bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Characterization of the
eis
Promoter of
Mycobacterium tuberculosis
Authors: Roberts Esteban A. , Clark Amanda , McBeth Sarah , Friedman Richard L. 
Published: August 2004

ABSTRACT

To further understand
Mycobacterium tuberculosis
pathogenesis, the regulation of potential virulence genes needs to be investigated. The
eis
gene of
M. tuberculosis
H37Rv enhances the intracellular survival of
Mycobacterium smegmatis
, which does not contain
eis
, within macrophages (J. Wei, J. L. Dahl, J. W. Moulder, E. A. Roberts, P. O'Gaora, D. B. Young, and R. L. Friedman, J. Bacteriol.
182:
377-384, 2000). Experiments were done to characterize the
eis
promoter in
M. smegmatis
and
M. tuberculosis
H37Ra. The putative −10 and −35 regions matched the
Escherichia coli
σ
70
consensus 67 and 83%, respectively, making it a group A/
SigA
-like mycobacterial promoter. Expression of site-directed variants of the core promoter region, determined by flow cytometry using
gfp
as a reporter, showed that the putative −10 region is essential for
eis
expression. In addition, site-directed alteration of the
eis
promoter to the consensus
E. coli
σ
70
promoter elements increased
gfp
transcription to levels similar to that driven by the heat shock promoter, p
hsp60
, of
Mycobacterium bovis
BCG. Upstream promoter deletion analysis showed that a 200- and 412-bp region of the promoter was necessary for maximum expression of
gfp
in
M. smegmatis
and
M. tuberculosis
H37Ra, respectively. Random mutagenesis of the 412-bp
eis
promoter, using a catechol 2,3-dioxygenase screen and activity assay, defined nucleotides upstream of the core promoter region that are essential to
eis
expression in both
M. smegmatis
and
M. tuberculosis
H37Ra, including a region homologous to a DinR
cis
element.
Differential Regulation of Soluble and Membrane-Bound Inorganic Pyrophosphatases in the Photosynthetic Bacterium
Rhodospirillum rubrum
Provides Insights into Pyrophosphate-Based Stress Bioenergetics
Authors: López-Marqués Rosa L. , Pérez-Castiñeira José R. , Losada Manuel , Serrano Aurelio 
Published: August 2004

ABSTRACT

Soluble and membrane-bound inorganic pyrophosphatases (sPPase and H
+
-PPase, respectively) of the purple nonsulfur bacterium
Rhodospirillum rubrum
are differentially regulated by environmental growth conditions. Both proteins and their transcripts were found in cells of anaerobic phototrophic batch cultures along all growth phases, although they displayed different time patterns. However, in aerobic cells that grow in the dark, which exhibited the highest growth rates, Northern and Western blot analyses as well as activity assays demonstrated high sPPase levels but no H
+
-PPase. It is noteworthy that H
+
-PPase is highly expressed in aerobic cells under acute salt stress (1 M NaCl). H
+
-PPase was also present in anaerobic cells growing at reduced rates in the dark under either fermentative or anaerobic respiratory conditions. Since H
+
-PPase was detected not only under all anaerobic growth conditions but also under salt stress in aerobiosis, the corresponding gene is not invariably repressed by oxygen. Primer extension analyses showed that, under all anaerobic conditions tested, the
R. rubrum
H
+
-PPase gene utilizes two activator-dependent tandem promoters, one with an FNR-like sequence motif and the other with a RegA motif, whereas in aerobiosis under salt stress, the H
+
-PPase gene is transcribed from two further tandem promoters involving other transcription factors. These results demonstrate a tight transcriptional regulation of the H
+
-PPase gene, which appears to be induced in response to a variety of environmental conditions, all of which constrain cell energetics.
Requirements for Nitric Oxide Generation from Isoniazid Activation In Vitro and Inhibition of Mycobacterial Respiration In Vivo
Authors: Timmins Graham S. , Master Sharon , Rusnak Frank , Deretic Vojo 
Published: August 2004

ABSTRACTIsoniazid (INH), a front-line antituberculosis agent, is activated by mycobacterial catalase-peroxidase KatG, converting INH into bactericidal reactive species. Here we investigated the requirements and the pathway of nitric oxide (NO˙) generation during oxidative activation of INH byMycobacterium tuberculosisKatG in vitro. We also provide in vivo evidence that INH-derived NO˙ can inhibit key mycobacterial respiratory enzymes, which may contribute to the overall antimycobacterial action of INH.
Analysis of the Genome Structure of the Nonpathogenic Probiotic
Escherichia coli
Strain Nissle 1917
Authors: Grozdanov Lubomir , Raasch Carsten , Schulze Jürgen , Sonnenborn Ulrich , Gottschalk Gerhard , Hacker Jörg , Dobrindt Ulrich 
Published: August 2004

ABSTRACT

Nonpathogenic
Escherichia coli
strain Nissle 1917 (O6:K5:H1) is used as a probiotic agent in medicine, mainly for the treatment of various gastroenterological diseases. To gain insight on the genetic level into its properties of colonization and commensalism, this strain's genome structure has been analyzed by three approaches: (i) sequence context screening of tRNA genes as a potential indication of chromosomal integration of horizontally acquired DNA, (ii) sequence analysis of 280 kb of genomic islands (GEIs) coding for important fitness factors, and (iii) comparison of Nissle 1917 genome content with that of other
E. coli
strains by DNA-DNA hybridization. PCR-based screening of 324 nonpathogenic and pathogenic
E. coli
isolates of different origins revealed that some chromosomal regions are frequently detectable in nonpathogenic
E. coli
and also among extraintestinal and intestinal pathogenic strains. Many known fitness factor determinants of strain Nissle 1917 are localized on four GEIs which have been partially sequenced and analyzed. Comparison of these data with the available knowledge of the genome structure of
E. coli
K-12 strain MG1655 and of uropathogenic
E. coli
O6 strains CFT073 and 536 revealed structural similarities on the genomic level, especially between the
E. coli
O6 strains. The lack of defined virulence factors (i.e., alpha-hemolysin, P-fimbrial adhesins, and the semirough lipopolysaccharide phenotype) combined with the expression of fitness factors such as microcins, different iron uptake systems, adhesins, and proteases, which may support its survival and successful colonization of the human gut, most likely contributes to the probiotic character of
E. coli
strain Nissle 1917.
DNA Microarray-Based Genome Comparison of a Pathogenic and a Nonpathogenic Strain of
Xylella fastidiosa
Delineates Genes Important for Bacterial Virulence
Authors: Koide Tie , Zaini Paulo A. , Moreira Leandro M. , Vêncio Ricardo Z. N. , Matsukuma Adriana Y. , Durham Alan M. , Teixeira Diva C. , El-Dorry Hamza , Monteiro Patrícia B. , da Silva Ana Claudia R. , Verjovski-Almeida Sergio , da Silva Aline M. , Gomes Suely L. 
Published: August 2004

ABSTRACT

Xylella fastidiosa
is a phytopathogenic bacterium that causes serious diseases in a wide range of economically important crops. Despite extensive comparative analyses of genome sequences of
Xylella
pathogenic strains from different plant hosts, nonpathogenic strains have not been studied. In this report, we show that
X. fastidiosa
strain J1a12, associated with citrus variegated chlorosis (CVC), is nonpathogenic when injected into citrus and tobacco plants. Furthermore, a DNA microarray-based comparison of J1a12 with 9a5c, a CVC strain that is highly pathogenic and had its genome completely sequenced, revealed that 14 coding sequences of strain 9a5c are absent or highly divergent in strain J1a12. Among them, we found an arginase and a fimbrial adhesin precursor of type III pilus, which were confirmed to be absent in the nonpathogenic strain by PCR and DNA sequencing. The absence of arginase can be correlated to the inability of J1a12 to multiply in host plants. This enzyme has been recently shown to act as a bacterial survival mechanism by down-regulating host nitric oxide production. The lack of the adhesin precursor gene is in accordance with the less aggregated phenotype observed for J1a12 cells growing in vitro. Thus, the absence of both genes can be associated with the failure of the J1a12 strain to establish and spread in citrus and tobacco plants. These results provide the first detailed comparison between a nonpathogenic strain and a pathogenic strain of
X. fastidiosa
, constituting an important step towards understanding the molecular basis of the disease.
spoIVH(ykvV), a Requisite Cortex Formation Gene, Is Expressed in Both Sporulating Compartments ofBacillus subtilis
Authors: Imamura Daisuke , Kobayashi Kazuo , Sekiguchi Junichi , Ogasawara Naotake , Takeuchi Michio , Sato Tsutomu 
Published: August 2004

ABSTRACTIt is well known that theykvU-ykvVoperon is under the regulation of the σE-associated RNA polymerase (EσE). In our study, we observed thatykvVis transcribed together with the upstreamykvUgene by EσEin the mother cell and monocistronically under EσGcontrol in the forespore. Interestingly, alternatively expressedykvVin either the forespore or the mother cell increased the sporulation efficiency in theykvVbackground. Studies show that the YkvV protein is a member of the thioredoxin superfamily and also contains a putative Sec-type secretion signal at the N terminus. We observed efficient sporulation in a mutant strain obtained by replacing the putative signal peptide of YkvV with the secretion signal sequence of SleB, indicating that the putative signal sequence is essential for spore formation. These results suggest that YkvV is capable of being transported by the putative Sec-type signal sequence into the space between the double membranes surrounding the forespore. The ability ofykvVexpression in either compartment to complement is indeed intriguing and further introduces a new dimension to the genetics ofB. subtilisspore formation. Furthermore, electron microscopic observation revealed a defective cortex in theykvVdisruptant. In addition, the expression levels of σK-directed genes significantly decreased despite normal σGactivity in theykvVmutant. However, immunoblotting with the anti-σKantibody showed that pro-σKwas normally processed in theykvVmutant, indicating that YkvV plays an important role in cortex formation, consistent with recent reports. We therefore propose thatykvVshould be renamedspoIVH.
The LuxR Homolog ExpR, in Combination with the Sin Quorum Sensing System, Plays a Central Role in
Sinorhizobium meliloti
Gene Expression
Authors: Hoang Hanh H. , Becker Anke , González Juan E. 
Published: August 2004

ABSTRACT

Quorum sensing, a population density-dependent mechanism for bacterial communication and gene regulation, plays a crucial role in the symbiosis between alfalfa and its symbiont
Sinorhizobium meliloti
. The Sin system, one of three quorum sensing systems present in
S. meliloti
, controls the production of the symbiotically active exopolysaccharide EPS II. Based on DNA microarray data, the Sin system also seems to regulate a multitude of
S. meliloti
genes, including genes that participate in low-molecular-weight succinoglycan production, motility, and chemotaxis, as well as other cellular processes. Most of the regulation by the Sin system is dependent on the presence of the ExpR regulator, a LuxR homolog. Gene expression profiling data indicate that ExpR participates in additional cellular processes that include nitrogen fixation, metabolism, and metal transport. Based on our microarray analysis we propose a model for the regulation of gene expression by the Sin/ExpR quorum sensing system and another possible quorum sensing system(s) in
S. meliloti
.
Comparative Whole-Genome Analysis of Virulent and Avirulent Strains ofPorphyromonas gingivalis
Authors: Chen Tsute , Hosogi Yumiko , Nishikawa Kiyoshi , Abbey Kevin , Fleischmann Robert D. , Walling Jennifer , Duncan Margaret J. 
Published: August 2004

ABSTRACTWe usedPorphyromonas gingivalisgene microarrays to compare the total gene contents of the virulent strain W83 and the avirulent type strain, ATCC 33277. Signal ratios and scatter plots indicated that the chromosomes were very similar, with approximately 93% of the predicted genes in common, while at least 7% of them showed very low or no signals in ATCC 33277. Verification of the array results by PCR indicated that several of the disparate genes were either absent from or variant in ATCC 33277. Divergent features included already reported insertion sequences andragB, as well as additional hypothetical and functionally assigned genes. Several of the latter were organized in a putative operon in W83 and encoded enzymes involved in capsular polysaccharide synthesis. Another cluster was associated with two paralogous regions of the chromosome with a low G+C content, at 41%, compared to that of the whole genome, at 48%. These regions also contained conserved and species-specific hypothetical genes, transposons, insertion sequences, and integrases and were located adjacent to tRNA genes; thus, they had several characteristics of pathogenicity islands. While this global comparative analysis showed the close relationship between W83 and ATCC 33277, the clustering of genes that are present in W83 but divergent in or absent from ATCC 33277 is suggestive of chromosomal islands that may have been acquired by lateral gene transfer.
Tra Proteins Characteristic of F-Like Type IV Secretion Systems Constitute an Interaction Group by Yeast Two-Hybrid Analysis
Authors: Harris Robin L. , Silverman Philip M. 
Published: August 2004

ABSTRACTUsing yeast two-hybrid screens, we have defined an interaction group of six Tra proteins encoded by the F plasmid and required by F+cells to elaborate F pili. The six proteins are TraH, TraF, TraW, TraU, TrbI, and TrbB. Except for TrbI, these proteins were all identified as hallmarks of F-like type IV secretion systems (TFSSs), with no homologues among TFSS genes of P-type or I-type systems (T. Lawley, W. Klimke, M. Gubbins, and L. Frost, FEMS Microbiol. Lett. 224:1-15, 2003). Also with the exception of TrbI, which is an inner membrane protein, the remaining proteins are or are predicted to be periplasmic. TrbI consists of one membrane-spanning segment near its N terminus and an 88-residue, hydrophilic domain that extends into the periplasm. Hence, the proteins of this group probably form a periplasmic cluster inEscherichia coli. The interaction network identifies TraH as the most highly connected node, with two-hybrid links to TrbI, TraU, and TraF. As measured by transcriptional activation oflacZ, the TrbI-TraH interaction inSaccharomyces cerevisiaerequires the TraH amino acid segment from residues 193 to 225. The TraU and TraF interactions are localized to C-terminal segments of TraH (amino acids 315 to 458 for TraF and amino acids 341 to 458 for TraU). The TrbI-TraH interaction with full-length (less the signal peptide) TraH is weak but increases 40-fold with N-terminal TraH deletions; the first 50 amino acids appear to be critical for inhibiting TrbI binding in yeast. Previous studies by others have shown that, with the exception oftrbBmutations, which do not affect the elaboration or function of F pili under laboratory conditions, a mutation in any of the other genes in this interaction group alters the number or length distribution of F pili. We propose a model whereby one function of the TraH interaction group is to control F-pilus extension and retraction.
Identification of the Secretion and Translocation Domain of the Enteropathogenic and EnterohemorrhagicEscherichia coliEffector Cif, Using TEM-1 β-Lactamase as a New Fluorescence-Based Reporter
Authors: Charpentier Xavier , Oswald Eric 
Published: August 2004

ABSTRACTEnteropathogenic and enterohemorrhagicEscherichia coli(EPEC and EHEC) strains are human and animal pathogens that inject effector proteins into host cells via a type III secretion system (TTSS). Cif is an effector protein which induces host cell cycle arrest and reorganization of the actin cytoskeleton. Cif is encoded by a lambdoid prophage present in most of the EPEC and EHEC strains. In this study, we analyzed the domain that targets Cif to the TTSS by using a new reporter system based on a translational fusion of the effector proteins with mature TEM-1 β-lactamase. Translocation was detected directly in living host cells by using the fluorescent β-lactamase substrate CCF2/AM. We show that the first 16 amino acids (aa) of Cif were necessary and sufficient to mediate translocation into the host cells. Similarly, the first 20 aa of the effector proteins Map, EspF, and Tir, which are encoded in the same region as the TTSS, mediated secretion and translocation in a type III-dependent but chaperone-independent manner. A truncated form of Cif lacking its first 20 aa was no longer secreted and translocated, but fusion with the first 20 aa of Tir, Map, or EspF restored both secretion and translocation. In addition, the chimeric proteins were fully able to trigger host cell cycle arrest and stress fiber formation. In conclusion, our results demonstrate that Cif is composed of a C-terminal effector domain and an exchangeable N-terminal translocation signal and that the TEM-1 reporter system is a convenient tool for the study of the translocation of toxins or effector proteins into host cells.
Multiple-Locus Variable-Number Tandem Repeat Analysis of Dutch
Bordetella pertussis
Strains Reveals Rapid Genetic Changes with Clonal Expansion during the Late 1990s
Authors: Schouls Leo M. , van der Heide Han G. J. , Vauterin Luc , Vauterin Paul , Mooi Frits R. 
Published: August 2004

ABSTRACT

Bordetella pertussis
, the causative agent of whooping cough, has remained endemic in The Netherlands despite extensive nationwide vaccination since 1953. In the 1990s, several epidemic periods have resulted in many cases of pertussis. We have proposed that strain variation has played a major role in the upsurges of this disease in The Netherlands. Therefore, molecular characterization of strains is important in identifying the causes of pertussis epidemiology. For this reason, we have developed a multiple-locus variable-number tandem repeat analysis (MLVA) typing system for
B. pertussis
. By combining the MLVA profile with the allelic profile based on multiple-antigen sequence typing, we were able to further differentiate strains. The relationships between the various genotypes were visualized by constructing a minimum spanning tree. MLVA of Dutch strains of
B. pertussis
revealed that the genotypes of the strains isolated in the prevaccination period were diverse and clearly distinct from the strains isolated in the 1990s. Furthermore, there was a decrease in diversity in the strains from the late 1990s, with a remarkable clonal expansion that coincided with the epidemic periods. Using this genotyping, we have been able to show that
B. pertussis
is much more dynamic than expected.
Escherichia coli
Serogroup O107/O117 Lipopolysaccharide Binds and Neutralizes Shiga Toxin 2
Authors: Gamage Shantini D. , McGannon Colleen M. , Weiss Alison A. 
Published: August 2004

ABSTRACT

The AB
5
toxin Shiga toxin 2 (Stx2) has been implicated as a major virulence factor of
Escherichia coli
O157:H7 and other Shiga toxin-producing
E. coli
strains in the progression of intestinal disease to more severe systemic complications. Here, we demonstrate that supernatant from a normal
E. coli
isolate, FI-29, neutralizes the effect of Stx2, but not the related Stx1, on Vero cells. Biochemical characterization of the neutralizing activity identified the lipopolysaccharide (LPS) of FI-29, a serogroup O107/O117 strain, as the toxin-neutralizing component. LPSs from FI-29 as well as from type strains
E. coli
O107 and
E. coli
O117 were able bind Stx2 but not Stx1, indicating that the mechanism of toxin neutralization may involve inhibition of the interaction between Stx2 and the Gb
3
receptor on Vero cells.
Novel Archaeal Alanine:Glyoxylate Aminotransferase from
Thermococcus litoralis
Authors: Sakuraba Haruhiko , Kawakami Ryushi , Takahashi Hajime , Ohshima Toshihisa 
Published: August 2004

ABSTRACT

A novel alanine:glyoxylate aminotransferase was found in a hyperthermophilic archaeon,
Thermococcus litoralis
. The amino acid sequence of the enzyme did not show a similarity to any alanine:glyoxylate aminotransferases reported so far. Homologues of the enzyme appear to be present in almost all hyperthermophilic archaea whose whole genomes have been sequenced.
Amino Acid Substitutions in Putative Selectivity Filter Regions III and IV in KdpA Alter Ion Selectivity of the KdpFABC Complex from
Escherichia coli
Authors: Bertrand Jessica , Altendorf Karlheinz , Bramkamp Marc 
Published: August 2004

ABSTRACT

When grown under conditions of potassium limitation or high osmolality,
Escherichia coli
synthesizes the K
+
-translocating KdpFABC complex. The KdpA subunit, which has sequence homology to potassium channels of the KcsA type, has been shown to be important for potassium binding and transport. Replacement of the glycine residues in KdpA at positions 345 and 470, members of putative selectivity filter regions III and IV, alters the ion selectivity of the KdpFABC complex.
Complete Genomic Nucleotide Sequence of the Temperate Bacteriophage AaΦ23 of
Actinobacillus actinomycetemcomitans
Authors: Resch Grégory , Kulik Eva M. , Dietrich Fred S. , Meyer Jürg 
Published: August 2004

ABSTRACT

The entire double-stranded DNA genome of the
Actinobacillus actinomycetemcomitans
bacteriophage AaΦ23 was sequenced. Linear DNA contained in the phage particles is circularly permuted and terminally redundant. Therefore, the physical map of the phage genome is circular. Its size is 43,033 bp with an overall molar G+C content of 42.5 mol%. Sixty-six potential open reading frames (ORFs) were identified, including an ORF resulting from a translational frameshift. A putative function could be assigned to 23 of them. Twenty-three other ORFs share homologies only with hypothetical proteins present in several bacteria or bacteriophages, and 20 ORFs seem to be specific for phage AaΦ23. The organization of the phage genome and several genetic functions share extensive similarities to that of the lambdoid phages. However, AaΦ23 encodes a DNA adenine methylase, and the DNA packaging strategy is more closely related to the P22 system. The attachment sites of AaΦ23 (
attP
) and several
A. actinomycetemcomitans
hosts (
attB
) are 49 bp long.
Characterization of Lipoteichoic Acids asLactobacillus delbrueckiiPhage Receptor Components
Authors: Räisänen Liisa , Schubert Karin , Jaakonsaari Tiina , Alatossava Tapani 
Published: August 2004

ABSTRACTLipoteichoic acids (LTAs) were purified fromLactobacillus delbrueckiisubsp.lactisATCC 15808 and its LL-H adsorption-resistant mutant, Ads-5, by hydrophobic interaction chromatography.L. delbrueckiiphages (LL-H, the LL-H host range mutant, and JCL1032) were inactivated by these poly(glycerophosphate) type of LTAs in vitro in accordance to their adsorption to intact ATCC 15808 and Ads-5 cells.