bluuebunny/crossref_metadata_2025_split huggingface dataset:
Targeted DNA Mutagenesis for the Cure of Chronic Viral Infections
Authors: Schiffer Joshua T. , Aubert Martine , Weber Nicholas D. , Mintzer Esther , Stone Daniel , Jerome Keith R. 
Published: June 2012

ABSTRACTHuman immunodeficiency virus type 1 (HIV-1), hepatitis B virus (HBV), and herpes simplex virus (HSV) have been incurable to date because effective antiviral therapies target only replicating viruses and do not eradicate latently integrated or nonreplicating episomal viral genomes. Endonucleases that can target and cleave critical regions within latent viral genomes are currently in development. These enzymes are being engineered with high specificity such that off-target binding of cellular DNA will be absent or minimal. Imprecise nonhomologous-end-joining (NHEJ) DNA repair following repeated cleavage at the same critical site may permanently disrupt translation of essential viral proteins. We discuss the benefits and drawbacks of three types of DNA cleavage enzymes (zinc finger endonucleases, transcription activator-like [TAL] effector nucleases [TALENs], and homing endonucleases [also called meganucleases]), the development of delivery vectors for these enzymes, and potential obstacles for successful treatment of chronic viral infections. We then review issues regarding persistence of HIV-1, HBV, and HSV that are relevant to eradication with genome-altering approaches.
The Threefold Protrusions of Adeno-Associated Virus Type 8 Are Involved in Cell Surface Targeting as Well as Postattachment Processing
Authors: Raupp Christina , Naumer Matthias , Müller Oliver J. , Gurda Brittney L. , Agbandje-McKenna Mavis , Kleinschmidt Jürgen A. 
Published: June 2012

ABSTRACT

Adeno-associated virus (AAV) has attracted considerable interest as a vector for gene therapy owing its lack of pathogenicity and the wealth of available serotypes with distinct tissue tropisms. One of the most promising isolates for vector development, based on its superior gene transfer efficiency to the liver in small animals compared to AAV type 2 (AAV2), is AAV8. Comparison of the
in vivo
gene transduction of rAAV2 and rAAV8 in mice showed that single amino acid exchanges in the 3-fold protrusions of AAV8 in the surface loops comprised of residues 581 to 584 and 589 to 592 to the corresponding amino acids of AAV2 and vice versa had a strong influence on transduction efficiency and tissue tropism. Surprisingly, not only did conversion of AAV8 to AAV2
cap
sequences increase the transduction efficiency and change tissue tropism but so did the reciprocal conversion of AAV2 to AAV8. Insertion of new peptide motifs at position 590 in AAV8 also enabled retargeting of AAV8 capsids to specific tissues, suggesting that these sequences can interact with receptors on the cell surface. However, a neutralizing monoclonal antibody that binds to amino acids
588
QQNTA
592
of AAV8 does not prevent cell binding and virus uptake, indicating that this region is not necessary for receptor binding but rather that the antibody interferes with an essential step of postattachment processing in which the 3-fold protrusion is also involved. This study supports a multifunctional role of the 3-fold region of AAV capsids in the infection process.
Expansion of Polyfunctional HIV-Specific T Cells upon Stimulation with mRNA Electroporated Dendritic Cells in the Presence of Immunomodulatory Drugs
Authors: De Keersmaecker Brenda , Allard Sabine D. , Lacor Patrick , Schots Rik , Thielemans Kris , Aerts Joeri L. 
Published: June 2012

ABSTRACTRecently, it has been demonstrated that disease progression during HIV infection is not determined merely by the number of HIV-specific T cells but also by their quality (J. R. Almeida, et al., J. Exp. Med. 204:2473–2485, 2007; C. T. Berger, et al., J. Virol. 85:9334–9345, 2011; M. R. Betts, et al., Blood 107:4781–4789, 2006; V. V. Ganusov, et al., J. Virol. 85:10518–10528, 2011; P. Kiepiela, et al., Nat. Med. 13:46–53, 2007; and F. Pereyra, et al., J. Infect. Dis. 197:563–571, 2008). Therefore, strategies to specifically enhance or induce high-quality, HIV-specific T-cell responses are necessary to develop effective immune therapies. Thalidomide, lenalidomide, and pomalidomide have a strong capacity to boost immune responses and are therefore referred to as immunomodulatory drugs (IMiDs). We evaluated the effects of lenalidomide and pomalidomide on HIV-specific T cells. We found that the presence of IMiDs duringin vitroT-cell stimulation with dendritic cells electroporated with Gag- or Nef-encoding mRNA resulted in higher numbers of cytokine-secreting HIV-specific CD8+T cells, particularly inducing polyfunctional HIV-specific CD8+T cells with an enhanced lytic capacity. Furthermore, CD8+T-cell responses were detected upon stimulation with lower antigenic peptide concentrations, and a higher number of Gag epitopes was recognized upon addition of IMiDs. Finally, IMiDs reduced the proliferation of the HIV-specific CD4+T cells while increasing the number of polyfunctional CD4+T cells. These results provide new information about the effects of IMiDs on antigen-specific T cells and suggest that these drugs increase the efficacy of immune therapies for infectious diseases and cancer.
DNA Prime-Adenovirus Boost Immunization Induces a Vigorous and Multifunctional T-Cell Response against Hepadnaviral Proteins in the Mouse and Woodchuck Model
Authors: Kosinska Anna D. , Johrden Lena , Zhang Ejuan , Fiedler Melanie , Mayer Anja , Wildner Oliver , Lu Mengji , Roggendorf Michael 
Published: June 2012

ABSTRACTInduction of hepatitis B virus (HBV)-specific cytotoxic T cells by therapeutic immunization may be a strategy to treat chronic hepatitis B. In the HBV animal model, woodchucks, the application of DNA vaccine expressing woodchuck hepatitis virus (WHV) core antigen (WHcAg) in combination with antivirals led to the prolonged control of viral replication. However, it became clear that the use of more potent vaccines is required to overcome WHV persistence. Therefore, we asked whether stronger and more functional T-cell responses could be achieved using the modified vaccines and an optimized prime-boost vaccination regimen. We developed a new DNA plasmid (pCGWHc) and recombinant adenoviruses (AdVs) showing high expression levels of WHcAg. Mice vaccinated with the improved plasmid pCGWHc elicited a stronger WHcAg-specific CD8+T-cell response than with the previously used vaccines. Using multicolor flow cytometry and anin vivocytotoxicity assay, we showed that immunization in a DNA prime-AdV boost regimen resulted in an even more vigorous and functional T-cell response than immunization with the new plasmid alone. Immunization of naïve woodchucks with pCGWHc plasmid or AdVs induced a significant WHcAg-specific degranulation response prior to the challenge, this response had not been previously detected. Consistently, this response led to a rapid control of infection after the challenge. Our results demonstrate that high antigen expression levels and the DNA prime-AdV boost immunization improved the T-cell response in mice and induced significant T-cell responses in woodchucks. Therefore, this new vaccination strategy may be a candidate for a therapeutic vaccine against chronic HBV infection.
Foot-and-Mouth Disease Virus 3C Protease Cleaves NEMO To Impair Innate Immune Signaling
Authors: Wang Dang , Fang Liurong , Li Kui , Zhong Huijuan , Fan Jinxiu , Ouyang Chao , Zhang Huan , Duan Erzhen , Luo Rui , Zhang Zhongming , Liu Xiangtao , Chen Huanchun , Xiao Shaobo 
Published: June 2012

ABSTRACT

Foot-and-mouth disease is a highly contagious viral illness of wild and domestic cloven-hoofed animals. The causative agent, foot-and-mouth disease virus (FMDV), replicates rapidly, efficiently disseminating within the infected host and being passed on to susceptible animals via direct contact or the aerosol route. To survive in the host, FMDV has evolved to block the host interferon (IFN) response. Previously, we and others demonstrated that the leader proteinase (L
pro
) of FMDV is an IFN antagonist. Here, we report that another FMDV-encoded proteinase, 3C
pro
, also inhibits IFN-α/β response and the expression of IFN-stimulated genes. Acting in a proteasome- and caspase-independent manner, the 3C
pro
of FMDV proteolytically cleaved nuclear transcription factor kappa B (NF-κB) essential modulator (NEMO), a bridging adaptor protein essential for activating both NF-κB and interferon-regulatory factor signaling pathways. 3C
pro
specifically targeted NEMO at the Gln 383 residue, cleaving off the C-terminal zinc finger domain from the protein. This cleavage impaired the ability of NEMO to activate downstream IFN production and to act as a signaling adaptor of the RIG-I/MDA5 pathway. Mutations specifically disrupting the cysteine protease activity of 3C
pro
abrogated NEMO cleavage and the inhibition of IFN induction. Collectively, our data identify NEMO as a substrate for FMDV 3C
pro
and reveal a novel mechanism evolved by a picornavirus to counteract innate immune signaling.
Latent HIV-1 Infection Occurs in Multiple Subsets of Hematopoietic Progenitor Cells and Is Reversed by NF-κB Activation
Authors: McNamara Lucy A. , Ganesh Janani A. , Collins Kathleen L. 
Published: June 2012

ABSTRACTThe ability of HIV-1 to establish a latent infection presents a barrier to curing HIV. The best-studied reservoir of latent virusin vivois resting memory CD4+T cells, but it has recently been shown that CD34+hematopoietic progenitor cells (HPCs) can also become latently infected by HIV-1in vitroandin vivo. CD34+cells are not homogenous, however, and it is not yet known which types of CD34+cells support a latent infection. Furthermore, the mechanisms through which latency is established in this cell type are not yet known. Here we report the development of a primary cell model for latent HIV-1 infection in HPCs. We demonstrate that in this model, latent infection can be established in all subsets of HPCs examined, including HPCs with cell surface markers consistent with immature hematopoietic stem and progenitor cells. We further show that the establishment of latent infection in these cells can be reversed by tumor necrosis factor alpha (TNF-α) through an NF-κB-dependent mechanism. In contrast, we do not find evidence for a role of positive transcription elongation factor b (P-TEFb) in the establishment of latent infection in HPCs. Finally, we demonstrate that prostratin and suberoylanilide hydroxamic acid (SAHA), but not hexamethylene bisacetamide (HMBA) or 5-aza-2′-deoxycytidine (Aza-CdR), reactivate latent HIV-1 in HPCs. These findings illuminate the mechanisms through which latent infection can be established in HPCs and suggest common pathways through which latent virus could be reactivated in both HPCs and resting memory T cells to eliminate latent reservoirs of HIV-1.
STAT4 Regulates Antiviral Gamma Interferon Responses and Recurrent Disease during Herpes Simplex Virus 2 Infection
Authors: Svensson Alexandra , Tunbäck Petra , Nordström Inger , Shestakov Andrey , Padyukov Leonid , Eriksson Kristina 
Published: June 2012

ABSTRACTSTAT4 is an important transcription factor that contributes to the incidence and severity of different autoimmune diseases and is implicated in the antiviral immune responses in mice. In this study, we evaluated the role of STAT4 in human and murine herpes simplex virus 2 (HSV-2) infections. We show that STAT4 regulates antiviral gamma interferon (IFN-γ) responses and disease severity during chronic HSV-2 infections in humans and vaccine-induced IFN-γ-mediated protection against HSV-2 infection in mice. In a cohort of 228 HSV-2-infected individuals, representing both patients with recurrent disease and asymptomatic HSV-2 carriers, we found that genetic variations in theSTAT4gene were associated with asymptomatic HSV-2 infection, as well as with increasedin vitrosecretion of IFN-γ in response to the virus. Mice that lacked STAT4 had impaired HSV-2-specific IFN-γ production and delayed-type hypersensitivity responses following vaccination, which led to impaired viral clearance in the genital tract of vaccinated animals after a genital HSV-2 challenge. We conclude that STAT4 plays an important role in IFN-γ-mediated HSV-2-specific immunity, affecting the severity of genital HSV-2 infection.
Large Ribosomal Protein 4 Increases Efficiency of Viral Recoding Sequences
Authors: Green Lisa , Houck-Loomis Brian , Yueh Andrew , Goff Stephen P. 
Published: June 2012

ABSTRACTExpression of retroviral replication enzymes (Pol) requires a controlled translational recoding event to bypass the stop codon at the end ofgag. This recoding event occurs either by direct suppression of termination via the insertion of an amino acid at the stop codon (readthrough) or by alteration of the mRNA reading frame (frameshift). Here we report the effects of a host protein, large ribosomal protein 4 (RPL4), on the efficiency of recoding. Using a dual luciferase reporter assay, we found that transfection of cells with a plasmid encoding RPL4 cDNA increases recoding efficiency in a dose-dependent manner, with a maximal enhancement of nearly twofold. Expression of RPL4 increases recoding of reporters containing retroviral readthrough and frameshift sequences, as well as the Sindbis virus leaky termination signal. RPL4-induced enhancement of recoding is cell line specific and appears to be specific to RPL4 among ribosomal proteins. Cotransfection of RPL4 cDNA with Moloney murine leukemia proviral DNA results in Gag processing defects and a reduction of viral particle formation, presumably caused by the RPL4-dependent alteration of the Gag-to-Gag-Pol ratio required for virion assembly and release.
Reevaluation of the Coding Potential and Proteomic Analysis of the BAC-Derived Rhesus Cytomegalovirus Strain 68-1
Authors: Malouli Daniel , Nakayasu Ernesto S. , Viswanathan Kasinath , Camp David G. , Chang W. L. William , Barry Peter A. , Smith Richard D. , Früh Klaus 
Published: June 2012

ABSTRACTCytomegaloviruses are highly host restricted, resulting in cospeciation with their hosts. As a natural pathogen of rhesus macaques (RM), rhesus cytomegalovirus (RhCMV) has therefore emerged as a highly relevant experimental model for pathogenesis and vaccine development due to its close evolutionary relationship to human CMV (HCMV). Mostin vivoexperiments performed with RhCMV employed strain 68-1 cloned as a bacterial artificial chromosome (BAC). However, the complete genome sequence of the 68-1 BAC has not been determined. Furthermore, the gene content of the RhCMV genome is unknown, and previous open reading frame (ORF) predictions relied solely on uninterrupted ORFs with an arbitrary cutoff of 300 bp. To obtain a more precise picture of the actual proteins encoded by the most commonly used molecular clone of RhCMV, we reevaluated the RhCMV 68-1 BAC genome by whole-genome shotgun sequencing and determined the protein content of the resulting RhCMV virions by proteomics. By comparing the RhCMV genome to those of several related Old World monkey (OWM) CMVs, we were able to filter out many unlikely ORFs and obtain a simplified map of the RhCMV genome. This comparative genomics analysis suggests a high degree of ORF conservation among OWM CMVs, thus decreasing the likelihood that ORFs found only in RhCMV comprise true genes. Moreover, virion proteomics independently validated the revised ORF predictions, since only proteins that were conserved across OWM CMVs could be detected. Taken together, these data suggest a much higher conservation of genome and virion structure between CMVs of humans, apes, and OWMs than previously assumed.
The First Full-Length Endogenous Hepadnaviruses: Identification and Analysis
Authors: Liu Wei , Pan Shaokun , Yang Huijuan , Bai Weiya , Shen Zhongliang , Liu Jing , Xie Youhua 
Published: June 2012

ABSTRACT

In silico
screening of metazoan genome data identified multiple endogenous hepadnaviral elements in the budgerigar (
Melopsittacus undulatus
) genome, most notably two elements comprising about 1.3× and 1.0× the full-length genome. Phylogenetic and molecular dating analyses show that endogenous budgerigar hepatitis B viruses (eBHBV) share an ancestor with extant avihepadnaviruses and infiltrated the budgerigar genome millions of years ago. Identification of full-length genomes with preserved key features like ε signals could enable resurrection of ancient BHBV.
A Nuclear Localization Signal in Herpesvirus Protein VP1-2 Is Essential for Infection via Capsid Routing to the Nuclear Pore
Authors: Abaitua F. , Hollinshead M. , Bolstad M. , Crump C. M. , O'Hare P. 
Published: June 2012

ABSTRACT
To initiate infection, herpesviruses must navigate to and transport their genomes across the nuclear pore. VP1-2 is a large structural protein of the virion that is conserved in all herpesviruses and plays multiple essential roles in virus replication, including roles in early entry. VP1-2 contains an N-terminal basic motif which functions as an efficient nuclear localization signal (NLS). In this study, we constructed a mutant HSV strain, K.VP1-2ΔNLS, which contains a 7-residue deletion of the core NLS at position 475. This mutant fails to spread in normal cells but can be propagated in complementing cell lines. Electron microscopy (EM) analysis of infection in noncomplementing cells demonstrated capsid assembly, cytoplasmic envelopment, and the formation of extracellular enveloped virions. Furthermore, extracellular virions isolated from noncomplementing cells had similar profiles and abundances of structural proteins. Virions containing VP1-2ΔNLS were able to enter and be transported within cells. However, further progress of infection was prevented, with at least a 500- to 1,000-fold reduction in the efficiency of initiating gene expression compared to that in the revertant. Ultrastructural and immunofluorescence analyses revealed that the K.VP1-2ΔNLS mutant was blocked at the microtubule organizing center or immediately upstream of nuclear pore docking and prior to gene expression. These results indicate that the VP1-2 NLS is not required for the known assembly functions of the protein but is a key requirement for the early routing to the nuclear pore that is necessary for successful infection. Given its conservation, we propose that this motif may also be critical for entry of other classes of herpesviruses.
Inhibition of Human Cytomegalovirus Immediate-Early Gene Expression by Cyclin A2-Dependent Kinase Activity
Authors: Oduro Jennifer D. , Uecker Ralf , Hagemeier Christian , Wiebusch Lüder 
Published: June 2012

ABSTRACT

Human cytomegalovirus (HCMV) starts its lytic replication cycle only in the G
0
/G
1
phase of the cell division cycle. S/G
2
cells can be infected but block the onset of immediate-early (IE) gene expression. This block can be overcome by inhibition of cyclin-dependent kinases (CDKs), suggesting that cyclin A2, the only cyclin with an S/G
2
-specific activity profile, may act as a negative regulator of viral gene expression. To directly test this hypothesis, we generated derivatives of an HCMV-permissive glioblastoma cell line that express cyclin A2 in a constitutive, cell cycle-independent manner. We demonstrate that even moderate cyclin A2 overexpression in G
1
was sufficient to severely compromise the HCMV replicative cycle after high-multiplicity infection. This negative effect was composed of a strong but transient inhibition of IE gene transcription and a more sustained alteration of IE mRNA processing, resulting in reduced levels of UL37 and IE2, an essential transactivator of viral early gene expression. Consistently, cyclin A2-overexpressing cells showed a strong delay of viral early and late gene expression, as well as virus reproduction. All effects were dependent on CDK activity, as a cyclin A2 mutant deficient in CDK binding was unable to interfere with the HCMV infectious cycle. Interestingly, murine CMV, whose IE gene expression is known to be cell cycle independent, is not affected by cyclin A2. Instead, it upregulates cyclin A2-associated kinase activity upon infection. Understanding the mechanisms behind the HCMV-specific action of cyclin A2-CDK might reveal new targets for antiviral strategies.
Clinicopathological significance of CADM4 expression, and its correlation with expression of E-cadherin and Ki-67 in colorectal adenocarcinomas
Authors: Jang Se Min , Han Hulin , Jun Young Jin , Jang Si-Hyong , Min Kyueng-Whan , Sim Jongmin , Ahn Hye In , Lee Kang Hong , Jang Ki-Seok , Paik Seung Sam 
Published: June 2012

AimsCell adhesion molecule 4 (CADM4) is a novel tumour suppressor. The purpose of this study was to investigate the correlation between its expression and the expression of E-cadherin and Ki-67 in colorectal adenocarcinomas, as well as its effect on patient survival.MethodsWe evaluated CADM4 expression in tissue microarrays of 513 colorectal adenocarcinomas by immunohistochemistry.ResultsCADM4 was highly expressed in 210 of the 513 colorectal adenocarcinomas; expression was reduced in 185 cases and absent in the remaining cases. Loss of CADM4 expression was correlated with larger tumour size (6.2±2.1 cm vs 5.3±2.0 cm, p<0.001), mucinous tumour type (61.5% vs 20.9%, p<0.001), lymph node metastasis (31.4% vs 20.9%, p=0.022), higher Dukes stage (25.5% vs 19.6%, p=0.044), poorer differentiation (38.5% vs 18.8%, p<0.001), absence of E-cadherin expression (28.5% vs 16.0%, p=0.007) and presence of Ki-67 expression (27.3% vs 12.3%, p<0.001). In univariable Cox regression analysis, absence of CADM4 expression was associated with poorer overall survival (HR 0.712; 95% CI 0.512 to 0.989, p=0.042) and disease-free survival (HR 0.732; 95% CI 0.546 to 0.981, p=0.037). In multivariate analysis with the Cox proportional hazards model, CADM4 expression was not an independent prognostic factor of overall survival (HR 0.726; 95% CI 0.516 to 1.021, p=0.066) and disease-free survival (HR 0.762; 95% CI 0.563 to 1.033, p=0.080).ConclusionsLoss of CADM4 expression is relatively frequent in colorectal adenocarcinomas and may play an important role in cancer progression and patient survival.
Pathology and prognosis in pseudomyxoma peritonei: a review of 274 cases
Authors: Carr Norman J , Finch Jenny , Ilesley Ian Charles , Chandrakumaran Kandiah , Mohamed Faheez , Mirnezami Alex , Cecil Tom , Moran Brendan 
Published: June 2012

AimsThe classification of abdominal mucinous neoplasia is a controversial area. In 2010, WHO published a classification which divides pseudomyxoma peritonei (PMP) into low and high grades. The aim of the authors was to correlate this classification with the prognosis and site of primary neoplasm.MethodsThe authors reviewed 274 patients with PMP who had undergone surgery at a single institution and classified them according to WHO criteria. The findings were correlated with clinical information and survival data.ResultsPMP was low grade in 78% of patients and high grade in 22%. The appendix accounted for 94% of lesions, and the most common primary tumour was a low grade appendiceal mucinous neoplasm. Colorectal primaries were more likely to be associated with high grade PMP. There was an excellent correlation between the grade of the PMP and the primary neoplasm; only two cases showed discordant morphology: both were high grade appendiceal adenocarcinomas that were associated with low grade PMP. Nodal metastases were more likely in high grade lesions, but there was no significant difference in the rate of parenchymal organ invasion between low grade and high grade. Low grade morphology was associated with significantly longer survival than high grade (overall 5-year survival of 63% for low grade and 23% for high grade).ConclusionsCategorisation as either low grade or high grade by WHO criteria correlates with prognosis. The grade of the PMP is generally consistent with the grade of the primary neoplasm. Colorectal primaries are more likely to be associated with high grade PMP.