bluuebunny/crossref_metadata_2025_split huggingface dataset:
Penicillium marneffeiCauses Osteopontin-Mediated Production of Interleukin-12 by Peripheral Blood Mononuclear Cells
Authors: Koguchi Yoshinobu , Kawakami Kazuyoshi , Kon Shigeyuki , Segawa Tatsuya , Maeda Masahiro , Uede Toshimitsu , Saito Atsushi 
Published: July 2002

ABSTRACTWe investigated the role of osteopontin (OPN) in interleukin-12 (IL-12) production from peripheral blood mononuclear cells (PBMCs) stimulated withPenicillium marneffei. Kinetic studies showed that OPN synthesis preceded that of IL-12 at both mRNA and protein levels when PBMCs were cocultured withP. marneffei. Treatment with anti-OPN monoclonal antibodies (MAb) significantly suppressed IL-12 secretion. Furthermore, native OPN induced a profound level of synthesis of IL-12 from noninfected PBMCs. The major cellular source of OPN was monocytes, because depletion of CD14+cells resulted in the abrogation of such production. We also examined the regulatory role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in OPN secretion fromP. marneffei-stimulated PBMCs. Neutralizing anti-GM-CSF MAb significantly reduced OPN secretion, and treatment with this cytokine induced OPN production from both infected and noninfected PBMCs. Finally, antagonists against the mannose receptor but not the β-glucan receptor almost completely abrogated the production of OPN. Our results demonstrated that OPN secreted from monocytes is involved in the production of IL-12 from PBMCs after stimulation withP. marneffeiand that OPN production is regulated by GM-CSF. Our results also indicated the possible involvement of the mannose receptor as a signal-transducing receptor for triggering the secretion of OPN byP. marneffei-stimulated PBMCs.
Safety and Immunogenicity of a Prototype EnterotoxigenicEscherichia coliVaccine Administered Transcutaneously
Authors: Güereña-Burgueño Fernando , Hall Eric R. , Taylor David N. , Cassels Frederick J. , Scott Daniel A. , Wolf Marcia K. , Roberts Zachary J. , Nesterova Galina V. , Alving Carl R. , Glenn Gregory M. 
Published: July 2002

ABSTRACTTranscutaneous immunization (TCI) is a new method for vaccine delivery that has been shown to induce immunity relevant to enteric disease vaccines. We evaluated the clinical safety and immunogenicity of a recombinant subunit vaccine against enterotoxigenicEscherichia coli(ETEC) delivered by TCI. Adult volunteers received patches containing the recombinant ETEC colonization factor CS6, either with heat-labile enterotoxin (LT) or patches containing CS6 alone. The vaccine was administered at 0, 1, and 3 months, and serum antibodies and antibody-secreting cells (ASCs) were assessed. Among the 26 volunteers that completed the trial, there were no responses to CS6 in the absence of LT. In the groups receiving both CS6 and LT, 68 and 53% were found to have serum anti-CS6 immunoglobulin G (IgG) and IgA, respectively; 37 and 42% had IgG and IgA anti-CS6 ASCs. All of the volunteers receiving LT had anti-LT IgG, and 90% had serum anti-LT IgA; 79 and 37% had anti-LT IgG and IgA ASCs. Delayed-type hypersensitivity (DTH), suggesting T-cell responses, was seen in 14 of 19 volunteers receiving LT and CS6; no DTH was seen in subjects receiving CS6 alone. This study demonstrated that protein antigens delivered by a simple patch could induce significant systemic immune responses but only in the presence of an adjuvant such as LT. The data suggest that an ETEC vaccine for travelers delivered by a patch may be a viable approach worthy of further evaluation.
TheBabesia bovisMerozoite Surface Antigen 2 Locus Contains Four Tandemly Arranged and Expressed Genes Encoding Immunologically Distinct Proteins
Authors: Florin-Christensen Monica , Suarez Carlos E. , Hines Stephen A. , Palmer Guy H. , Brown Wendy C. , McElwain Terry F. 
Published: July 2002

ABSTRACTMembers of the variable merozoite surface antigen (vmsa) gene family ofBabesia bovisencode membrane proteins involved in erythrocyte invasion. In this study, we have identified and sequenced the complete 8.3-kb genomic locus containingmsa-2, a member of thevmsafamily, in the biologically cloned Mexico Mo7 strain. Four tandemly arranged copies ofmsa-2-related genes were found in the locus. The four genes, designatedmsa-2a1(which corresponds to the originally describedmsa-2gene),msa-2a2,msa-2b, andmsa-2c, were shown to be transcribed and expressed and encode proteins with open reading frames ranging in size from 266 (MSA-2c) to 317 (MSA-2a1) amino acids. MSA-2a1and -2a2are the most closely related of the four proteins (90% identity), differing by (i) the number of 24-amino-acid repeats that comprise a surface-exposed B-cell epitope and (ii) the presence of a 32-amino-acid area of recombination between MSA-2a2and -2b. In contrast,msa-2cis most closely related to the previously describedbabr 0.8gene in Australia strains ofB. bovis. Comparison of MSA-2 proteins in the Argentina R1A strain ofB. boviswith the Mexico Mo7 clone revealed a relatively high degree of conservation (83.6, 69.4, 79.1, and 88.7% amino acid identity for MSA-2a1, -2a2, -2b, and -2c, respectively), in contrast to the extensive MSA-1 sequence variation (52% identity) between the same two strains. Postinfection bovine immune serum contains antibodies that bound to each of the recombinant MSA-2 proteins. Blocking assays demonstrated the presence of unique B-cell epitopes in MSA-2a1, -2b, and -2c. The results support the evolution of themsa-2locus through at least two gene duplications, with selection for multiple related but antigenically distinct merozoite surface proteins.
Inhibition of Leukocyte Rolling by Nitric Oxide during Sepsis Leads to Reduced Migration of Active Microbicidal Neutrophils
Authors: Benjamim Claudia Farias , Silva João Santana , Fortes Zuleica Bruno , Oliveira Maria Aparecida , Ferreira Sérgio Henrique , Cunha Fernando Queiroz 
Published: July 2002

ABSTRACTWe developed two models of sepsis with different degrees of severity, sublethal and lethal sepsis, induced by cecal ligation and puncture. Lethal sepsis induced by cecal ligation and puncture (L-CLP) resulted in failure of neutrophil migration to the infection site and high mortality. Treatment of septic animals with aminoguanidine (AG), a nitric oxide (NO) synthase inhibitor, precluded the failure of neutrophil migration and protected the animals from death. However, cytokine-induced NO synthase (iNOS)-deficient (iNOS−/−) mice subjected to L-CLP did not present neutrophil migration failure, but 100% lethality occurred. iNOS−/−mice subjected to sublethal sepsis induced by cecal ligation and puncture (SL-CLP) also suffered high mortality despite the occurrence of neutrophil migration. This apparent paradox could be explained by the lack of microbicidal activity in neutrophils of iNOS−/−mice present at the infection site due to their inability to produce NO. Notably, SL- and L-CLP iNOS−/−mice showed high bacterial numbers in exudates. The inhibition of neutrophil migration by NO is due to inhibition of a neutrophil/endothelium adhesion mechanism, since a reduction in leukocyte rolling, adhesion, and emigration was observed in L-CLP wild-type mice. These responses were prevented by AG treatment and were not observed in the iNOS−/−L-CLP group. There was no significant change in L-selectin expression in neutrophils from L-CLP mice. Thus, it seems that the decrease in leukocyte rolling is due to a defect in the expression of adhesion molecules on endothelial surfaces mediated by iNOS-derived NO. In conclusion, the results indicate that despite the importance of NO in neutrophil microbicidal activity, its generation in severe sepsis reduces neutrophil migration by inhibiting leukocyte rolling and their firm adhesion to the endothelium, in effect impairing the migration of leukocytes and consequently their fundamental role in host cell defense mechanisms.
Candida albicans
Expresses a Focal Adhesion Kinase-Like Protein That Undergoes Increased Tyrosine Phosphorylation upon Yeast Cell Adhesion to Vitronectin and the EA.hy 926 Human Endothelial Cell Line
Authors: Santoni Giorgio , Lucciarini Roberta , Amantini Consuelo , Jacobelli Jordan , Spreghini Elisabetta , Ballarini Patrizia , Piccoli Mario , Gismondi Angela 
Published: July 2002

ABSTRACT

The signaling pathways triggered by adherence of
Candida albicans
to the host cells or extracellular matrix are poorly understood. We provide here evidence in
C. albicans
yeasts of a p105 focal adhesion kinase (Fak)-like protein (that we termed CaFak), antigenically related to the vertebrate p125Fak, and its involvement in integrin-like-mediated fungus adhesion to vitronectin (VN) and EA.hy 926 human endothelial cell line. Biochemical analysis with different anti-chicken Fak antibodies identified CaFak as a 105-kDa protein and immunofluorescence and cytofluorimetric analysis on permeabilized cells specifically stain
C. albicans
yeasts; moreover, confocal microscopy evidences CaFak as a cytosolic protein that colocalizes on the membrane with the integrin-like VN receptors upon yeast adhesion to VN. The protein tyrosine kinase (PTK) inhibitors genistein and herbimycin A strongly inhibited
C. albicans
yeast adhesion to VN and EA.hy 926 endothelial cells. Moreover, engagement of αvβ3 and αvβ5 integrin-like on
C. albicans
either by specific monoclonal antibodies or upon adhesion to VN or EA.hy 926 endothelial cells stimulates CaFak tyrosine phosphorylation that is blocked by PTK inhibitor. A role for CaFak in
C. albicans
yeast adhesion was also supported by the failure of VN to stimulate its tyrosine phosphorylation in a
C. albicans
mutant showing normal levels of CaFak and VNR-like integrins but displaying reduced adhesiveness to VN and EA.hy 926 endothelial cells. Our results suggest that
C. albicans
Fak-like protein is involved in the control of yeast cell adhesion to VN and endothelial cells.
Enzymology of Type IV Macromolecule Secretion Systems: the Conjugative Transfer Regions of Plasmids RP4 and R388 and the
cag
Pathogenicity Island of
Helicobacter pylori
Encode Structurally and Functionally Related Nucleoside Triphosphate Hydrolases
Authors: Krause Sabine , Pansegrau Werner , Lurz Rudi , de la Cruz Fernando , Lanka Erich 
Published: July 2002

ABSTRACT

Type IV secretion systems direct transport of protein or nucleoprotein complexes across the cell envelopes of prokaryotic donor and eukaryotic or prokaryotic recipient cells. The process is mediated by a membrane-spanning multiprotein assembly. Potential NTPases belonging to the VirB11 family are an essential part of the membrane-spanning complex. Three representatives of these NTPases originating from the conjugative transfer regions of plasmids RP4 (TrbB) and R388 (TrwD) and from the
cag
pathogenicity island of
Helicobacter pylori
(HP0525) were overproduced and purified in native form. The proteins display NTPase activity with distinct substrate specificities in vitro. TrbB shows its highest specific hydrolase activity with dATP, and the preferred substrate for HP0525 is ATP. Analysis of defined TrbB mutations altered in motifs conserved within the VirB11 protein family shows that there is a correlation between the loss or reduction of NTPase activity and transfer frequency. Tryptophan fluorescence spectroscopy of TrbB and HP0525 suggests that both interact with phospholipid membranes, changing their conformation. NTPase activity of both proteins was stimulated by the addition of certain phospholipids. According to our results, Virb11-like proteins seem to most likely be involved in the assembly of the membrane-spanning multiprotein complex.
Branched-Chain α-Keto Acid Catabolism via the Gene Products of the
bkd
Operon in
Enterococcus faecalis
: a New, Secreted Metabolite Serving as a Temporary Redox Sink
Authors: Ward Donald E. , van der Weijden Coen C. , van der Merwe Marthinus J. , Westerhoff Hans V. , Claiborne Al , Snoep Jacky L. 
Published: July 2002

ABSTRACT

Recently the
bkd
gene cluster from
Enterococcus faecalis
was sequenced, and it was shown that the gene products constitute a pathway for the catabolism of branched-chain α-keto acids. We have now investigated the regulation and physiological role of this pathway. Primer extension analysis identified the presence of a single promoter upstream of the
bkd
gene cluster. Furthermore, a putative catabolite-responsive element was identified in the promoter region, indicative of catabolite repression. Consistent with this was the observation that expression of the
bkd
gene cluster is repressed in the presence of glucose, fructose, and lactose. It is proposed that the conversion of the branched-chain α-keto acids to the corresponding free acids results in the formation of ATP via substrate level phosphorylation. The utilization of the α-keto acids resulted in a marked increase of biomass, equivalent to a net production of 0.5 mol of ATP per mol of α-keto acid metabolized. The pathway was active under aerobic as well as anaerobic conditions. However, under anaerobic conditions the presence of a suitable electron acceptor to regenerate NAD
+
from the NADH produced by the branched-chain α-keto acid dehydrogenase complex was required for complete conversion of α-ketoisocaproate. Interestingly, during the conversion of the branched-chain α-keto acids an intermediate was always detected extracellularly. With α-ketoisocaproic acid as the substrate this intermediate was tentatively identified as 1,1-dihydroxy-4-methyl-2-pentanone. This reduced form of α-ketoisocaproic acid was found to serve as a temporary redox sink.
Regulation of Cation-Coupled High-Affinity Phosphate Uptake in the Yeast
Saccharomyces cerevisiae
Authors: Pattison-Granberg Johanna , Persson Bengt L. 
Published: July 2002

ABSTRACT

Studies of the high-affinity phosphate transporters in the yeast
Saccharomyces cerevisiae
using mutant strains lacking either the Pho84 or the Pho89 permease revealed that the transporters are differentially regulated. Although both genes are induced by phosphate starvation, activation of the Pho89 transporter precedes that of the Pho84 transporter early in the growth phase in a way which may possibly reflect a fine tuning of the phosphate uptake process relative to the availability of external phosphate.
ZipA-Induced Bundling of FtsZ Polymers Mediated by an Interaction between C-Terminal Domains
Authors: Hale Cynthia A. , Rhee Amy C. , de Boer Piet A. J. 
Published: July 2002

ABSTRACTFtsZ and ZipA are essential components of the septal ring apparatus, which mediates cell division inEscherichia coli. FtsZ is a cytoplasmic tubulin-like GTPase that forms protofilament-like homopolymers in vitro. In the cell, the protein assembles into a ring structure at the prospective division site early in the division cycle, and this marks the first recognized event in the assembly of the septal ring. ZipA is an inner membrane protein which is recruited to the nascent septal ring at a very early stage through a direct interaction with FtsZ. Using affinity blotting and protein localization techniques, we have determined which domain on each protein is both sufficient and required for the interaction between the two proteins in vitro as well as in vivo. The results show that ZipA binds to residues confined to the 20 C-terminal amino acids of FtsZ. The FtsZ binding (FZB) domain of ZipA is significantly larger and encompasses the C-terminal 143 residues of ZipA. Significantly, we find that the FZB domain of ZipA is also required and sufficient to induce dramatic bundling of FtsZ protofilaments in vitro. Consistent with the notion that the ability to bind and bundle FtsZ polymers is essential to the function of ZipA, we find that ZipA derivatives lacking an intact FZB domain fail to support cell division in cells depleted for the native protein. Interestingly, ZipA derivatives which do contain an intact FZB domain but which lack the N-terminal membrane anchor or in which this anchor is replaced with the heterologous anchor of the DjlA protein also fail to rescue ZipA−cells. Thus, in addition to the C-terminal FZB domain, the N-terminal domain of ZipA is required for ZipA function. Furthermore, the essential properties of the N domain may be more specific than merely acting as a membrane anchor.
A Region of the Transmembrane Regulatory Protein ToxR That Tethers the Transcriptional Activation Domain to the Cytoplasmic Membrane Displays Wide Divergence among
Vibrio
Species
Authors: Osorio Carlos R. , Klose Karl E. 
Published: July 2002

ABSTRACT

The virulence regulatory protein ToxR of
Vibrio cholerae
is unique in that it contains a cytoplasmic DNA-binding–transcriptional activation domain, a transmembrane domain, and a periplasmic domain. Although ToxR and other transmembrane transcriptional activators have been discovered in other bacteria, little is known about their mechanism of activation. Utilizing degenerate oligonucleotides and PCR, we have amplified internal
toxR
gene sequences from seven
Vibrio
and
Photobacterium
species and subspecies, demonstrating that
toxR
is an ancestral gene of the family
Vibrionaceae
. Sequence alignment of all available ToxR amino acid sequences revealed a region between the transcriptional activation and transmembrane domains that displays wide divergence among
Vibrio
species. We hypothesize that this region merely tethers the transcriptional activation domain to the cytoplasmic membrane and thus can tolerate wide divergence and multiple insertions and deletions. The divergence in the tether region at the nucleotide level may provide a useful tool for the distinction of
Vibrio
and
Photobacterium
species.
Leafy Gall Formation Is Controlled by
fasR
, an AraC-Type Regulatory Gene in
Rhodococcus fascians
Authors: Temmerman Wim , Vereecke Danny , Dreesen Rozemarijn , Van Montagu Marc , Holsters Marcelle , Goethals Koen 
Published: July 2002

ABSTRACT

Rhodococcus fascians
can interact with many plant species and induce the formation of either leafy galls or fasciations. To provoke symptoms,
R. fascians
strain D188 requires pathogenicity genes that are located on a linear plasmid, pFiD188. The
fas
genes are essential for virulence and constitute an operon that encodes, among other functions, a cytokinin synthase gene. Expression of the
fas
genes is induced by extracts of infected plant tissue only. We have isolated an AraC-type regulatory gene,
fasR
, located on pFiD188, which is indispensable for pathogenesis and for
fas
gene expression. The combined results of our experiments show that in vitro expression of the
fas
genes in a defined medium is strictly regulated and that several environmental factors (pH, carbon and nitrogen sources, phosphate and oxygen content, and cell density) and regulatory proteins are involved. We further show that expression of the
fas
genes is controlled at both the transcriptional and the translational levels. The complex expression pattern probably reflects the necessity of integrating a multitude of signals and underlines the importance of the
fas
operon in the pathogenicity of
R. fascians
.
Sequence of the Genome of
Salmonella
Bacteriophage P22
Authors: Byl Carolyn Vander , Kropinski Andrew M. 
Published: July 2002

ABSTRACT

The sequence of the nonredundant region of the
Salmonella enterica
serovar Typhimurium temperate, serotype-converting bacteriophage P22 has been completed. The genome is 41,724 bp with an overall moles percent GC content of 47.1%. Numerous examples of potential integration host factor and C1-binding sites were identified in the sequence. In addition, five potential rho-independent terminators were discovered. Sixty-five genes were identified and annotated. While many of these had been described previously, we have added several new ones, including the genes involved in serotype conversion and late control. Two of the serotype conversion gene products show considerable sequence relatedness to GtrA and -B from
Shigella
phages SfII, SfV, and SfX. We have cloned the serotype-converting cassette (
gtrABC
) and demonstrated that it results in
Salmonella
serovar Typhimurium LT2 cells which express antigen O1. Many of the putative proteins show sequence relatedness to proteins from a great variety of other phages, supporting the hypothesis that this phage has evolved through the recombinational exchange of genetic information with other viruses.
Surface Signaling in Ferric Citrate Transport Gene Induction: Interaction of the FecA, FecR, and FecI Regulatory Proteins
Authors: Enz Sabine , Mahren Susanne , Stroeher Uwe H. , Braun Volkmar 
Published: July 2002

ABSTRACTInEscherichia coli, transcription of the ferric citrate transport genesfecABCDEis controlled by a novel signal transduction mechanism that starts at the cell surface. Binding of ferric citrate to the outer membrane protein FecA initiates a signal that is transmitted by FecR across the cytoplasmic membrane into the cytoplasm where FecI, the sigma factor, is activated. Interaction between the signaling proteins was demonstrated by utilizing two methods. In in vitro binding assays, FecR that was His tagged at the N terminus [(His)10-FecR] and bound to a Ni-nitrilotriacetic acid agarose column was able to retain FecA, and FecR that was His tagged at the C terminus [FecR-(His)6] retained FecI on the column. An N-terminally truncated, induction-negative but transport-active FecA protein did not bind to (His)10-FecR. The in vivo assay involved the determination of the FecA, FecR, and FecI interacting domains with the bacterial two-hybrid Lex-based system. FecA1–79interacts with FecR101–317and FecR1–85interacts with FecI1–173. These data clearly support a model that proposes interaction of the periplasmic N terminus of FecA with the periplasmic C-terminal portion of FecR and interaction of the cytoplasmic N terminus of FecR with FecI, which results in FecI activation.
Bacteriophage T4 Self-Assembly: Localization of gp3 and Its Role in Determining Tail Length
Authors: Vianelli A. , Wang G. R. , Gingery M. , Duda R. L. , Eiserling F. A. , Goldberg E. B. 
Published: July 2002

ABSTRACT
Gene 3 of bacteriophage T4 participates at a late stage in the T4 tail assembly pathway, but the hypothetical protein product, gp3, has never been identified in extracts of infected cells or in any tail assembly intermediate. In order to overcome this difficulty, we expressed gp3 in a high-efficiency plasmid expression vector and subsequently purified it for further analysis. The N-terminal sequence of the purified protein showed that the initial methionine had been removed. Variant C-terminal amino acid sequences were resolved by determining the cysteine content of the protein. The molecular mass of 20.6 kDa for the pure protein was confirmed by Western blotting, using a specific anti-gp3 serum for which the purified protein was the immunogen. We also demonstrated, for the first time, the physical presence of gp3 in the mature T4 phage particle and localized it to the tail tube. By finding a nonleaky, nonpermissive host for a gene 3 mutant, we could clearly demonstrate a new phenotype: the slow, aberrant elongation of the tail tube in the absence of gp3.