bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tpr Homologs inTreponema paraluiscuniculiCuniculi A Strain
Authors: Giacani Lorenzo , Sun Eileen S. , Hevner Karin , Molini Barbara J. , Van Voorhis Wesley C. , Lukehart Sheila A. , Centurion-Lara Arturo 
Published: October 2004

ABSTRACTTreponema paraluiscuniculi, the etiologic agent of rabbit venereal syphilis, is morphologically indistinguishable fromTreponema pallidumsubsp.pallidum(T. pallidum), the human syphilis treponeme, and induces similar immune responses and histopathologic changes in the infected host. Because of their high degree of relatedness, comparative studies are likely to identify genetic determinants that contribute to pathogenesis or virulence in human syphilis. Thetpr(Treponema pallidumrepeat) genes are believed to code for potential virulence factors. In this study, we identified 10tprhomologs inTreponema paraluiscuniculiCuniculi A strain and determined their sequence architecture. Half of this group of paralogous genes were predicted to be nonfunctional due to the presence of frameshifts and premature stop codons. Furthermore, the immune response against theT. paraluiscuniculiTpr homologs in long-term-infected rabbits was studied by enzyme-linked immunosorbent assay and lymphocyte proliferation assay, showing that TprK is the only target of the antibody and T-cell responses during experimental infection and emphasizing the importance of this putative virulence factor in venereal treponematosis.
Effects of
vlsE
Complementation on the Infectivity of
Borrelia burgdorferi
Lacking the Linear Plasmid lp28-1
Authors: Lawrenz Matthew B. , Wooten R. Mark , Norris Steven J. 
Published: October 2004

ABSTRACT

The loss of linear plasmid lp28-1, which contains the
vls
antigenic variation locus, is associated with reduced infectivity of
Borrelia burgdorferi
in immunocompetent mice. The recombinant shuttle vector pBBE22, which includes the virulence determinant BBE22 from lp25 and restores infectivity to readily transformable
B. burgdorferi
lacking lp25 and lp56, was used to determine the effect of
trans
expression of
vlsE
on virulence. Spirochetes lacking lp28-1 were complemented with the plasmid pBBE22:vlsE, containing both BBE22 and
vlsE
. VlsE protein produced by this construct was expressed and surface accessible in in vitro-cultured
B. burgdorferi
, as determined by surface proteolysis and immunoblot analysis. Clones lacking lp25 but containing lp28-1 and either pBBE22 or pBBE22:vlsE were reisolated consistently from immunocompetent mice 8 weeks after infection. In contrast, a clone lacking both lp25 and lp28-1 and complemented with pBBE22:vlsE was isolated from only a single tissue of one of six C3H/HeN mice 8 weeks postinfection. These results indicate that either an intact
vls
antigenic variation locus or another determinant on lp28-1 is required to restore complete infectivity. In addition, an isogenic clone that retained lp28-1 was complemented with the
vlsE
shuttle plasmid and was examined for
vlsE
sequence variation and infectivity. Sequence variation was not observed for the shuttle plasmid, indicating that the
cis
arrangement of
vlsE
and the
vls
silent cassettes in lp28-1 facilitate
vlsE
gene conversion. Lack of
vlsE
sequence variation on the shuttle plasmid thus did not result in clearance of the
trans
-complemented strain in immunocompetent mice under the conditions tested.
Distinct Trafficking and Localization of STEVOR Proteins in Three Stages of the
Plasmodium falciparum
Life Cycle
Authors: McRobert Louisa , Preiser Peter , Sharp Sarah , Jarra William , Kaviratne Mallika , Taylor Martin C. , Renia Laurent , Sutherland Colin J. 
Published: October 2004

ABSTRACT

The genome of
Plasmodium falciparum
harbors three extensive multigene families,
var
,
rif
, and
stevor
(for subtelomeric variable open reading frame), located mainly in the subtelomeric regions of the parasite's 14 chromosomes. STEVOR variants are known to be expressed in asexual parasites, but no function has as yet been ascribed to this protein family. We have examined the expression of STEVOR proteins in intraerythrocytic sexual stages, gametocytes, and extracellular sporozoites isolated from infected
Anopheles
mosquitoes. In gametocytes,
stevor
transcripts appear transiently early in development but STEVOR proteins persist for several days and are transported out of the parasite, travel through the host cell cytoplasm, and localize to the erythrocyte plasma membrane. In contrast to asexual parasites, gametocytes move STEVOR to the periphery via a trafficking pathway independent of Maurer's clefts. In sporozoites, STEVOR appear dispersed throughout the cytoplasm in vesicle-like structures. The pattern of STEVOR localization we have observed in gametocytes and sporozoites differs significantly from that in asexual parasite stages. STEVOR variants are therefore likely to perform different functions in each stage of the parasites life cycle in which they occur.
Prolonged Toll-Like Receptor Signaling byMycobacterium tuberculosisand Its 19-Kilodalton Lipoprotein Inhibits Gamma Interferon-Induced Regulation of Selected Genes in Macrophages
Authors: Pai Rish K. , Pennini Meghan E. , Tobian Aaron A. R. , Canaday David H. , Boom W. Henry , Harding Clifford V. 
Published: October 2004

ABSTRACTInfection of macrophages withMycobacterium tuberculosisor exposure toM. tuberculosis19-kDa lipoprotein for >16 h inhibits gamma interferon (IFN-γ)-induced major histocompatibility complex class II (MHC-II) expression by a mechanism involving Toll-like receptors (TLRs).M. tuberculosiswas found to inhibit murine macrophage MHC-II antigen (Ag) processing activity induced by IFN-γ but not by interleukin-4 (IL-4), suggesting inhibition of IFN-γ-induced gene regulation. We designed an approach to test the ability ofM. tuberculosis-infected cells to respond to IFN-γ. To model chronic infection withM. tuberculosiswith accompanying prolonged TLR signaling, macrophages were infected withM. tuberculosisor incubated withM. tuberculosis19-kDa lipoprotein for 24 h prior to the addition of IFN-γ. Microarray gene expression studies were then used to determine whether prolonged TLR signaling byM. tuberculosisbroadly inhibits IFN-γ regulation of macrophage gene expression. Of 347 IFN-γ-induced genes,M. tuberculosisand 19-kDa lipoprotein inhibited induction of 42 and 36%, respectively. Key genes or gene products were also examined by quantitative reverse transcription-PCR and flow cytometry, confirming and extending the results obtained by microarray studies.M. tuberculosisinhibited IFN-γ induction of genes involved in MHC-II Ag processing, Ag presentation, and recruitment of T cells. These effects were largely dependent on myeloid differentiation factor 88, implying a role for TLRs. Thus, prolonged TLR signaling byM. tuberculosisinhibits certain macrophage responses to IFN-γ, particularly those related to MHC-II Ag presentation. This inhibition may promoteM. tuberculosisevasion of T-cell responses and persistence of infection in tuberculosis.
Toll-Like Receptor 4-Dependent Early Elicited Tumor Necrosis Factor Alpha Expression Is Critical for Innate Host Defense againstBordetella bronchiseptica
Authors: Mann Paul B. , Elder Kelly D. , Kennett Mary J. , Harvill Eric T. 
Published: October 2004

ABSTRACTToll-like receptor 4 (TLR4) mediates the response to lipopolysaccharide, and its activation induces the expression of a large number of inflammatory genes, many of which are also induced by other pathogen-associated molecular patterns. Interestingly, the subset of genes that are dependent on TLR4 for optimal expression during gram-negative bacterial infection has not been determined. We have previously shown that TLR4-deficient mice rapidly develop acute pneumonia after inoculation withBordetella bronchiseptica, suggesting that TLR4 is required for expression of early elicited gene products in this model. Microarray analysis with macrophages derived from wild-type and TLR4-deficient mice was used to identify genes whose expression, within 1 h of bacterial exposure, is dependent on TLR4. The results of this investigation suggest that TLR4 is not required for the majority of the transcriptional response toB. bronchiseptica. However, early tumor necrosis factor alpha (TNF-α) mRNA expression is primarily dependent on TLR4 and in vitro and in vivo protein levels substantiate this finding. TLR4-deficient mice and TNF-α−/−mice are similarly susceptible to infection with relatively low doses ofB. bronchisepticaand in vivo neutralization studies indicate that it is the TLR4-dependent early elicited TNF-α response that is critical for preventing severe pneumonia and limiting bacterial growth. These results suggest that one critical role for TLR4 is the generation of a robust but transient TNF-α response that is critical to innate host defense during acute gram-negative respiratory infection.
Mucosal Administration of Flagellin Induces Innate Immunity in the Mouse Lung
Authors: Honko Anna N. , Mizel Steven B. 
Published: October 2004

ABSTRACTNonsurgical intratracheal instillation of 1 μg of purified, recombinant flagellin in several strains of mice stimulated a transient innate immune response in the lung characterized by the infiltration of neutrophils and the rapid production of tumor necrosis factor alpha, interleukin 6, granulocyte colony-stimulating factor, and the chemokines keratinocyte-derived chemokine, MIP1α, and MIP-2.
LcrV Synthesis Is Altered by DNA Adenine Methylase Overproduction in
Yersinia pseudotuberculosis
and Is Required To Confer Immunity in Vaccinated Hosts
Authors: Badie Golnaz , Heithoff Douglas M. , Mahan Michael J. 
Published: October 2004

ABSTRACT

Yersinia pseudotuberculosis
mutants that overproduce the DNA adenine methylase (Dam
OP
Yersinia
) are attenuated, confer robust protective immune responses, and synthesize or secrete several
Yersinia
outer proteins (Yops) under conditions that are nonpermissive for synthesis and secretion in wild-type strains. To understand the molecular basis of immunity elicited by Dam
OP
Yersinia
, we investigated the effects of Dam overproduction on the synthesis and localization of a principal
Yersinia
immunogen, LcrV, a low-calcium-responsive virulence factor involved in Yop synthesis, localization, and suppression of host inflammatory activities. Dam overproduction relaxed the stringent temperature and calcium regulation of LcrV synthesis. Moreover, the LcrV-dependent synthesis and localization of the actin cytotoxin, YopE, were shown to be relaxed in Dam
OP
cells, suggesting that the synthesis and localization of Yops can occur via both LcrV-dependent and -independent mechanisms. Last, the immunity conferred by Dam
OP
Yersinia
was strictly dependent on the presence of LcrV, which may result from its role (i) as an immunogen, (ii) as an immunomodulator of host anti-inflammatory activities, or (iii) in the altered synthesis and localization of Yops that could contribute to immunogen repertoire expansion.
Comparative Analysis of Locus of Enterocyte Effacement Pathogenicity Islands of Atypical Enteropathogenic
Escherichia coli
Authors: Gärtner Julia F. , Schmidt M. Alexander 
Published: October 2004

ABSTRACT

The pathogenicity of enteropathogenic
Escherichia coli
(EPEC) is linked to the locus of enterocyte effacement, or LEE, encoding a type III secretion system (T3SS) that directly transfers bacterial effector proteins into eukaryotic cells. Atypical diffusely adhering EPEC (DA-EPEC) strains that harbor homologues of the LEE but lack the EPEC adherence factor plasmid have been increasingly associated with outbreaks of diarrhea. In this study, we have completely sequenced and functionally characterized LEE pathogenicity islands derived from the clinical DA-EPEC isolates 3431 (O8:H
−
) and 0181 (O119:H9:K61). LEE
3431
and LEE
0181
exhibit genetic organization analogous to that of the prototype LEE
E2348/69
. Genes constituting the T3SS apparatus are highly conserved. However, LEE-encoded effector proteins exhibit major differences. Transfer and functional expression of LEE
0181
in an
E. coli
XL1 blue MR background demonstrated that LEE
0181
contains all the information for signal transduction and pedestal formation.
Lack of Stage-Specific Proteins in Coccoid
Helicobacter pylori
Cells
Authors: Bumann Dirk , Habibi Hajar , Kan Biao , Schmid Monika , Goosmann Christian , Brinkmann Volker , Meyer Thomas F. , Jungblut Peter R. 
Published: October 2004

ABSTRACT

Helicobacter pylori
exists in two distinct forms, rod shaped or coccoid, in stomachs of infected patients. Based on in vitro proteome comparisons, there are no detectable coccoid-specific proteins, which argues against the specific adaptation of coccoid
Helicobacter
to distinct biological functions, such as enhanced persistence or transmission to other hosts.
The Cauliflower Mosaic Virus 35S Promoter Extends into the Transcribed Region
Authors: Pauli Sandra , Rothnie Helen M. , Chen Gang , He Xiaoyuan , Hohn Thomas 
Published: October 2004

ABSTRACT
A 60-nucleotide region (S1) downstream of the transcription start site of the cauliflower mosaic virus 35S RNA can enhance gene expression. By using transient expression assays with plant protoplasts, this activity was shown to be at least partially due to the effect of transcriptional enhancers within this region. We identify sequence motifs with enhancer function, which are normally masked by the powerful upstream enhancers of the 35S promoter. A repeated CT-rich motif is involved both in enhancer function and in interaction with plant nuclear proteins. The S1 region can also enhance expression from heterologous promoters.
Expression and Localization of the Epstein-Barr Virus-Encoded Protein Kinase
Authors: Gershburg E. , Marschall M. , Hong K. , Pagano J. S. 
Published: October 2004

ABSTRACT

The protein kinase (PK) encoded by the Epstein-Barr Virus (EBV) BGLF4 gene is the only EBV protein kinase. The expression pattern of EBV PK during the reactivation of the viral lytic cycle and the subcellular localization of the protein were analyzed with a polyclonal antiserum raised against a peptide corresponding to the N terminus of EBV PK. Based on previously published data (E. Gershburg and J. S. Pagano, J. Virol.
76:
998-1003, 2002) and the expression pattern described here, we conclude that EBV PK is an early protein that requires viral-DNA replication for maximum expression. By biochemical fractionation, the protein could be detected mainly in the nuclear fraction 4 h after viral reactivation in Akata cells. Nuclear localization could be visualized by indirect immunofluorescence in HeLa cells transiently expressing EBV BGLF4 in the absence of other viral products. Transient expression of 3′-terminal deletion mutants of EBV BGLF4 resulted in cytoplasmic localization, confirming the presence of a nuclear localization site in the C-terminal region of the protein. In contrast to the wild-type EBV PK, all of the mutants were unable to hyperphosphorylate EA-D during coexpression or to phosphorylate ganciclovir, as measured by an in-cell activity assay. Thus, the results demonstrate that the nuclear localization, as well as the kinase activity, of BGFL4 is dependent on an intact C-terminal region.
A Retroviral Promoter and a Cellular Enhancer Define a Bipartite Element Which Controls
env
ERVWE1 Placental Expression
Authors: Prudhomme Sarah , Oriol Guy , Mallet François 
Published: October 2004

ABSTRACT
The HERV-W family contains hundreds of loci diversely expressed in several physiological and pathological contexts. A unique locus termed ERVWE1 encodes an envelope glycoprotein (syncytin) involved in hominoid placental physiology. Here we show that syncytin expression is regulated by a bipartite element consisting of a cyclic AMP (cAMP)-inducible long terminal repeat (LTR) retroviral promoter adjacent to a cellular enhancer conferring a high level of expression and placental tropism. Deletion mutant analysis showed that the ERVWE1 5′ LTR contains binding sites essential for basal placental activity in the region from positions +1 to +125. The region from positions +125 to +310 represents a cAMP-responsive core HERV-W promoter active in all cell types. Site-directed mutagenesis analysis highlighted the complexity of U3 regulation. ERVWE1 placenta-specific positive (e.g., T240) and negative (e.g., G71) regulatory sites were identified, as were essential sites required for basic activity (e.g., A247). The flanking sequences of the ERVWE1 provirus contain several putative regulatory elements. The upstream HERV-H and HERV-P LTRs were found to be inactive. Conversely, the 436-bp region located between the HERV-P LTR and ERVWE1 was shown to be an upstream regulatory element (URE) which is significantly active in placenta cells. This URE acts as a tissue-specific enhancer. Genetic and functional analyses of hominoid UREs revealed large differences between UREs of members of the Hominidae and the Hylobatidae. These data allowed the identification of a positive regulatory region from positions −436 to −128, a mammalian apparent LTR retrotransposon negative regulatory region from positions −128 to −67, and a trophoblast-specific enhancer (TSE) from positions −67 to −35. Putative AP-2, Sp-1, and GCMa binding sites are essential constituents of the 33-bp TSE.
De Novo Initiation Pocket Mutations Have Multiple Effects on Hepatitis C Virus RNA-Dependent RNA Polymerase Activities
Authors: Ranjith-Kumar C. T. , Sarisky R. T. , Gutshall L. , Thomson M. , Kao C. C. 
Published: October 2004

ABSTRACT

The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp) has several distinct biochemical activities, including initiation of RNA synthesis by a de novo mechanism, extension from a primed template, nontemplated nucleotide addition, and synthesis of a recombinant RNA product from two or more noncovalently linked templates (template switch). All of these activities require specific interaction with nucleoside triphosphates (NTPs). Based on the structure of the HCV RdRp bound to NTP (S. Bressanelli, L. Tomei, F. A. Rey, and R. DeFrancesco, J. Virol.
76:
3482-3492, 2002), we mutated the amino acid residues that contact the putative initiation GTP and examined the effects on the various activities. Although all mutations retained the ability for primer extension, alanine substitution at R48, R158, R386, R394, or D225 decreased de novo initiation, and two or more mutations abolished de novo initiation. While the prototype enzyme had a

K
m

for GTP of 3.5 μM, all of the mutations except one had

K
m

s that were three- to sevenfold higher. These results demonstrate that the affected residues are functionally required to interact with the initiation nucleotide. Unexpectedly, many of the mutations also affected the addition of nontemplated nucleotide, indicating that residues in the initiating NTP (NTPi)-binding pocket are required for nontemplated nucleotide additions. Interestingly, mutations in D225 are dramatically affected in template switch, indicating that this residue of the NTPi pocket also interacts with components in the elongation complex. We also examined the interaction of ribavirin triphosphate with the NTPi-binding site.
Conditionally Replicating Adenoviruses Kill Tumor Cells via a Basic Apoptotic Machinery-Independent Mechanism That Resembles Necrosis-Like Programmed Cell Death
Authors: Abou El Hassan Mohamed A. I. , van der Meulen-Muileman Ida , Abbas Saman , Kruyt Frank A. E. 
Published: October 2004

ABSTRACT

Conditionally replicating adenoviruses (CRAds) represent a promising class of novel anticancer agents that are used for virotherapy. The E1AΔ24 mutation-based viruses, Ad5-
Δ
24 [CRAd(E3−); E3 region deleted] and infectivity-enhanced Ad5-
Δ
24RGD [CRAd(E3+)] have been shown to potently eradicate tumor cells. The presence of the E3 region in the latter virus is known to improve cell killing that can be attributed to the presence of the oncolysis-enhancing Ad death protein. The more precise mechanism by which CRAds kill tumor cells is unclear, and the role of the host cell apoptotic machinery in this process has been addressed only in a limited way. Here, we examine the role of several major apoptotic pathways in the CRAd-induced killing of non-small-cell lung cancer H460 cells. As expected, CRAd(E3+) was more potent than CRAd(E3−). No evidence for the involvement of the p53-Bax apoptotic pathway was found. Western blot analyses demonstrated strong suppression of p53 expression and unchanged Bax levels during viral replication, and stable overexpression of human papillomavirus type 16-E6 in H460 cells did not affect killing by both CRAds. CRAd activity was also not hampered by stable overexpression of anti-apoptotic Bcl2 or BclXL, and endogenous Bcl2/BclXL protein levels remained constant during the oncolytic cycle. Some evidence for caspase processing was obtained at late time points after infection; however, the inhibition of caspases by the X-linked inhibitor of apoptosis protein overexpression or cotreatment with zVAD-fmk did not inhibit CRAd-dependent cell death. Analyses of several apoptotic features revealed no evidence for nuclear fragmentation or DNA laddering, although phosphatidylserine externalization was detected. We conclude that despite the known apoptosis-modulating abilities of individual Ad proteins, Ad5-
Δ
24-based CRAds trigger necrosis-like cell death. In addition, we propose that deregulated apoptosis in cancer cells, a possible drug resistance mechanism, provides no barrier for CRAd efficacy.
Multivesicular Bodies as a Platform for Formation of the Marburg Virus Envelope
Authors: Kolesnikova Larissa , Berghöfer Beate , Bamberg Sandra , Becker Stephan 
Published: October 2004

ABSTRACT
The Marburg virus (MARV) envelope consists of a lipid membrane and two major proteins, the matrix protein VP40 and the glycoprotein GP. Both proteins use different intracellular transport pathways: GP utilizes the exocytotic pathway, while VP40 is transported through the retrograde late endosomal pathway. It is currently unknown where the proteins combine to form the viral envelope. In the present study, we identified the intracellular site where the two major envelope proteins of MARV come together as peripheral multivesicular bodies (MVBs). Upon coexpression with VP40, GP is redistributed from the trans-Golgi network into the VP40-containing MVBs. Ultrastructural analysis of MVBs suggested that they provide the platform for the formation of membrane structures that bud as virus-like particles from the cell surface. The virus-like particles contain both VP40 and GP. Single expression of GP also resulted in the release of particles, which are round or pleomorphic. Single expression of VP40 led to the release of filamentous structures that closely resemble viral particles and contain traces of endosomal marker proteins. This finding indicated a central role of VP40 in the formation of the filamentous structure of MARV particles, which is similar to the role of the related Ebola virusVP40. In MARV-infected cells, VP40 and GP are colocalized in peripheral MVBs as well. Moreover, intracellular budding of progeny virions into MVBs was frequently detected. Taken together, these results demonstrate an intracellular intersection between GP and VP40 pathways and suggest a crucial role of the late endosomal compartment for the formation of the viral envelope.