bluuebunny/crossref_metadata_2025_split huggingface dataset:
Role of Cysteines in Stabilizing the Randomized Receptor Binding Domains within Feline Leukemia Virus Envelope Proteins
Authors: Valdivieso-Torres Leonardo , Sarangi Anindita , Whidby Jillian , Marcotrigiano Joseph , Roth Monica J. 
Published: December 2015

ABSTRACT
Retargeting of gammaretroviral envelope proteins has shown promising results in the isolation of novel isolates with therapeutic potential. However, the optimal conditions required to obtain high-affinity retargeted envelope proteins with narrow tropism are not understood. This study highlights the advantage of constrained peptides within receptor binding domains and validates the random library screening technique of obtaining novel retargeted Env proteins. Using a modified vector backbone to screen the envelope libraries on 143B osteosarcoma cells, three novel and unique retargeted envelopes were isolated. The use of complex disulfide bonds within variable regions required for receptor binding is found within natural gammaretroviral envelope isolates. Interestingly, two of the isolates, named AII and BV2, have a pair of cysteines located within the randomized region of 11 amino acids similar to that identified within the CP Env, an isolate identified in a previous Env library screen on the human renal carcinoma Caki-1 cell line. The amino acids within the randomized region of AII and BV2 envelopes that are essential for viral infection have been identified in this study and include these cysteine residues. Through mutagenesis studies, the putative disulfide bond pairs including and beyond the randomized region were examined. In parallel, the disulfide bonds of CP Env were identified using mass spectrometry. The results indicate that this pair of cysteines creates the structural context to position key hydrophobic (F and W) and basic (K and H) residues critical for viral titer and suggest that AII, BV2, and CP internal cysteines bond together in distinct ways.

IMPORTANCE
Retargeted gammaretroviral particles have broad applications for therapeutic use. Although great advances have been achieved in identifying new Env-host cell receptor pairs, the rules for designing optimal Env libraries are still unclear. We have found that isolates with an additional pair of cysteines within the randomized region have the highest transduction efficiencies. This emphasizes the importance of considering cysteine pairs in the design of new libraries. Furthermore, our data clearly indicate that these cysteines are essential for viral infectivity by presenting essential residues to the host cell receptor. These studies facilitate the screening of Env libraries for functional entry into target cells, allowing the identification of novel gammaretroviral Envs targeting alternative host cell receptors for gene and protein delivery.
Isolation and Characterization of a Novel Bat Coronavirus Closely Related to the Direct Progenitor of Severe Acute Respiratory Syndrome Coronavirus
Authors: Yang Xing-Lou , Hu Ben , Wang Bo , Wang Mei-Niang , Zhang Qian , Zhang Wei , Wu Li-Jun , Ge Xing-Yi , Zhang Yun-Zhi , Daszak Peter , Wang Lin-Fa , Shi Zheng-Li 
Published: December 2015

ABSTRACT
We report the isolation and characterization of a novel bat coronavirus which is much closer to the severe acute respiratory syndrome coronavirus (SARS-CoV) in genomic sequence than others previously reported, particularly in its S gene. Cell entry and susceptibility studies indicated that this virus can use ACE2 as a receptor and infect animal and human cell lines. Our results provide further evidence of the bat origin of the SARS-CoV and highlight the likelihood of future bat coronavirus emergence in humans.
Defining the Microglia Response during the Time Course of Chronic Neurodegeneration
Authors: Vincenti James E. , Murphy Lita , Grabert Kathleen , McColl Barry W. , Cancellotti Enrico , Freeman Tom C. , Manson Jean C. 
Published: December 2015

ABSTRACTInflammation has been proposed as a major component of neurodegenerative diseases, although the precise role it plays has yet to be defined. We examined the role of key contributors to this inflammatory process, microglia, the major resident immune cell population of the brain, in a prion disease model of chronic neurodegeneration. Initially, we performed an extensive reanalysis of a large study of prion disease, where the transcriptome of mouse brains had been monitored throughout the time course of disease. Our analysis has provided a detailed classification of the disease-associated genes based on cell type of origin and gene function. This revealed that the genes upregulated during disease, regardless of the strain of mouse or prion protein, are expressed predominantly by activated microglia. In order to study the microglia contribution more specifically, we established a mouse model of prion disease in which the 79A murine prion strain was introduced by an intraperitoneal route into BALB/cJFms-EGFP/−mice, which express enhanced green fluorescent protein under the control of thec-fmsoperon. Samples were taken at time points during disease progression, and histological analysis of the brain and transcriptional analysis of isolated microglia was carried out. The analysis of isolated microglia revealed a disease-specific, highly proinflammatory signature in addition to an upregulation of genes associated with metabolism and respiratory stress. This study strongly supports the growing recognition of the importance of microglia within the prion disease process and identifies the nature of the response through gene expression analysis of isolated microglia.IMPORTANCEInflammation has been proposed as a major component of neurodegenerative diseases. We have examined the role of key contributors to this inflammatory process, microglia, the major resident immune cell population of the brain, in a murine prion disease model of chronic neurodegeneration. Our study demonstrates that genes upregulated throughout the disease process are expressed predominantly by microglia. A disease-specific, highly proinflammatory signature was observed in addition to an upregulation of genes associated with metabolism and respiratory stress. This study strongly supports the growing recognition of the important contribution of microglia to a chronic neurodegenerative disease process.
Membrane Perturbation-Associated Ca
2+
Signaling and Incoming Genome Sensing Are Required for the Host Response to Low-Level Enveloped Virus Particle Entry
Authors: Hare David N. , Collins Susan E. , Mukherjee Subhendu , Loo Yueh-Ming , Gale Michael , Janssen Luke J. , Mossman Karen L. 
Published: December 2015

ABSTRACT

The type I interferon (IFN) response is an important aspect of innate antiviral defense, and the transcription factor IRF3 plays an important role in its induction. Membrane perturbation during fusion, a necessary step for enveloped virus particle entry, appears sufficient to induce transcription of a subset of IFN-stimulated genes (ISGs) in an IRF3-dependent, IFN-independent fashion. IRF3 is emerging as a central node in host cell stress responses, although it remains unclear how different forms of stress activate IRF3. Here, we investigated the minimum number of Sendai virus (SeV) and human cytomegalovirus (HCMV) particles required to activate IRF3 and trigger an antiviral response. We found that Ca
2+
signaling associated with membrane perturbation and recognition of incoming viral genomes by cytosolic nucleic acid receptors are required to activate IRF3 in response to fewer than 13 particles of SeV and 84 particles of HCMV per cell. Moreover, it appears that Ca
2+
signaling is important for activation of STING and IRF3 following HCMV particle entry, suggesting that Ca
2+
signaling sensitizes cells to recognize genomes within incoming virus particles. To our knowledge, this is the first evidence that cytosolic nucleic acid sensors recognize genomes within incoming virus particles prior to virus replication. These studies highlight the exquisite sensitivity of the cellular response to low-level stimuli and suggest that virus particle entry is sensed as a stress signal.

IMPORTANCE
The mechanism by which replicating viruses trigger IRF3 activation and type I IFN induction through the generation and accumulation of viral pathogen-associated molecular patterns has been well characterized. However, the mechanism by which enveloped virus particle entry mediates a stress response, leading to IRF3 activation and the IFN-independent response, remained elusive. Here, we find that Ca
2+
signaling associated with membrane perturbation appears to sensitize cells to recognize genomes within incoming virus particles. To our knowledge, this is the first study to show that cytosolic receptors recognize genomes within incoming virus particles prior to virus replication. These findings not only highlight the sensitivity of cellular responses to low-level virus particle stimulation, but provide important insights into how nonreplicating virus vectors or synthetic lipid-based carriers used as clinical delivery vehicles activate innate immune responses.
Structure-Guided Redesign Increases the Propensity of HIV Env To Generate Highly Stable Soluble Trimers
Authors: Guenaga Javier , Dubrovskaya Viktoriya , de Val Natalia , Sharma Shailendra K. , Carrette Barbara , Ward Andrew B. , Wyatt Richard T. 
Published: December 2015

ABSTRACT
Due to high viral diversity, an effective HIV-1 vaccine will likely require Envs derived from multiple subtypes to generate broadly neutralizing antibodies (bNAbs). Soluble Env mimics, like the native flexibly linked (NFL) and SOSIP trimers, derived from the subtype A BG505 Env, form homogeneous, stable native-like trimers. However, other Env sequences, such as JRFL and 16055 from subtypes B and C, do so to a lesser degree. The high-resolution BG505 SOSIP crystal structures permit the identification and redesign of Env elements involved in trimer stability. Here, we identified structure trimer-derived (TD) residues that increased the propensity of the subtype B JRFL and subtype C 16055 Env sequences to form well-ordered, homogenous, and highly stable soluble trimers. The generation of these spike mimics no longer required antibody-based selection, positive or negative. Using the redesigned subtype B and C trimer representatives as respective foundations, we further stabilized the NFL TD trimers by engineering an intraprotomer disulfide linkage in the prebridging sheet, I201C-A433C (CC), that locks the gp120 in the receptor nontriggered state. We demonstrated that this disulfide pair prevented CD4 induced-conformational rearrangements in NFL trimers derived from the prototypic subtype A, B, and C representatives. Coupling the TD-based design with the engineered disulfide linkage, CC, increased the propensity of Env to form soluble highly stable spike mimics that are resistant to CD4-induced changes. These advances will allow testing of the hypothesis that such stabilized immunogens will more efficiently elicit neutralizing antibodies in small-animal models and primates.

IMPORTANCE
HIV-1 displays unprecedented global diversity circulating in the human population. Since the envelope glycoprotein (Env) is the target of neutralizing antibodies, Env-based vaccine candidates that address such diversity are needed. Soluble well-ordered Env mimics, typified by NFL and SOSIP trimers, are attractive vaccine candidates. However, the current designs do not allow most Envs to form well-ordered trimers. Here, we made design modifications to increase the propensity of representatives from two of the major HIV subtypes to form highly stable trimers. This approach should be applicable to other viral Envs, permitting the generation of a repertoire of homogeneous, highly stable trimers. The availability of such an array will allow us to assess if sequential or cocktail immune strategies can overcome some of the vaccine challenges presented by HIV diversity.
Tumor Suppressor Interferon-Regulatory Factor 1 Counteracts the Germinal Center Reaction Driven by a Cancer-Associated Gammaherpesvirus
Authors: Mboko Wadzanai P. , Olteanu Horatiu , Ray Avijit , Xin Gang , Darrah Eric J. , Kumar Suresh N. , Kulinski Joseph M. , Cui Weiguo , Dittel Bonnie N. , Gauld Stephen B. , Tarakanova Vera L. 
Published: December 2015

ABSTRACT
Gammaherpesviruses are ubiquitous pathogens that are associated with the development of B cell lymphomas. Gammaherpesviruses employ multiple mechanisms to transiently stimulate a broad, polyclonal germinal center reaction, an inherently mutagenic stage of B cell differentiation that is thought to be the primary target of malignant transformation in virus-driven lymphomagenesis. We found that this gammaherpesvirus-driven germinal center expansion was exaggerated and lost its transient nature in the absence of interferon-regulatory factor 1 (IRF-1), a transcription factor with antiviral and tumor suppressor functions. Uncontrolled and persistent expansion of germinal center B cells led to pathological changes in the spleens of chronically infected IRF-1-deficient animals. Additionally, we found decreased IRF-1 expression in cases of human posttransplant lymphoproliferative disorder, a malignant condition associated with gammaherpesvirus infection. The results of our study define an unappreciated role for IRF-1 in B cell biology and provide insight into the potential mechanism of gammaherpesvirus-driven lymphomagenesis.

IMPORTANCE
Gammaherpesviruses establish lifelong infection in most adults and are associated with B cell lymphomas. While the infection is asymptomatic in many hosts, it is critical to identify individuals who may be at an increased risk of virus-induced cancer. Such identification is currently impossible, as the host risk factors that predispose individuals toward viral lymphomagenesis are poorly understood. The current study identifies interferon-regulatory factor 1 (IRF-1) to be one of such candidate host factors. Specifically, we found that IRF-1 enforces long-term suppression of an inherently mutagenic stage of B cell differentiation that gammaherpesviruses are thought to target for transformation. Correspondingly, in the absence of IRF-1, chronic gammaherpesvirus infection induced pathological changes in the spleens of infected animals. Further, we found decreased IRF-1 expression in human gammaherpesvirus-induced B cell malignancies.
Cucumber Necrosis Virus Recruits Cellular Heat Shock Protein 70 Homologs at Several Stages of Infection
Authors: Alam Syed Benazir , Rochon D'Ann 
Published: December 2015

ABSTRACT

RNA viruses often depend on host factors for multiplication inside cells due to the constraints of their small genome size and limited coding capacity. One such factor that has been exploited by several plant and animal viruses is heat shock protein 70 (HSP70) family homologs which have been shown to play roles for different viruses in viral RNA replication, viral assembly, disassembly, and cell-to-cell movement. Using next generation sequence analysis, we reveal that several isoforms of Hsp70 and Hsc70 transcripts are induced to very high levels during cucumber necrosis virus (CNV) infection of
Nicotiana benthamiana
and that HSP70 proteins are also induced by at least 10-fold. We show that HSP70 family protein homologs are co-opted by CNV at several stages of infection. We have found that overexpression of Hsp70 or Hsc70 leads to enhanced CNV genomic RNA, coat protein (CP), and virion accumulation, whereas downregulation leads to a corresponding decrease. Hsc70-2 was found to increase solubility of CNV CP
in vitro
and to increase accumulation of CNV CP independently of viral RNA replication during coagroinfiltration in
N. benthamiana
. In addition, virus particle assembly into virus-like particles in CP agroinfiltrated plants was increased in the presence of Hsc70-2. HSP70 was found to increase the targeting of CNV CP to chloroplasts during infection, reinforcing the role of HSP70 in chloroplast targeting of host proteins. Hence, our findings have led to the discovery of a highly induced host factor that has been co-opted to play multiple roles during several stages of the CNV infection cycle.

IMPORTANCE
Because of the small size of its RNA genome, CNV is dependent on interaction with host cellular components to successfully complete its multiplication cycle. We have found that CNV induces HSP70 family homologs to a high level during infection, possibly as a result of the host response to the high levels of CNV proteins that accumulate during infection. Moreover, we have found that CNV co-opts HSP70 family homologs to facilitate several aspects of the infection process such as viral RNA, coat protein and virus accumulation. Chloroplast targeting of the CNV CP is also facilitated, which may aid in CNV suppression of host defense responses. Several viruses have been shown to induce HSP70 during infection and others to utilize HSP70 for specific aspects of infection such as replication, assembly, and disassembly. We speculate that HSP70 may play multiple roles in the infection processes of many viruses.
Naturally Occurring Major and Minor Capsid Protein Variants of Human Papillomavirus 45 (HPV45): Differential Recognition by Cross-Neutralizing Antibodies Generated by HPV Vaccines
Authors: Godi Anna , Facchetti Alessandra , Bissett Sara L. , Cocuzza Clementina , Miller Elizabeth , Beddows Simon 
Published: December 2015

ABSTRACT
We investigated naturally occurring variation within the major (L1) and minor (L2) capsid proteins of human papillomavirus genotype 45 (HPV45). Pseudoviruses (PsVs) representing HPV45 sublineages A1, A2, A3, B1, and B2 exhibited comparable particle-to-infectivity ratios and morphologies but demonstrated both increased (A2, A3, and B1) and decreased (B2) sensitivities to cross-neutralization by HPV vaccine antibodies compared to that of the A1 sublineage. Mutant PsVs identified HI loop residue 357 as being critical for conferring this differential sensitivity.
Nef Proteins from HIV-1 Elite Controllers Are Inefficient at Preventing Antibody-Dependent Cellular Cytotoxicity
Authors: Alsahafi Nirmin , Ding Shilei , Richard Jonathan , Markle Tristan , Brassard Nathalie , Walker Bruce , Lewis George K. , Kaufmann Daniel E. , Brockman Mark A. , Finzi Andrés 
Published: December 2015

ABSTRACTImpairment of Nef function, including reduced CD4 downregulation, was described in a subset of HIV-1-infected individuals that control viral replication without antiretroviral treatment (elite controllers [EC]). Elimination of HIV-1-infected cells by antibody-dependent cellular cytotoxicity (ADCC) requires the presence of envelope glycoproteins (Env) in the CD4-bound conformation, raising the possibility that accumulating CD4 at the surface of virus-infected cells in EC could interact with Env and thereby sensitize these cells to ADCC. We observed a significant increase in the exposure of Env epitopes targeted by ADCC-mediating antibodies at the surface of cells expressing Nef isolates from EC; this correlated with enhanced susceptibility to ADCC. Altogether, our results suggest that enhanced susceptibility of HIV-1-infected cells to ADCC may contribute to the EC phenotype.IMPORTANCENef clones derived from elite controllers (EC) have been shown to be attenuated for CD4 downregulation; how this contributes to the nonprogressor phenotype of these infected individuals remains uncertain. Increasing evidence supports a role for HIV-specific antibody-dependent cellular cytotoxicity (ADCC) in controlling viral infection and replication. Here, we show that residual CD4 left at the surface of cells expressing Nef proteins isolated from ECs are sufficient to allow Env-CD4 interaction, leading to increased exposure of Env CD4-induced epitopes and increased susceptibility of infected cells to ADCC. Our results suggest that ADCC might be an active immune mechanism in EC that helps to maintain durable suppression of viral replication and low plasma viremia level in this rare subset of infected individuals. Therefore, targeting Nef's ability to downregulate CD4 could render HIV-1-infected cells susceptible to ADCC and thus have therapeutic utility.
Network Analysis of the Chronic Hepatitis C Virome Defines Hypervariable Region 1 Evolutionary Phenotypes in the Context of Humoral Immune Responses
Authors: Palmer Brendan A. , Schmidt-Martin Daniel , Dimitrova Zoya , Skums Pavel , Crosbie Orla , Kenny-Walsh Elizabeth , Fanning Liam J. 
Published: December 2015

ABSTRACT
Hypervariable region 1 (HVR1) of hepatitis C virus (HCV) comprises the first 27 N-terminal amino acid residues of E2. It is classically seen as the most heterogeneous region of the HCV genome. In this study, we assessed HVR1 evolution by using ultradeep pyrosequencing for a cohort of treatment-naive, chronically infected patients over a short, 16-week period. Organization of the sequence set into connected components that represented single nucleotide substitution events revealed a network dominated by highly connected, centrally positioned master sequences. HVR1 phenotypes were observed to be under strong purifying (stationary) and strong positive (antigenic drift) selection pressures, which were coincident with advancing patient age and cirrhosis of the liver. It followed that stationary viromes were dominated by a single HVR1 variant surrounded by minor variants comprised from conservative single amino acid substitution events. We present evidence to suggest that neutralization antibody efficacy was diminished for stationary-virome HVR1 variants. Our results identify the HVR1 network structure during chronic infection as the preferential dominance of a single variant within a narrow sequence space.

IMPORTANCE
HCV infection is often asymptomatic, and chronic infection is generally well established in advance of initial diagnosis and subsequent treatment. HVR1 can undergo rapid sequence evolution during acute infection, and the variant pool is typically seen to diverge away from ancestral sequences as infection progresses from the acute to the chronic phase. In this report, we describe HVR1 viromes in chronically infected patients that are defined by a dominant epitope located centrally within a narrow variant pool. Our findings suggest that weakened humoral immune activity, as a consequence of persistent chronic infection, allows for the acquisition and maintenance of host-specific adaptive mutations at HVR1 that reflect virus fitness.