bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of an Insulator in AAVS1, a Preferred Region for Integration of Adeno-Associated Virus DNA
Authors: Ogata Toshihiko , Kozuka Takuyo , Kanda Tadahito 
Published: July 2003

ABSTRACT
In latent adeno-associated virus (AAV) infection, the viral genome is integrated preferentially into the human chromosome 19 q arm at a specific region designated AAVS1, which has an open chromatin conformation as indicated by the presence of a DNase I-hypersensitive site (DHS-S1). We examined whether an insulator, which defines the domain of gene expression by directionally blocking the action of enhancers and by preventing the spread of heterochomatin, is present near the DHS-S1 in the middle of a 2.6-kbp AAVS1-related DNA fragment used in this study. The fragment, cloned into an Epstein-Barr virus (EBV)-based eukaryotic episomal plasmid, was introduced into HEK293 cells. The DHS-S1 on the plasmid replicating in the nuclei was hypersensitive to DNase I digestion, and thus, the EBV plasmid system was used in an enhancer-blocking assay with the 2.6-kbp DNA and two shortened DNAs, of 1.6 kbp and 336 bp, containing DHS-S1. The three DNA fragments, when inserted in the proper direction between the cytomegalovirus immediate-early enhancer and minimal promoter, repressed the expression of a reporter gene. Thus, the enhancer-blocking activity was located within the 336-bp DNA containing the entire region (300 bp) of DHS-S1. To investigate the prevention of repression caused by heterochromatin, a transgene-expressing cassette flanked by the two 336-bp DNAs placed in the enhancer-blocking direction was introduced into HEK293 and HeLa cells. All the cell clones examined with the cassette integrated into cell DNA continued to express the transgene, which indicates that the pair of 336-bp DNA apparently prevented the spread of heterochromatin. The results show that an insulator lies between nucleotides 17 and 354 near the DHS-S1 in AAVS1. In a gel shift test, the 336-bp DNA did not bind an in vitro-prepared CCCTC-binding factor that binds to the chicken β-globin insulator, suggesting that the AAVS1 insulator requires an as yet unidentified binding protein. The newly identified AAVS1 insulator is likely to contribute to the maintenance of an open chromatin conformation that affects the life cycle of AAV.
Intracellular Trafficking of a Palmitoylated Membrane-Associated Protein Component of Enveloped Vaccinia Virus
Authors: Husain Matloob , Moss Bernard 
Published: July 2003

ABSTRACTThe F13L protein of vaccinia virus, an essential and abundant palmitoylated peripheral membrane component of intra- and extracellular enveloped virions, associates with Golgi, endosomal, and plasma membranes in the presence or absence of other viral proteins. In the present study, the trafficking of a fully functional F13L-green fluorescent protein (GFP) chimera in transfected and productively infected cells was analyzed using specific markers and inhibitors. We found that Sar1H79G, atrans-dominant-negative protein inhibitor of cargo transport from the endoplasmic reticulum, had no apparent effect on the intracellular distribution of F13L-GFP, suggesting that the initial membrane localization occurs at a downstream compartment of the secretory pathway. Recycling of F13L-GFP from the plasma membrane was demonstrated by partial colocalization with FM4-64, a fluorescent membrane marker of endocytosis. Punctate F13L-GFP fluorescence overlapped with clathrin and Texas red-conjugated transferrin, suggesting that endocytosis occurred via clathrin-coated pits. The inhibitory effects of chlorpromazine andtrans-dominant-negative forms of dynamin and Eps15 protein on the recycling of F13L-GFP provided further evidence for clathrin-mediated endocytosis. In addition, the F13L protein was specifically coimmunoprecipitated with α-adaptin, a component of the AP-2 complex that interacts with Eps15. Nocodazole and wortmannin perturbed the intracellular trafficking of F13L-GFP, consistent with its entry into late and early endosomes through the secretory and endocytic pathways, respectively. The recycling pathway described here provides a mechanism for the reutilization of the F13L protein following its deposition in the plasma membrane during the exocytosis of enveloped virions.
Identification of a C-Terminal Regulatory Motif in Hepatitis C Virus RNA-Dependent RNA Polymerase: Structural and Biochemical Analysis
Authors: Lévêque Vincent J.-P. , Johnson Robert B. , Parsons Stephen , Ren Jianxin , Xie Congping , Zhang Faming , Wang Q. May 
Published: July 2003

ABSTRACT
The NS5B RNA-dependent RNA polymerase encoded by the hepatitis C virus (HCV) is a key component of the viral replicase. Reported here is the three-dimensional structure of HCV NS5B polymerase, with the highlight on its C-terminal folding, determined by X-ray crystallography at 2.1-Å resolution. Structural analysis revealed that a stretch of C-terminal residues of HCV NS5B inserted into the putative RNA binding cleft, where they formed a hydrophobic pocket and interacted with several important structural elements. This region was found to be conserved and unique to the RNA polymerases encoded by HCV and related viruses. Through biochemical analyses, we confirmed that this region interfered with the binding of HCV NS5B to RNA. Deletion of this fragment from HCV NS5B enhanced the RNA synthesis rate up to ∼50-fold. These results provide not only direct experimental insights into the role of the C-terminal tail of HCV NS5B polymerase but also a working model for the RNA synthesis mechanism employed by HCV and related viruses.
Major Histocompatibility Complex Class I Alleles Associated with Slow Simian Immunodeficiency Virus Disease Progression Bind Epitopes Recognized by Dominant Acute-Phase Cytotoxic-T-Lymphocyte Responses
Authors: O'Connor David H. , Mothe Bianca R. , Weinfurter Jason T. , Fuenger Sarah , Rehrauer William M. , Jing Peicheng , Rudersdorf Richard R. , Liebl Max E. , Krebs Kendall , Vasquez Joshua , Dodds Elizabeth , Loffredo John , Martin Sarah , McDermott Adrian B. , Allen Todd M. , Wang Chenxi , Doxiadis G. G. , Montefiori David C. , Hughes Austin , Burton Dennis R. , Allison David B. , Wolinsky Steven M. , Bontrop Ronald , Picker Louis J. , Watkins David I. 
Published: July 2003

ABSTRACTCertain major histocompatibility complex class I (MHC-I) alleles are associated with delayed disease progression in individuals infected with human immunodeficiency virus (HIV) and in macaques infected with simian immunodeficiency virus (SIV). However, little is known about the influence of these MHC alleles on acute-phase cellular immune responses. Here we follow 51 animals infected with SIVmac239 and demonstrate a dramatic association between Mamu-A*01 and -B*17 expression and slowed disease progression. We show that the dominant acute-phase cytotoxic T lymphocyte (CTL) responses in animals expressing these alleles are largely directed against two epitopes restricted by Mamu-A*01 and one epitope restricted by Mamu-B*17. One Mamu-A*01-restricted response (Tat28-35SL8) and the Mamu-B*17-restricted response (Nef165-173IW9) typically select for viral escape variants in early SIVmac239 infection. Interestingly, animals expressing Mamu-A*1 and -B*17 have less variation in the Tat28-35SL8 epitope during chronic infection than animals that express only Mamu-A*01. Our results show that MHC-I alleles that are associated with slow progression to AIDS bind epitopes recognized by dominant CTL responses during acute infection and underscore the importance of understanding CTL responses during primary HIV infection.
Distinct Roles of Cellular Lck and p80 Proteins in Herpesvirus Saimiri Tip Function on Lipid Rafts
Authors: Park Junsoo , Cho Nam-Hyuk , Choi Joong-Kook , Feng Pinghui , Choe Joonho , Jung Jae U. 
Published: July 2003

ABSTRACT
Lipid rafts are proposed to function as platforms for both receptor signaling and trafficking. Following interaction with antigenic peptides, the T-cell receptor (TCR) rapidly translocates to lipid rafts, where it transmits signals and subsequently undergoes endocytosis. The Tip protein of herpesvirus saimiri (HVS), which is a T-lymphotropic tumor virus, interacts with cellular Lck tyrosine kinase and p80, a WD domain-containing endosomal protein. Interaction of Tip with p80 induces enlarged vesicles and recruits Lck and TCR complex into these vesicles for trafficking. We report here that Tip is constitutively present in lipid rafts and that Tip interaction with p80 but not with Lck is necessary for its efficient localization in lipid rafts. The Tip-Lck interaction was required for recruitment of the TCR complex to lipid rafts, and the Tip-p80 interaction was critical for the aggregation and internalization of lipid rafts. These results suggest the potential mechanism for Tip-mediated TCR downregulation: Tip interacts with Lck to recruit TCR complex to lipid rafts, and it subsequently interacts with p80 to initiate the aggregation and internalization of the lipid raft domain and thereby downregulate the TCR complex. Thus, the signaling and targeting functions of HVS Tip rely on two functionally and genetically separable mechanisms that independently target cellular Lck tyrosine kinase and p80 endosomal protein.
Redistribution of Cyclophilin A to Viral Factories during Vaccinia Virus Infection and Its Incorporation into Mature Particles
Authors: Castro Ana Paula V. , Carvalho Técia M. U. , Moussatché Nissin , Damaso Clarissa R. A. 
Published: July 2003

ABSTRACTCyclophilins are peptidyl-prolylcis-transisomerases involved in catalyzing conformational changes and accelerating the rate of protein folding and refolding in several cellular systems. In the present study, we analyzed the expression pattern and intracellular distribution of the cellular isomerase cyclophilin A (CypA) during vaccinia virus (VV) infection. An impressive increase in CypA stability was observed, leading to a practically unchanged accumulation of CypA during infection, although its synthesis was completely inhibited at late times. By confocal microscopy, we observed that CypA went through an intense reorganization in the cell cytoplasm and colocalized with the virosomes late in infection. CypA relocation to viral factories required the synthesis of viral postreplicative proteins, and treatment of infected cells with cyclosporine (CsA) prevented CypA relocation, clearly excluding the virosomes from CypA staining. Immunoelectron microscopy of VV-infected cells showed that CypA was incorporated into VV particles during morphogenesis. Biochemical and electron microscopic assays with purified virions confirmed that CypA was encapsidated within the virus particle and localized specifically in the core. This work suggests that CypA may develop an important role in VV replication.
Role of the Human Immunodeficiency Virus Type 1 Envelope Gene in Viral Fitness
Authors: Rangel Hector R. , Weber Jan , Chakraborty Bikram , Gutierrez Arantxa , Marotta Michael L. , Mirza Muneer , Kiser Patti , Martinez Miguel A. , Este Jose A. , Quiñones-Mateu Miguel E. 
Published: July 2003

ABSTRACT

A human host offers a variety of microenvironments to the infecting human immunodeficiency virus type 1 (HIV-1), resulting in various selective pressures, most of them directed against the envelope (
env
) gene. Therefore, it seems evident that the replicative capacity of the virus is largely related to viral entry. In this study we have used growth competition experiments and TaqMan real-time PCR detection to measure the fitness of subtype B HIV-1 primary isolates and autologous
env
-recombinant viruses in order to analyze the contribution of wild-type
env
sequences to overall HIV-1 fitness. A significant correlation was observed between fitness values obtained for wild-type HIV-1 isolates and those for the corresponding
env
-recombinant viruses (
r
= 0.93;
P
= 0.002). Our results suggest that the
env
gene, which is linked to a myriad of viral characteristics (e.g., entry into the host cell, transmission, coreceptor usage, and tropism), plays a major role in fitness of wild-type HIV-1. In addition, this new recombinant assay may be useful for measuring the contribution of HIV-1
env
to fitness in viruses resistant to novel antiretroviral entry inhibitors.
Pseudorabies Virus US3 Protein Kinase Mediates Actin Stress Fiber Breakdown
Authors: Minnebruggen Geert Van , Favoreel Herman W. , Jacobs Liesbeth , Nauwynck Hans J. 
Published: July 2003

ABSTRACT
Disruption of specific components of the host cytoskeleton has been reported for several viruses and is thought to be beneficial for viral replication and spread. Our previous work demonstrated that infection of swine kidney (SK-6) cells with pseudorabies virus (PRV), a swine alphaherpesvirus, induced actin stress fiber breakdown. In the present study, using several PRV deletion mutants, we found that the US3 serine/threonine (S/T) protein kinase is involved in breakdown of actin stress fibers in different PRV-infected cell lines. Further, by transfection assays, we showed that PRV US3 itself, in the absence of other viral proteins, is able to trigger actin stress fiber breakdown when it is localized in sufficient amounts in the nucleus.
TT Virus Loads and Lymphocyte Subpopulations in Children with Acute Respiratory Diseases
Authors: Maggi Fabrizio , Pifferi Massimo , Tempestini Elena , Fornai Claudia , Lanini Letizia , Andreoli Elisabetta , Vatteroni Marialinda , Presciuttini Silvano , Pietrobelli Angelo , Boner Attilio , Pistello Mauro , Bendinelli Mauro 
Published: July 2003

ABSTRACT
TT virus (TTV) produces chronic plasma viremia in around 90% of healthy individuals of all ages and has, therefore, been proposed as a commensal human virus. We recently demonstrated that in children hospitalized for acute respiratory diseases high TTV loads were associated with severe forms of disease. Here, we report that in such children TTV loads showed an inverse correlation with the percentage of circulating total T and helper T cells and a direct correlation with the percentage of B cells. Thus, florid TTV replication might contribute to lymphocyte imbalances and, possibly, immunosuppressive effects, thus resembling related animal viruses.
Recombinant Avian Infectious Bronchitis Virus Expressing a Heterologous Spike Gene Demonstrates that the Spike Protein Is a Determinant of Cell Tropism
Authors: Casais Rosa , Dove Brian , Cavanagh David , Britton Paul 
Published: July 2003

ABSTRACT

A recombinant infectious bronchitis virus (IBV), BeauR-M41(S), was generated using our reverse genetics system (R. Casais, V. Thiel, S. G. Siddell, D. Cavanagh, and P. Britton, J. Virol.
75:
12359-12369, 2001), in which the ectodomain region of the spike gene from IBV M41-CK replaced the corresponding region of the IBV Beaudette genome. BeauR-M41(S) acquired the same cell tropism phenotype as IBV M41-CK in four different cell types, demonstrating that the IBV spike glycoprotein is a determinant of cell tropism.
Expression of Immunogenic S1 Glycoprotein of Infectious Bronchitis Virus in Transgenic Potatoes
Authors: Zhou Ji-Yong , Wu Jian-Xiang , Cheng Li-Qin , Zheng Xiao-Juan , Gong Hui , Shang Shao-Bin , Zhou En-Min 
Published: July 2003

ABSTRACT

The expression of infectious bronchitis virus (IBV) S1 glycoprotein in potatoes and its immunogenicity in mice and chickens were investigated. Potato plants were genetically transformed with a cDNA construct encoding the IBV S1 glycoprotein with the
Agrobacterium
system. Genomic DNA and mRNA analyses of the transformed plantlets confirmed the integration of the foreign cDNA into the potato genome, as well as its transcription. Mice and chickens vaccinated with the expressed IBV S1 glycoprotein produced antibodies that neutralized IBV infectivity. After three immunizations, vaccinated chickens were completely protected from virulent IBV infection. These results demonstrate that transgenic potatoes expressing IBV S1 glycoprotein can be used as a source of recombinant antigen for vaccine production.
Vaccinia Virus Recombinant Expressing an 87-Kilodalton Polyprotein That Is Sufficient To Form Astrovirus-Like Particles
Authors: Dalton Rosa M. , Pastrana Esperanza P. , Sánchez-Fauquier Alicia 
Published: July 2003

ABSTRACT
Human astrovirus is an important cause of acute gastroenteritis. We have generated, for the first time, a vaccinia virus recombinant expressing the astrovirus 87-kDa structural polyprotein. The results demonstrate that this expression results in the formation of virus-like particles in the absence of other astrovirus proteins and genomic RNA. The purified trypsin-activated virus-like particles strongly resemble the complete astrovirus particles.
Novel Gamma-1 Herpesviruses Identified in Free-Ranging New World Monkeys (Golden-Handed Tamarin [Saguinus midas], Squirrel Monkey [Saimiri sciureus], and White-Faced Saki [Pithecia pithecia]) in French Guiana
Authors: de Thoisy Benoit , Pouliquen Jean-François , Lacoste Vincent , Gessain Antoine , Kazanji Mirdad 
Published: July 2003

ABSTRACTThe recent finding of a novel Epstein-Barr virus-related lymphocryptovirus (CalHV-3) in a captive colony of common marmoset (Callithrix jacchus) in the United States modifies the view that the host range of lymphocryptovirus is restricted to humans and Old World primates. We investigated the presence of Epstein-Barr virus-related viruses in 79 samples of New World monkeys caught in the wild, including six species of theCebidaefamily and one of theCallitrichidae, living in the rain forest of French Guiana. Using a degenerate consensus PCR method for the herpesvirus DNA polymerase gene, we identified three novel lymphocryptoviruses from golden-handed tamarin (Saguinus midas) of theCallitrichidaefamily and squirrel monkey (Saimiri sciureus) and white-faced saki (Pithecia pithecia) of theCebidaefamily. With the CalHV-3 strain, these three novel viruses constitute a well-supported phylogenetic clade in theLymphocryptovirusgenus, which is clearly distinct from the lineage of Old World lymphocryptovirus, hosted by catarrhine monkeys and humans. In tamarins, the prevalence of the novel lymphocryptovirus was more than 50%, indicating that it circulates well in the wild population, perhaps due to specific ecoethological patterns such as confrontations and intergroup migration. The detection and partial molecular characterization of the polymerase gene of three novelGamma-1-Herpesvirinaefrom New World monkeys caught in the wild clearly indicate that free-ranging populations of platyrrhine are natural hosts of lymphocryptoviruses. Further characterization of these novel viruses will provide new insight not only into the origin and evolution ofGammaherpesvirinaebut also into their pathogenicity.
Risk of hypospadias in relation to maternal occupational exposure to potential endocrine disrupting chemicals
Authors: Vrijheid M , Armstrong B , Dolk H , van Tongeren M , Botting B 
Published: July 2003

Background:Reported rises in the prevalence of hypospadias and other abnormalities of the male reproductive system may be a result of exposure to endocrine disrupting chemicals.Aims:To analyse the relation between risk of hypospadias and maternal occupation, particularly with regard to exposure to potential endocrine disrupting chemicals (EDCs).Methods:Data (1980–96) from the National Congenital Anomaly System (NCAS) were used to analyse the proportion of all congenital anomaly cases (n = 35 962) which were notified with hypospadias (n = 3471) by occupational codes (348 individual job titles) and by categories of exposure to potential EDCs from a job exposure matrix.Results:Five individual occupations (of 348) showed nominally statistically significant excesses, none of which had possible or probable exposure to potential EDCs. Odds ratios for “possible” or “probable” compared to “unlikely” exposure to potential EDCs did not show statistically significant increases in any of the EDC categories after adjustment for social class of the mother and father, nor was there evidence of an upward trend in risk with likelihood of exposure. In the 1992–96 time period odds ratios were increased for hairdressers (the largest group exposed to potential EDCs) and for probable exposure to phthalates (of which hairdressers form the largest group) before social class adjustment.Conclusions:There was little evidence for a relation between risk of hypospadias and maternal occupation or occupational exposure to potential EDCs, but as the exposure classification was necessarily crude, these findings should be interpreted with caution.
Associations of lead biomarkers with renal function in Korean lead workers
Authors: Weaver V M , Lee B-K , Ahn K-D , Lee G-S , Todd A C , Stewart W F , Wen J , Simon D J , Parsons P J , Schwartz B S 
Published: July 2003

Aims:To compare associations of lead biomarkers with renal function in current and former lead workers.Methods:Cross sectional analysis of first year results from a longitudinal study of 803 lead workers and 135 controls in South Korea. Clinical renal function was assessed by blood urea nitrogen (BUN), serum creatinine, and measured and calculated creatinine clearance. Urinary N-acetyl-β-D-glucosaminidase (NAG) and retinol-binding protein were also measured.Results:Mean (SD) tibia lead, blood lead, and DMSA chelatable lead levels in lead workers were 37.2 (40.4) μg/g bone mineral, 32.0 (15.0) μg/dl, and 767.8 (862.1) μg/g creatinine, respectively. Higher lead measures were associated with worse renal function in 16/42 models. When influential outliers were removed, higher lead measures remained associated with worse renal function in nine models. An additional five associations were in the opposite direction. Effect modification by age was observed. In 3/16 models, associations between higher lead measures and worse clinical renal function in participants in the oldest age tertile were significantly different from associations in those in the youngest age tertile which were in the opposite direction. Mean urinary cadmium (CdU) was 1.1 μg/g creatinine (n = 191). Higher CdU levels were associated with higher NAG.Conclusions:These data suggest that lead has an adverse effect on renal function in the moderate dose range, particularly in older workers. Associations between higher lead measures and lower BUN and serum creatinine and higher creatinine clearances may represent lead induced hyperfiltration. Environmental cadmium may also have an adverse renal impact, at least on NAG.