 90%. Thus, cross-protective immunotherapeutics are urgently needed. Here, we describe monoclonal antibodies with cross-reactivity to several filoviruses, including the first report of a cross-neutralizing antibody that exhibits protection against Ebola virus and Sudan virus in mice. Our results further describe a novel combination of antibodies with enhanced protective efficacy. These results form a basis for further development of effective immunotherapeutics against filoviruses for human use. Understanding the cross-protective epitopes are also important for rational design of pan-ebolavirus and pan-filovirus vaccines.
TRIM5 Retroviral Restriction Activity Correlates with the Ability To Induce Innate Immune Signaling
Authors: Lascano Josefina , Uchil Pradeep D. , Mothes Walther , Luban Jeremy 
Published: October 2015

ABSTRACT
Host restriction factor TRIM5 inhibits retroviral transduction in a species-specific manner by binding to and destabilizing the retroviral capsid lattice before reverse transcription is completed. However, the restriction mechanism may not be that simple since TRIM5 E3 ubiquitin ligase activity, the proteasome, autophagy, and TAK1-dependent AP-1 signaling have been suggested to contribute to restriction. Here, we show that, among a panel of seven primate and Carnivora TRIM5 orthologues, each of which has potential for potent retroviral restriction activity, all activated AP-1 signaling. In contrast, TRIM family paralogues most closely related to TRIM5 did not. While each primate species has a single TRIM5 gene, mice have at least seven TRIM5 homologues that cluster into two groups, Trim12a, -b, and -c and Trim30a, -b, -c, and -d. The three Trim12 proteins activated innate immune signaling, while the Trim30 proteins did not, though none of the murine Trim5 homologues restricted any of a panel of cloned retroviruses. To determine if any mouse TRIM5 homologues had potential for restriction activity, each was fused to the human immunodeficiency virus type 1 (HIV-1) CA binding protein cyclophilin A (CypA). The three Trim12-CypA fusions all activated AP-1 and restricted HIV-1 transduction, whereas the Trim30-CypA fusions did neither. AP-1 activation and HIV-1 restriction by the Trim12-CypA fusions were inhibited by disruption of TAK1. Overall then, these experiments demonstrate that there is a strong correlation between TRIM5 retroviral restriction activity and the ability to activate TAK1-dependent innate immune signaling.

IMPORTANCE
The importance of retroviruses for the evolution of susceptible host organisms cannot be overestimated. Eight percent of the human genome is retrovirus sequence, fixed in the germ line during past infection. Understanding how metazoa protect their genomes from mutagenic retrovirus infection is therefore of fundamental importance to biology. TRIM5 is a cellular protein that protects host genome integrity by disrupting the retroviral capsid as it transports viral nucleic acid to the host cell nucleus. Previous data suggest that innate immune signaling contributes to TRIM5-mediated restriction. Here, we show that activation of innate immune signaling is conserved among primate and carnivore TRIM5 orthologues and among 3 of the 7 mouse Trim5 homologues and that such activity is required for TRIM5-mediated restriction activity.
Do patients with gastrointestinal cancer want to decide where they have tests and surgery? A questionnaire study of provider choice
Authors: Byrne Ben E , Faiz Omar D , Vincent Charles 
Published: October 2015

BackgroundsChoice of provider has been an important strategy among policy makers, intended, in part, to drive improvements in quality and efficiency of healthcare. This study examined the information requirements, and decision-making experiences and preferences of patients who have had surgery for gastrointestinal cancer, to assess the status of provider choice in current practice.MethodsThe single-item Control Preferences Scale was used to determine patients’ experiences and preferences when being referred for tests, and choosing where to have surgery. Participants used a Likert scale to rate the importance of 23 information items covering a variety of structures, processes and outcomes at the hospital level and the department level. Participants were recruited by post and online.Results463 responses were analysed. Patients reported very low levels of involvement in provider choice, with their doctor deciding where they underwent tests or surgery in 77.0% and 81.8% of cases, respectively. Over two-thirds of participants would have preferred greater involvement in provider choice than they experienced. Of note, patient age and education were not associated with reported preferences. Information on how long patients with cancer wait for treatment, annual operative volume and postoperative mortality rate, as well as retained foreign bodies and infection rates were considered very important.ConclusionsThere was a substantial unmet desire for greater involvement in provider choice among study participants. Respondents attached particular importance to surgery-specific information. Efforts should be made to increase involvement of patients with gastrointestinal cancer in provider decisions, across primary and secondary care, to deliver more patient-centred care. The reported lack of patient involvement in provider choice suggests it is unlikely to be working as an effective lever to drive quality improvement at present.
Underlying risk factors for prescribing errors in long-term aged care: a qualitative study
Authors: Tariq Amina , Georgiou Andrew , Raban Magdalena , Baysari Melissa Therese , Westbrook Johanna 
Published: October 2015

Objective(s)To identify system-related risk factors perceived to contribute to prescribing errors in Australian long-term care settings, that is, residential aged care facilities (RACFs).Design and settingThe study used qualitative methods to explore factors that contribute to unsafe prescribing in RACFs. Data were collected at three RACFs in metropolitan Sydney, Australia between May and November 2011. Participants included RACF managers, doctors, pharmacists and RACF staff actively involved in prescribing-related processes. Methods included non-participant observations (74 h), in-depth semistructured interviews (n=25) and artefact analysis. Detailed process activity models were developed for observed prescribing episodes supplemented by triangulated analysis using content analysis methods.ResultsSystem-related factors perceived to increase the risk of prescribing errors in RACFs were classified into three overarching themes: communication systems, team coordination and staff management. Factors associated with communication systems included limited point-of-care access to information, inadequate handovers, information storage across different media (paper, electronic and memory), poor legibility of charts, information double handling, multiple faxing of medication charts and reliance on manual chart reviews. Team factors included lack of established lines of responsibility, inadequate team communication and limited participation of doctors in multidisciplinary initiatives like medication advisory committee meetings. Factors related to staff management and workload included doctors’ time constraints and their accessibility, lack of trained RACF staff and high RACF staff turnover.ConclusionsThe study highlights several system-related factors including laborious methods for exchanging medication information, which often act together to contribute to prescribing errors. Multiple interventions (eg, technology systems, team communication protocols) are required to support the collaborative nature of RACF prescribing.
Expression of EphB6 in ovarian serous carcinoma is associated with grade, TNM stage and survival
Authors: Gu Yun , Li Fengshan , Qian Ning , Chen Xiao , Wang Hai , Wang Jiandong 
Published: October 2015

BackgroundOvarian serous carcinoma (OSC) is the most common ovarian epithelial malignancy. Despite new medical and surgical advances, the overall 5-year survival for OSC remains poor. There is an important need to determine diagnostic and prognostic markers for this disease. Eph receptors are the largest known family of receptor tyrosines characterised in humans. These receptors are involved in the development and progression of various diseases including cancer. EphB6 contains kinase domains that are altered in several conserved amino acids and is catalytically inactive. The aim of the present study was to correlate the immunohistological expression of EphB6 in a cohort of patients with epithelial ovarian tumours with clinicopathological parameters and survival.MethodsIn this study we examined the expression of EphB6 protein in 55 cases of OSC, 24 cases of benign ovarian serous tumours, 37 cases of serous borderline tumours and 20 cases with normal fallopian tubes by immunohistochemical staining with a polyclonal anti-EphB6 antibody. The relationship between EphB6 expression and pathological parameters was analysed. Kaplan–Meier survival function was used to analyse the prognosis.ResultsHigh expression of EphB6 was observed in 100% (20/20) of normal fallopian tube samples, 100% (24/24) of benign epithelial ovarian tumours, 78% (29/37) of ovarian serous borderline tumours and 18% (10/55) of OSCs (p<0.001). The expression of EphB6 was significantly associated with grade (p<0.001) and TNM stage (p=0.017). EphB6 expression was reversely related to Ki-67 (p=0.021). The survival analysis showed that patients with negative or weak expression of EphB6 protein had a poorer outcome than those with positive expression (p=0.046).ConclusionsOur results show that EphB6 is a new biomarker for distinguishing high- and low-grade OSC, and may be a potential prognostic marker in OSCs.
Sociodemographic risk factors of alcohol drinking in Hong Kong adolescents
Authors: Huang Rong , Ho Sai Yin , Wang Man Ping , Lo Wing Sze , Lam Tai Hing 
Published: October 2015

BackgroundAdolescent drinking in Western countries has been associated with older age and high socioeconomic status, but the association with family structure was inconsistent.MethodsIn a 2012–2013 school-based survey in Hong Kong, 23 096 students (mean age 14.7 years, SD 1.8 years) completed an anonymous questionnaire. Current drinking was defined as any drinking and binge drinking as consuming at least 5 drinks on one occasion, both in the past 30 days. Multilevel logistic regression was used to examine the association of sociodemographic characteristics with current drinking, binge drinking and type of alcohol consumed.ResultsCurrent drinking was associated with age (≥15 vs ≤14 years) (adjusted OR 1.87, 95% CI 1.74 to 2.02), higher perceived family affluence versus low affluence (1.11, 1.02 to 1.21 for medium affluence; 1.55, 1.38 to 1.75 for high affluence), private housing versus public housing (1.11, 1.01 to 1.21) and non-intact family versus intact family (1.31, 1.19 to 1.45 for separated/divorced parents; 1.40, 1.21 to 1.62 for one or both deceased parents). Similar risk factors were observed for binge drinking. Girls were more likely to drink fruit wine (1.48, 1.36 to 1.62), while they were less likely to drink beer (0.85, 0.79 to 0.92) and spirits (0.69, 0.54 to 0.87). Students who reported high family affluence were more likely to drink wine (1.91, 1.59 to 2.30) and spirits (2.23, 1.54 to 3.24).ConclusionsGenerally, adolescents who were older and had higher socioeconomic status were more likely to drink. High family affluence was associated with wine and spirits drinking. Beer and spirits were preferred more by boys, and fruit wine by girls. These results indicated high-risk groups for adolescent alcohol interventions.