bluuebunny/crossref_metadata_2025_split huggingface dataset:
tmRNA in
Caulobacter crescentus
Is Cell Cycle Regulated by Temporally Controlled Transcription and RNA Degradation
Authors: Keiler Kenneth C. , Shapiro Lucy 
Published: March 2003

ABSTRACT

SsrA, or tmRNA, is a small RNA found in all bacteria that intervenes in selected translation reactions to target the nascent polypeptide for rapid proteolysis. We have found that the abundance of SsrA RNA in
Caulobacter crescentus
is regulated with respect to the cell cycle. SsrA RNA abundance increases in late G
1
phase, peaks during the G
1
-S transition, and declines in early S phase, in keeping with the reported role for SsrA in the timing of DNA replication initiation. Cell cycle regulation of SsrA RNA is accomplished by a combination of temporally controlled transcription and regulated RNA degradation. Transcription from the
ssrA
promoter peaks late in G
1
, just before the peak in SsrA RNA abundance. SsrA RNA is stable in G
1
-phase cells and late S-phase cells but is degraded with a half-life of 4 to 5 min at the onset of S phase. This degradation is surprising, since SsrA RNA is both highly structured and highly abundant. This is the first observation of a structural RNA that is cell cycle regulated.
Novel Genomic Rearrangement That Affects Expression of theStreptococcus pyogenesStreptolysin O (slo) Gene
Authors: Savic Dragutin J. , Ferretti Joseph J. 
Published: March 2003

ABSTRACTA RecA-independent chromosomal rearrangement in the upstream region of the streptolysin O (slo) gene ofStreptococcus pyogeneswhich affectssloexpression was identified. PCR analysis was used to demonstrate that this kind of rearrangement was found in several strains of different lineages. Chromosomal loci involved in the recombination were found to be 746 kb apart on the 1.85-Mb-long chromosome. The primary structure of the splicing region, the reproducibility of the rearrangement, and the fact that reconstructed recombinant molecules fused toermandlacZreporter genes affected their expression indicate that this event is not accidental but may play a role in the expression of theslogene. In addition, the product of the recombining DNAs, including the splicing site, does not follow any example of a known recombination mechanism. The implications of this rearrangement forsloexpression are discussed.
Leucine-Responsive Regulatory Protein-Mediated Repression of
clp
(Encoding CS31A) Expression by
l
-Leucine and
l
-Alanine in
Escherichia coli
Authors: Crost Cécile , Garrivier Annie , Harel Josée , Martin Christine 
Published: March 2003

ABSTRACT

CS31A produced by septicemic and diarrheic
Escherichia coli
belongs to the Pap-regulatory family of adhesive factors, which are under methylation-dependent transcriptional regulation. Common features of operons encoding members of this family include two conserved GATC sites in the upstream regulatory region, and transcriptional regulators homologue to the PapB and PapI proteins. Methylation protection of GATC sites was previously shown to be dependent on the leucine-responsive regulatory protein (Lrp). Lrp and ClpB, the PapB equivalent, repressed
clp
basal transcription. A PapI homologue (AfaF) was required together with Lrp to establish the phase variation control, which gave rise to phase-ON cells that expressed CS31A and phase-OFF cells that did not express CS31A. In phase-OFF cells, the GATC
dist
site was methylated and the GATC
prox
site was protected from methylation, whereas in phase-ON cells, the inverse situation was found. Unlike Pap fimbriae, CS31A synthesis was dramatically reduced in media containing
l
-alanine or
l
-leucine.
l
-Alanine prevented the OFF-to-ON switch, locking
clp
expression in the OFF phase, whereas
l
-leucine repressed transcription without obvious effect on the switch frequency of phase variation. In phase-variable cells, leucine and alanine promoted methylation of GATC
dist
and methylation protection of GATC
prox
, increasing the methylation pattern characteristic of repressed cells. Furthermore, alanine prevented the AfaF-dependent methylation protection of GATC
dist
and thus the appearance of phase-ON cells. In addition, analysis of
clp
expression in a Lrp-negative background indicated that alanine and leucine also repressed
clp
transcription by a methylation-independent mechanism.
Operon Structure and Regulation of the
nos
Gene Region of
Pseudomonas stutzeri
, Encoding an ABC-Type ATPase for Maturation of Nitrous Oxide Reductase
Authors: Honisch Ulrike , Zumft Walter G. 
Published: March 2003

ABSTRACT

The synthesis of a functional nitrous oxide reductase requires an assembly apparatus for the insertion of the prosthetic copper. Part of the system is encoded by maturation genes located in
Pseudomonas stutzeri
immediately downstream of the structural gene for the enzyme. We have studied the transcriptional organization and regulation of this region and found a
nosDFYL tatE
operon structure. In addition to a putative ABC transporter, consisting of NosD, NosF, and NosY, the operon encodes a Cu chaperone, NosL, and a component of the Tat translocon, TatE. The
nosD
operon was activated in response to anaerobiosis and nitrate denitrification. The membrane-bound regulator NosR was required for operon expression; in addition, DnrD, a regulator of the Crp-Fnr family, enhanced expression under anaerobic conditions. This establishes a likely signal transduction sequence of NO → DnrD →
nosR
/NosR →
nosD
operon. DnrD-dependent expression was also observed for the
nnrS
operon (located immediately downstream of the
nosD
operon), which encodes a putative heme-Cu protein (NnrS) and a member of the short-chain dehydrogenase family (ORF247). The NosF protein, encoded within the
nosD
operon, exhibits sequence similarity to ABC-type ATPases. It was fused to the
Escherichia coli
maltose-binding protein and overexpressed in soluble form. The fusion protein was purified and shown to have ATPase activity. NosF is the first maturation factor for which a catalytic function has been demonstrated in vitro.
Identification of the Immunodominant Protein and Other Proteins of theBacillus anthracisExosporium
Authors: Steichen Christopher , Chen Ping , Kearney John F. , Turnbough, Charles L. 
Published: March 2003

ABSTRACTSpores ofBacillus anthracis, the causative agent of anthrax, are enclosed by a prominent loose-fitting, balloon-like layer called the exosporium. Although the exosporium serves as the source of surface antigens and a primary permeability barrier of the spore, its molecular structure and function are not well characterized. In this study, we identified five major proteins in purifiedB. anthracis(Sterne strain) exosporia. One protein was the recently identified collagen-like glycoprotein BclA, which appears to be a structural component of the exosporium hair-like nap. Using a large panel of unique antispore monoclonal antibodies, we demonstrated that BclA is the immunodominant antigen on theB. anthracisspore surface. We also showed that the BclA protein and not a carbohydrate constituent directs the dominant immune response. In addition, the length of the central (GXX)nrepeat region of BclA appears to be strain specific. Two other unique proteins, BxpA and BxpB, were identified. BxpA is unusually rich in Gln and Pro residues and contains several different tandem repeats, which also exhibit strain-specific variation. In addition, BxpA was found to be cleaved approximately in half. BxpB appears to be glycosylated or associated with glycosylated material and is encoded by a gene that (along withbclA) may be part of an exosporium genomic island. The other two proteins identified were alanine racemase and superoxide dismutase, both of which were reported to be associated with the surface of otherBacillusspores. Possible functions of the newly identified proteins are discussed.
Additional Targets of theBacillus subtilisGlobal Regulator CodY Identified by Chromatin Immunoprecipitation and Genome-Wide Transcript Analysis
Authors: Molle Virginie , Nakaura Yoshiko , Shivers Robert P. , Yamaguchi Hirotake , Losick Richard , Fujita Yasutaro , Sonenshein Abraham L. 
Published: March 2003

ABSTRACTAdditional targets of CodY, a GTP-activated repressor of early stationary-phase genes inBacillus subtilis, were identified by combining chromatin immunoprecipitation, DNA microarray hybridization, and gel mobility shift assays. The direct targets of CodY newly identified by this approach included regulatory genes for sporulation, genes that are likely to encode transporters for amino acids and sugars, and the genes for biosynthesis of branched-chain amino acids.
Design of Antisense RNA Constructs for Downregulation of the Acetone Formation Pathway of
Clostridium acetobutylicum
Authors: Tummala Seshu B. , Welker Neil E. , Papoutsakis Eleftherios T. 
Published: March 2003

ABSTRACT

We investigated the effect of antisense RNA (asRNA) structural properties on the downregulation efficacy of enzymes in the acetone-formation pathway (acetoacetate decarboxylase [AADC] and coenzyme A-transferase [CoAT]) of
Clostridium acetobutylicum
strain ATCC 824. First, we generated three strains,
C. acetobutylicum
ATCC 824 (pADC38AS), 824(pADC68AS), and 824(pADC100AS), which contain plasmids that produce asRNAs of various lengths against the AADC (
adc
) transcript. Western analysis showed that all three strains exhibit low levels of AADC compared to the plasmid control [ATCC 824(pSOS95del)]. By using computational algorithms, the three different asRNAs directed toward AADC, along with previously reported clostridial asRNAs, were examined for structural features (free nucleotides and components). When the normalized metrics of these structural features were plotted against percent downregulation, only the component/nucleotide ratio correlated well with in vivo asRNA effectiveness. Despite the significant downregulation of AADC in these strains, there were no concomitant effects on acetone formation. These findings suggest that AADC does not limit acetone formation and, thus, we targeted next the CoAT. Using the component/nucleotide ratio as a selection parameter, we developed three strains [ATCC 824 (pCTFA2AS), 824(pCTFB1AS), and 824(pCOAT11AS)] which express asRNAs to downregulate either or both of the CoAT subunits. Compared to the plasmid control strain, these strains produced substantially low levels of acetone and butanol and Western blot analyses showed significantly low levels of both CoAT subunits. These results show that CoAT is the rate-limiting enzyme in acetone formation and strengthen the hypothesis that the component/nucleotide ratio is a predictive indicator of asRNA effectiveness.
Mutational Analysis of the Residue at Position 48 in the
Salmonella enterica
Serovar Typhimurium PhoQ Sensor Kinase
Authors: Sanowar Sarah , Martel Alexandre , Moual Hervé Le 
Published: March 2003

ABSTRACT

The PhoP/PhoQ two-component regulatory system of
Salmonella enterica
serovar Typhimurium plays an essential role in controlling virulence by mediating the adaptation to Mg
2+
depletion. The
pho
-
24
allele of
phoQ
harbors a single amino acid substitution (T48I) in the periplasmic domain of the PhoQ histidine kinase sensor. This mutation has been shown to increase net phosphorylation of the PhoP response regulator. We analyzed the effect on signaling by PhoP/PhoQ of various amino acid substitutions at this position (PhoQ-T48X [X = A, S, V, I, or L]). Mutations T48V, T48I, and T48L were found to affect signaling by PhoP/PhoQ both in vivo and in vitro. Mutations PhoQ-T48V and PhoQ-T48I increased both the expression of the
mgtA
::
lacZ
transcriptional fusion and the net phosphorylation of PhoP, conferring to cells a PhoP constitutively active phenotype. In contrast, mutation PhoQ-T48L barely responded to changes in the concentration of external Mg
2+
, in vivo and in vitro, conferring to cells a PhoP constitutively inactive phenotype. By analyzing in vitro the individual catalytic activities of the PhoQ-T48X sensors, we found that the PhoP constitutively active phenotype observed for the PhoQ-T48V and PhoQ-T48I proteins is solely due to decreased phosphatase activity. In contrast, the PhoP constitutively inactive phenotype observed for the PhoQ-T48L mutant resulted from both decreased autokinase activity and increased phosphatase activity. Our data are consistent with a model in which the residue at position 48 of PhoQ contributes to a conformational switch between kinase- and phosphatase-dominant states.
High Levels of Intracellular Cysteine Promote Oxidative DNA Damage by Driving the Fenton Reaction
Authors: Park Sunny , Imlay James A. 
Published: March 2003

ABSTRACT

Escherichia coli
is generally resistant to H
2
O
2
, with >75% of cells surviving a 3-min challenge with 2.5 mM H
2
O
2
. However, when cells were cultured with poor sulfur sources and then exposed to cystine, they transiently exhibited a greatly increased susceptibility to H
2
O
2
, with <1% surviving the challenge. Cell death was due to an unusually rapid rate of DNA damage, as indicated by their filamentation, a high rate of mutation among the survivors, and DNA lesions by a direct assay. Cell-permeable iron chelators eliminated sensitivity, indicating that intracellular free iron mediated the conversion of H
2
O
2
into a hydroxyl radical, the direct effector of DNA damage. The cystine treatment caused a temporary loss of cysteine homeostasis, with intracellular pools increasing about eightfold. In vitro analysis demonstrated that cysteine reduces ferric iron with exceptional speed. This action permits free iron to redox cycle rapidly in the presence of H
2
O
2
, thereby augmenting the rate at which hydroxyl radicals are formed. During routine growth, cells maintain small cysteine pools, and cysteine is not a major contributor to DNA damage. Thus, the homeostatic control of cysteine levels is important in conferring resistance to oxidants. More generally, this study provides a new example of a situation in which the vulnerability of cells to oxidative DNA damage is strongly affected by their physiological state.
Molecular Analysis of the Multiple GroEL Proteins of Chlamydiae
Authors: Karunakaran Karuna P. , Noguchi Yasuyuki , Read Timothy D. , Cherkasov Artem , Kwee Jeffrey , Shen Caixia , Nelson Colleen C. , Brunham Robert C. 
Published: March 2003

ABSTRACT

Genome sequencing revealed that all six chlamydiae genomes contain three
groEL
-like genes (
groEL1
,
groEL2
, and
groEL3
). Phylogenetic analysis of
groEL1
,
groEL2
, and
groEL3
indicates that these genes are likely to have been present in chlamydiae since the beginning of the lineage. Comparison of deduced amino acid sequences of the three
groEL
genes with those of other organisms showed high homology only for
groEL1
, although comparison of critical amino acid residues that are required for polypeptide binding of the
Escherichia coli
chaperonin GroEL revealed substantial conservation in all three chlamydial GroELs. This was further supported by three-dimensional structural predictions. All three genes are expressed constitutively throughout the developmental cycle of
Chlamydia trachomatis
, although
groEL1
is expressed at much higher levels than are
groEL2
and
groEL3
. Transcription of
groEL1
, but not
groEL2
and
groEL3
, was elevated when HeLa cells infected with
C. trachomatis
were subjected to heat shock. Western blot analysis with polyclonal antibodies raised against recombinant GroEL1, GroEL2, and GroEL3 demonstrated the presence of the three proteins in
C. trachomatis
elementary bodies, with GroEL1 being present in the largest amount. Only
C. trachomatis groEL1
and
groES
together complemented a temperature-sensitive
E. coli groEL
mutant. Complementation did not occur with
groEL2
or
groEL3
alone or together with
groES
. The role for each of the three GroELs in the chlamydial developmental cycle and in disease pathogenesis requires further study.
Mass Distribution and Spatial Organization of the Linear Bacterial Motor ofSpiroplasma citriR8A2
Authors: Trachtenberg Shlomo , Andrews S. Brian , Leapman Richard D. 
Published: March 2003

ABSTRACTIn the simple, helical, wall-less bacterial genusSpiroplasma, chemotaxis and motility are effected by a linear, contractile motor arranged as a flat cytoskeletal ribbon attached to the inner side of the membrane along the shortest helical line. With scanning transmission electron microscopy and diffraction analysis, we determined the hierarchical and spatial organization of the cytoskeleton ofSpiroplasma citriR8A2. The structural unit appears to be a fibril, ∼5 nm wide, composed of dimers of a 59-kDa protein; each ribbon is assembled from seven fibril pairs. The functional unit of the intact ribbon is a pair of aligned fibrils, along which pairs of dimers form tetrameric ring-like repeats. On average, isolated and purified ribbons contain 14 fibrils or seven well-aligned fibril pairs, which are the same structures observed in the intact cell. Scanning transmission electron microscopy mass analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified cytoskeletons indicate that the 59-kDa protein is the only constituent of the ribbons.
Identification and Biosynthesis of Cyclic Enterobacterial Common Antigen in
Escherichia coli
Authors: Erbel Paul J. A. , Barr Kathleen , Gao Ninguo , Gerwig Gerrit J. , Rick Paul D. , Gardner Kevin H. 
Published: March 2003

ABSTRACT

Phosphoglyceride-linked enterobacterial common antigen (ECA
PG
) is a cell surface glycolipid that is synthesized by all gram-negative enteric bacteria. The carbohydrate portion of ECA
PG
consists of linear heteropolysaccharide chains comprised of the trisaccharide repeat unit Fuc4NAc-ManNAcA-GlcNAc, where Fuc4NAc is 4-acetamido-4,6-dideoxy-
d
-galactose, ManNAcA is
N
-acetyl-
d
-mannosaminuronic acid, and GlcNAc is
N
-acetyl-
d
-glucosamine. The potential reducing terminal GlcNAc residue of each polysaccharide chain is linked via phosphodiester linkage to a phosphoglyceride aglycone. We demonstrate here the occurrence of a water-soluble cyclic form of enterobacterial common antigen, ECA
CYC
, purified from
Escherichia coli
strains B and K-12 with solution nuclear magnetic resonance (NMR) spectroscopy, electrospray ionization mass spectrometry (ESI-MS), and additional biochemical methods. The ECA
CYC
molecules lacked an aglycone and contained four trisaccharide repeat units that were nonstoichiometrically substituted with up to four O-acetyl groups. ECA
CYC
was not detected in mutant strains that possessed null mutations in the
wecA
,
wecF
, and
wecG
genes of the
wec
gene cluster. These observations corroborate the structural data obtained by NMR and ESI-MS analyses and show for the first time that the trisaccharide repeat units of ECA
CYC
and ECA
PG
are assembled by a common biosynthetic pathway.
Gene Expression Profile of
Campylobacter jejuni
in Response to Growth Temperature Variation
Authors: Stintzi Alain 
Published: March 2003

ABSTRACT

The foodborne pathogen
Campylobacter jejuni
is the primary causative agent of gastroenteritis in humans. In the present study a whole genome microarray of
C. jejuni
was constructed and validated. These DNA microarrays were used to measure changes in transcription levels over time, as
C. jejuni
cells responded to a temperature increase from 37 to 42°C. Approximately 20% of the
C. jejuni
genes were significantly up- or downregulated over a 50-min period after the temperature increase. The global change in
C. jejuni
transcriptome was found to be essentially transient, with only a small subset of genes still differentially expressed after 50 min. A substantial number of genes with a downregulated coexpression pattern were found to encode for ribosomal proteins. This suggests a short growth arrest upon temperature stress, allowing the bacteria to reshuffle their energy toward survival and adaptation to the new growth temperature. Genes encoding chaperones, chaperonins, and heat shock proteins displayed the most dramatic and rapid upregulation immediately after the temperature change. Interestingly, genes encoding proteins involved in membrane structure modification were differentially expressed, either up- or downregulated, suggesting a different protein membrane makeup at the two different growth temperatures. Overall, these data provide new insights into the primary response of
C. jejuni
to surmount a sudden temperature upshift, allowing the bacterium to survive and adapt its transcriptome to a new steady state.
Definition of the
Escherichia coli
MC4100 Genome by Use of a DNA Array
Authors: Peters Joseph E. , Thate Timothy E. , Craig Nancy L. 
Published: March 2003

ABSTRACT

We have used an
Escherichia coli
K-12 whole-genome array based on the DNA sequence of strain MG1655 as a tool to identify deletions in another
E. coli
K-12 strain, MC4100, by probing the array with labeled chromosomal DNA. Despite the continued widespread use of MC4100 as an experimental system, the specific genetic relationship of this strain to the sequenced K-12 derivative MG1655 has not been resolved. MC4100 was found to contain four deletions, ranging from 1 to 97 kb in size. The exact nature of three of the deletions was previously unresolved, and the fourth deletion was altogether unknown.
Plasmid R16 ArdA Protein Preferentially Targets Restriction Activity of the Type I Restriction-Modification System
Eco
KI
Authors: Thomas Angela T. , Brammar William J. , Wilkins Brian M. 
Published: March 2003

ABSTRACT

The ArdA antirestriction protein of the IncB plasmid R16 selectively inhibited the restriction activity of
Eco
KI, leaving significant levels of modification activity under conditions in which restriction was almost completely prevented. The results are consistent with the hypothesis that ArdA functions in bacterial conjugation to allow an unmodified plasmid to evade restriction in the recipient bacterium and yet acquire cognate modification.
Identification of an Unknown Promoter,
OUTIIp
, within the IS
10
R Element
Authors: Martínez-García Esteban , Navarro-Lloréns Juana María , Tormo Antonio 
Published: March 2003

ABSTRACT

A novel promoter in IS
10
R (
OUTIIp
) has been found in one of its ends in an inverted position relative to promoter pOUT.
OUTIIp
shows characteristics similar to those of
rpoS
-dependent promoters such as a gearbox expression pattern. It is under catabolite repression and positively regulated by ppGpp or conditioned media. This opens new challenges in IS
10
R transposition.