bluuebunny/crossref_metadata_2025_split huggingface dataset:
Yersinia pseudotuberculosis
Produces a Cytotoxic Necrotizing Factor
Authors: Lockman Hank A. , Gillespie Rebecca A. , Baker Beth D. , Shakhnovich Elizabeth 
Published: July 2002

ABSTRACT

Cell extracts from
Yersinia pseudotuberculosis
induced multinucleation in HEp-2 cells in a manner similar to the effect caused by
Escherichia coli
cytotoxic necrotizing factor (CNF). The activity was not dependent on the
Yersinia
70-kb virulence plasmid, and the activity was not inhibited by antibodies capable of neutralizing
E. coli
CNF type 1. The nucleotide sequence of the
Yersinia cnf
gene was 65.1% identical to the
E. coli cnf
gene.
Regulation of Antigen-Specific Immunoglobulin G Subclasses in Response to Conserved and PolymorphicPlasmodium falciparumAntigens in an In Vitro Model
Authors: Garraud Olivier , Perraut Ronald , Diouf Ababacar , Nambei Wilfrid S. , Tall Adama , Spiegel André , Longacre Shirley , Kaslow David C. , Jouin Hélène , Mattei Denise , Engler Gina M. , Nutman Thomas B. , Riley Eleanor M. , Mercereau-Puijalon Odile 
Published: July 2002

ABSTRACTCytophilic antibodies (Abs) play a critical role in protection againstPlasmodium falciparumblood stages, yet little is known about the parameters regulating production of these Abs. We used an in vitro culture system to study the subclass distribution of antigen (Ag)-specific immunoglobulin G (IgG) produced by peripheral blood mononuclear cells (PBMCs) from individuals exposed toP. falciparumor unexposed individuals. PBMCs, cultivated with or without cytokines and exogenous CD40/CD40L signals, were stimulated with a crude parasite extract, recombinant vaccine candidates derived from conserved Ags (19-kDa C terminus of merozoite surface protein 1 [MSP119], R23, and PfEB200), or recombinant Ags derived from the polymorphic Ags MSP1 block 2 and MSP2. NoP. falciparum-specific Ab production was detected in PBMCs from unexposed individuals. PBMCs from donors exposed frequently toP. falciparuminfections produced multiple IgG subclasses when they were stimulated with the parasite extract but usually only one IgG subclass when they were stimulated with a recombinant Ag. Optimal Ab production required addition of interleukin-2 (IL-2) and IL-10 for all antigenic preparations. The IgG subclass distribution was both donor and Ag dependent and was only minimally influenced by the exogenous cytokine environment. In vitro IgG production and subclass distribution correlated with plasma Abs to some Ags (MSP119, R23, and MSP2) but not others (PfEB200 and the three MSP1 block 2-derived Ags). Data presented here suggest that intrinsic properties of the protein Ag itself play a major role in determining the subclass of the Ab response, which has important implications for rational design of vaccine delivery.
Expression and Characterization of StreptococcalrgpGenes Required for Rhamnan Synthesis inEscherichia coli
Authors: Shibata Yukie , Yamashita Yoshihisa , Ozaki Kazuhisa , Nakano Yoshio , Koga Toshihiko 
Published: July 2002

ABSTRACTSix genes (rgpAthroughrgpF) that were involved in assembling the rhamnose-glucose polysaccharide (RGP) inStreptococcus mutanswere previously identified (Y. Yamashita, Y. Tsukioka, K. Tomihisa, Y. Nakano, and T. Koga, J. Bacteriol.180:5803-5807, 1998). The group-specific antigens of Lancefield group A, C, and E streptococci and the polysaccharide antigen ofStreptococcus sobrinushave the same rhamnan backbone as the RGP ofS. mutans. Escherichia coliharboring plasmid pRGP1 containing all sixrgpgenes did not synthesize complete RGP. However,E. colicarrying a plasmid with all of thergpgenes except forrgpEsynthesized the rhamnan backbone of RGP without glucose side chains, suggesting that in addition torgpE, another gene is required for glucose side-chain formation. Synthesis of the rhamnan backbone inE. colirequired the initiation of transfer ofN-acetylglucosamine to a lipid carrier and the expression of thergpCandrgpDgenes encoding the putative ABC transporter specific for RGP. The similarities in RGP synthesis betweenE. coliandS. mutanssuggest common pathways for rhamnan synthesis. Therefore, we evaluated the rhamnosyl polymerization process inE. coliby high-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the lipooligosaccharide (LOS). AnE. colitransformant harboringrgpAproduced the LOS modified by the addition of a single rhamnose residue. Furthermore, thergpA,rgpB, andrgpFgenes of pRGP1 were independently mutated by an internal deletion, and the LOS chemotypes of their transformants were examined. The transformant with anrgpAdeletion showed the same LOS profile asE. coliwithout a plasmid. The transformant with anrgpBdeletion showed the same LOS profile asE. coliharboringrgpAalone. The transformant with anrgpFdeletion showed the LOS band with the most retarded migration. On the basis of these results, we speculated that RgpA, RgpB, and RgpF, in that order, function in rhamnan polymerization.
Oral Vaccination with Subunit Vaccines Protects Animals against Aerosol Infection withMycobacterium tuberculosis
Authors: Doherty T. Mark , Olsen Anja Weinrich , van Pinxteren Laurens , Andersen Peter 
Published: July 2002

ABSTRACTImmunity againstMycobacterium tuberculosisdepends largely on activation of cell-mediated responses, and gamma interferon has been shown to play a crucial role in this process in both humans and animal models. Since the lung is normally the organ in which infection is initiated and is the major site of pathology, immune responses in the lung play a significant role in restricting initial infection withM. tuberculosis. The aim of the present study was to stimulate efficient immunity in the lung by targeting the gut mucosa. Detoxified monophosphoryl lipid A (MPL) has been shown to be a relatively nontoxic adjuvant which efficiently promotes the induction of type 1 responses when it is given by the traditional subcutaneous route. We have therefore compared subcutaneous immunization of mice to oral immunization by using a model subunit vaccine carrying two immunodominant proteins fromM. tuberculosis, in combination with MPL-based adjuvants. While less effective when used to prime a response, a heterologous priming and boosting vaccination strategy employing oral boosting induced significant systemic type 1 responses which equaled and surpassed those attained by subcutaneous immunization protocols. Moreover, the increased immune responses observed correlated with the induction of substantial protection against subsequent aerosol infection with virulentM. tuberculosisat levels comparable to, or better than, those obtained by multiple subcutaneous vaccinations. These results demonstrate that booster vaccinations via mucosal surfaces, by combining efficient subunit vaccines with the potent adjuvant MPL, may be an effective method of addressing some of the shortcomings of current vaccination strategies.
DNA Immunization in a Mouse Model of Latent Tuberculosis: Effect of DNA Vaccination on Reactivation of Disease and on Reinfection with a Secondary Challenge
Authors: Repique Charlene J. , Li Amy , Collins Frank M. , Morris Sheldon L. 
Published: July 2002

ABSTRACT

Individuals who are latently infected with
Mycobacterium tuberculosis
can develop active disease via either endogenous reactivation of the latent bacilli or exogenous reinfection with a second mycobacterial strain. In this study, we investigated whether immunization with a tuberculosis DNA vaccine cocktail that induces significant protective responses in mice could prevent reactivation of disease in a murine latent-tuberculosis model. In addition, we assessed whether DNA vaccination could retard the growth of a secondary aerogenic infection with
M. tuberculosis
(exogenous reinfection) in latently infected mice. In the reactivation studies, administration of the DNA vaccine combination did not prevent recrudescence of the latent infection after injection of dexamethasone. Moreover, for the reinfection experiments, only a modest decrease in the growth of a secondary
M. tuberculosis
challenge in DNA-vaccinated animals, compared to controls, was observed 14 and 28 days after the reinfection of previously exposed mice. Interestingly, although proliferation of the secondary challenge was reduced significantly in a nonvaccinated chronic-infection group relative to the naïve controls, the number of bacilli still increased by 500-fold 1 month after the secondary challenge in mice with active tuberculosis. These results indicate that novel immunotherapeutic approaches will be required to prevent reactivation of infection or reinfection of individuals with latent tuberculosis.
Multiple-Locus Variable-Number Tandem Repeat Analysis Reveals Genetic Relationships within
Bacillus anthracis
Authors: Keim P. , Price L. B. , Klevytska A. M. , Smith K. L. , Schupp J. M. , Okinaka R. , Jackson P. J. , Hugh-Jones M. E. 
Published: July 2002

ABSTRACT

Bacillus anthracis
is one of the most genetically homogeneous pathogens described, making strain discrimination particularly difficult. In this paper, we present a novel molecular typing system based on rapidly evolving variable-number tandem repeat (VNTR) loci. Multiple-locus VNTR analysis (MLVA) uses the combined power of multiple alleles at several marker loci. In our system, fluorescently labeled PCR primers are used to produce PCR amplification products from eight VNTR regions in the
B. anthracis
genome. These are detected and their sizes are determined using an ABI377 automated DNA sequencer. Five of these eight loci were discovered by sequence characterization of molecular markers (
vrrC
1
,
vrrC
2
,
vrrB
1
,
vrrB
2
, and CG3), two were discovered by searching complete plasmid nucleotide sequences (pXO1-aat and pXO2-at), and one was known previously (
vrrA
). MLVA characterization of 426
B. anthracis
isolates identified 89 distinct genotypes. VNTR markers frequently identified multiple alleles (from two to nine), with Nei's diversity values between 0.3 and 0.8. Unweighted pair-group method arithmetic average cluster analysis identified six genetically distinct groups that appear to be derived from clones. Some of these clones show worldwide distribution, while others are restricted to particular geographic regions. Human commerce doubtlessly has contributed to the dispersal of particular clones in ancient and modern times.
Hyperrecombination in
Streptococcus pneumoniae
Depends on an Atypical
mutY
Homologue
Authors: Samrakandi Moulay Mustapha , Pasta Franck 
Published: July 2002

ABSTRACT

The unusual behavior of the mutation
ami36
, which generates hyperrecombination in two point crosses, was previously attributed to a localized conversion process changing A/G mispairs into CG pairs. Although the mechanism was found to be dependent on the DNA polymerase I, the specific function responsible for this correction was still unknown. Analysis of the pneumococcal genome sequence has revealed the presence of an open reading frame homologous to the gene
mutY
of
Escherichia coli
. The gene
mutY
encodes an adenine glycosylase active on A/G and A/7,8-dihydro-8-oxoguanine (8-OxoG) mismatches, inducing their repair to CG and C/8-OxoG, respectively. Here we report that disrupting the pneumococcal
mutY
homologue abolishes the hyperrecombination induced by
ami36
and leads to a mutator phenotype specifically enhancing AT-to-CG transversions. The deduced amino acid sequence of the pneumococcal MutY protein reveals the absence of four cysteines, highly conserved in the endonuclease III/MutY glycosylase family, which ligate a [4Fe-4S]
2+
cluster. The actual function of this cluster is still intriguing, inasmuch as we show that the pneumococcal gene complements a
mutY
strain of
E. coli.
The Hydrogenase Cytochrome
b
Heme Ligands of
Azotobacter vinelandii
Are Required for Full H
2
Oxidation Capability
Authors: Meek Laura , Arp Daniel J. 
Published: July 2002

ABSTRACT

The hydrogenase in
Azotobacter vinelandii
, like other membrane-bound [NiFe] hydrogenases, consists of a catalytic heterodimer and an integral membrane cytochrome
b
. The histidines ligating the hemes in this cytochrome
b
were identified by H
2
oxidation properties of altered proteins produced by site-directed mutagenesis. Four fully conserved and four partially conserved histidines in HoxZ were substituted with alanine or tyrosine. The roles of these histidines in HoxZ heme binding and hydrogenase were characterized by O
2
-dependent H
2
oxidation and H
2
-dependent methylene blue reduction in vivo. Mutants H33A/Y (H33 replaced by A or Y), H74A/Y, H194A, H208A/Y, and H194,208A lost O
2
-dependent H
2
oxidation activity, H194Y and H136A had partial activity, and H97Y,H98A and H191A had full activity. These results suggest that the fully conserved histidines 33, 74, 194, and 208 are ligands to the hemes, tyrosine can serve as an alternate ligand in position 194, and H136 plays a role in H
2
oxidation. In mutant H194A/Y, imidazole (Imd) rescued H
2
oxidation activity in intact cells, which suggests that Imd acts as an exogenous ligand. The heterodimer activity, quantitatively determined as H
2
-dependent methylene blue reduction, indicated that the heterodimers of all mutants were catalytically active. H33A/Y had wild-type levels of methylene blue reduction, but the other HoxZ ligand mutants had significantly less than wild-type levels. Imd reconstituted full methylene blue reduction activity in mutants H194A/Y and H208A/Y and partial activity in H194,208A. These results indicate that structural and functional integrity of HoxZ is required for physiologically relevant H
2
oxidation, and structural integrity of HoxZ is necessary for full heterodimer-catalyzed H
2
oxidation.
First Evidence for Existence of an Uphill Electron Transfer through the
bc
1
and NADH-Q Oxidoreductase Complexes of the Acidophilic Obligate Chemolithotrophic Ferrous Ion-Oxidizing Bacterium
Thiobacillus ferrooxidans
Authors: Elbehti A. , Brasseur G. , Lemesle-Meunier D. 
Published: July 2002

ABSTRACT

The energy-dependent electron transfer pathway involved in the reduction of pyridine nucleotides which is required for CO
2
fixation to occur in the acidophilic chemolithotrophic organism
Thiobacillus ferrooxidans
was investigated using ferrocytochrome
c
as the electron donor. The experimental results show that this uphill pathway involves a
bc
1
and an NADH-Q oxidoreductase complex functioning in reverse, using an electrochemical proton gradient generated by ATP hydrolysis. Based on these results, a model is presented to explain the balance of the reducing equivalent from ferrocytochrome
c
between the exergonic and endergonic electron transfer pathways.
Saccharomyces cerevisiae
Σ1278b Has Novel Genes of the
N
-Acetyltransferase Gene Superfamily Required for
l
-Proline Analogue Resistance
Authors: Takagi Hiroshi , Shichiri Mika , Takemura Miho , Mohri Miho , Nakamori Shigeru 
Published: July 2002

ABSTRACT

We discovered on the chromosome of
Saccharomyces cerevisiae
Σ1278b novel genes involved in
l
-proline analogue
l
-azetidine-2-carboxylic acid resistance which are not present in the standard laboratory strains. The 5.4 kb-DNA fragment was cloned from the genomic library of the
l
-azetidine-2-carboxylic acid-resistant mutant derived from a cross between
S. cerevisiae
strains S288C and Σ1278b. The nucleotide sequence of a 4.5-kb segment exhibited no identity with the sequence in the genome project involving strain S288C. Deletion analysis indicated that one open reading frame encoding a predicted protein of 229 amino acids is indispensable for
l
-azetidine-2-carboxylic acid resistance. The protein sequence was found to be a member of the
N
-acetyltransferase superfamily. Genomic Southern analysis and gene disruption showed that two copies of the novel gene with one amino acid change at position 85 required for
l
-azetidine-2-carboxylic acid resistance were present on chromosomes X and XIV of Σ1278b background strains. When this novel
MPR1
or
MPR2
gene (sigma 1278b gene for
l
-proline analogue resistance) was introduced into the other
S. cerevisiae
strains, all of the recombinants were resistant to
l
-azetidine-2-carboxylic acid, indicating that both
MPR1
and
MPR2
are expressed and have a global function in
S. cerevisiae
.
ς
BldN
, an Extracytoplasmic Function RNA Polymerase Sigma Factor Required for Aerial Mycelium Formation in
Streptomyces coelicolor
A3(2)
Authors: Bibb Maureen J. , Molle Virginie , Buttner Mark J. 
Published: July 2002

ABSTRACT

Sporulation mutants of
Streptomyces coelicolor
appear white because they are defective in the synthesis of the gray polyketide spore pigment, and such white (
whi
) mutants have been used to define 13 sporulation loci.
whiN
, one of five new
whi
loci identified in a recent screen of NTG (
N
-methyl-
N
′-nitro-
N
-nitrosoguanidine)-induced
whi
strains (N. J. Ryding et al., J. Bacteriol. 181:5419–5425, 1999), was defined by two mutants, R112 and R650. R650 produced frequent spores that were longer than those of the wild type. In contrast, R112 produced long, straight, undifferentiated hyphae, although rare spore chains were observed, sometimes showing highly irregular septum placement. Subcloning and sequencing showed that
whiN
encodes a member of the extracytoplasmic function subfamily of RNA polymerase sigma factors and that the sigma factor has an unusual N-terminal extension of approximately 86 residues that is not present in other sigma factors. A constructed
whiN
null mutant failed to form aerial mycelium (the “bald” phenotype) and, as a consequence,
whiN
was renamed
bldN
. This observation was not totally unexpected because, on some media, the R112 point mutant produced substantially less aerial mycelium than its parent, M145. The
bldN
null mutant did not fit simply into the extracellular signaling cascade proposed for
S. coelicolor bld
mutants. Expression of
bldN
was analyzed during colony development in wild-type and aerial mycelium-deficient
bld
strains.
bldN
was transcribed from a single promoter,
bldNp. bldN
transcription was developmentally regulated, commencing approximately at the time of aerial mycelium formation, and depended on
bldG
and
bldH
, but not on
bldA
,
bldB
,
bldC
,
bldF
,
bldK
, or
bldJ
or on
bldN
itself. Transcription from the
p1
promoter of the response-regulator gene
bldM
depended on
bldN
in vivo, and the
bldMp1
promoter was shown to be a direct biochemical target for ς
BldN
holoenzyme in vitro.
Recovery of Hydrogen Peroxide-Sensitive Culturable Cells of
Vibrio vulnificus
Gives the Appearance of Resuscitation from a Viable but Nonculturable State
Authors: Bogosian Gregg , Aardema Noelle D. , Bourneuf Edward V. , Morris Patricia J. L. , O'Neil Julia P. 
Published: July 2002

ABSTRACT

The viabilities of five strains of
Vibrio vulnificus
were evaluated during the storage of the organisms in sterile seawater at 5°C. The number of CFU was measured by plate count methods on rich media. The total cell numbers were determined by direct microscopic count methods. The titer of CFU declined logarithmically to undetectable levels over a period of 2 to 3 weeks, while the total cell numbers were unchanged. Midway through each study, higher culturable cell counts began to be observed on plates containing catalase or sodium pyruvate; during the latter stages of the study, the plate counts on such media were up to 1,000-fold higher than those on unsupplemented plates. Because autoclaving is known to generate hydrogen peroxide in rich media, and because catalase and sodium pyruvate are known to eliminate hydrogen peroxide, it appears that the conditions of the experiments led to the selection of a hydrogen peroxide-sensitive culturable cell subpopulation. At the time of the final stage of the decline in viability of each culture, hydrogen peroxide-sensitive cells were the only culturable cells present. Warming samples of the cultures to room temperature led to the growth of these residual culturable cells, utilizing nutrients provided by the nonculturable cells. The cells that grew recovered hydrogen peroxide resistance. When mixtures of culturable and nonculturable cells were diluted to the point where only nonculturable cells were present, or when the hydrogen peroxide-sensitive culturable cells had declined to undetectable levels, warming had no effect; no culturable cells were recovered. Warming has been reported to “resuscitate” nonculturable cells. Recognition of the existence of hydrogen peroxide-sensitive culturable cell populations, as well as their ability to grow to high levels in the warmed seawater microcosms, leads instead to the conclusion that while warming permits culturable cells to grow, it has no effect on nonculturable cells.
Revised Translation Start Site for
secM
Defines an Atypical Signal Peptide That Regulates
Escherichia coli secA
Expression
Authors: Sarker Shameema , Rudd Kenneth E. , Oliver Donald 
Published: July 2002

ABSTRACT

The secretion-responsive regulation of
Escherichia coli secA
occurs by coupling its translation to the translation and secretion of an upstream regulator,
secM
(formerly geneX). We revise the translational start site for
secM
, defining a new signal peptide sequence with an extended amino-terminal region. Mutational studies indicate that certain atypical amino acyl residues within this extended region are critical for proper
secA
regulation.
Characterization of Specific Nucleotide Substitutions in DtxR-Specific Operators of
Corynebacterium diphtheriae
That Dramatically Affect DtxR Binding, Operator Function, and Promoter Strength
Authors: Lee John H. , Holmes Randall K. 
Published: July 2002

ABSTRACT

The diphtheria toxin repressor (DtxR) of
Corynebacterium diphtheriae
uses Fe
2+
as a corepressor. Holo-DtxR inhibits transcription from the iron-regulated promoters (IRPs) designated IRP1 through IRP5 as well as from the promoters for the
tox
and
hmuO
genes. DtxR binds to 19-bp operators with the consensus sequence 5′-TTAGGTTAGCCTAACCTAA-3′, a perfect 9-bp palindrome interrupted by a single C · G base pair. Among the seven known DtxR-specific operators, IRP3 exhibits the weakest binding to DtxR. The message (sense) strand of the IRP3 operator (5′-TTAGGT
G
AG
A
C
GC
ACC
C
A
T
-3′ [nonconsensus nucleotides underlined]) overlaps by 2 nucleotides at its 5′ end with the putative −10 sequence of the IRP3 promoter. The underlined C at position +7 from the center of the IRP3 operator [C(+7)] is unique, because T is conserved at that position in other DtxR-specific operators. The present study examined the effects of nucleotide substitutions at position +7 or −7 in the IRP3 operator. In gel mobility shift assays, only the change of C(+7) to the consensus nucleotide T caused a dramatic increase in the binding of DtxR, whereas other nucleotide substitutions for C(+7) or replacements for A(−7) had only small positive or negative effects on DtxR binding. All substitutions for C(+7) or A(−7) except for A(−7)C dramatically decreased IRP3 promoter strength. In contrast, the A(−7)C variant caused increased promoter strength at the cost of nearly eliminating repressibility by DtxR. The message (sense) strand of the IRP1 operator (5′-TTAGGTTAGCCAAACCTTT-3′) includes the −35 region of the IRP3 promoter. A T(+7)C variant of the IRP1 operator was also constructed, and it was shown to exhibit decreased binding to DtxR, decreased repressibility by DtxR, and increased promoter strength. The nucleotides at positions +7 and −7 in DtxR-specific operators are therefore important determinants of DtxR binding and repressibility of transcription by DtxR, and they also have significant effects on promoter activity for IRP3 and IRP1.