bluuebunny/crossref_metadata_2025 huggingface dataset:
Turning behavior in Drosophila larvae: a role for the small scribbler transcript
Authors: Suster M. L. , Karunanithi S. , Atwood H. L. , Sokolowski M. B.
Published: September 2004

The Drosophila larva is extensively used for studies of neural development and function, yet the mechanisms underlying the appropriate development of its stereotypic motor behaviors remain largely unknown. We have previously shown that mutations in scribbler (sbb), a gene encoding two transcripts widely expressed in the nervous system, cause abnormally frequent episodes of turning in the third instar larva. Here we report that hypomorphic sbb mutant larvae display aberrant turning from the second instar stage onwards. We focus on the smaller of the two sbb transcripts and show that its pan‐neural expression during early larval life, but not in later larval life, restores wild type turning behavior. To identify the classes of neurons in which this sbb transcript is involved, we carried out transgenic rescue experiments. Targeted expression of the small sbb transcript using the cha‐GAL4 driver was sufficient to restore wild type turning behavior. In contrast, expression of this sbb transcript in motoneurons, sensory neurons or large numbers of unidentified interneurons was not sufficient. Our data suggest that the expression of the smaller sbb transcript may be needed in a subset of neurons for the maintenance of normal turning behavior in Drosophila larvae.
Reduced aggression in AMPA‐type glutamate receptor GluR‐A subunit‐deficient mice
Authors: Vekovischeva O. Y. , Aitta‐aho T. , Echenko O. , Kankaanpää A. , Seppälä T. , Honkanen A. , Sprengel R. , Korpi E. R.
Published: September 2004

The importance of AMPA‐type glutamate receptors has been demonstrated in neuronal plasticity and in adaptation to drugs of abuse. We studied the involvement of AMPA receptors in social interaction and anxiety and found that in several paradigms of agonistic behavior naïve male mice deficient for the GluR‐A subunit‐ containing AMPA receptors are less aggressive than wild‐type littermates. GluR‐A deficient mice and wild‐type littermates exhibited similar basic behavior and reflexes as monitored by observational Irwin's test, but they tended to be less anxious in elevated plus‐maze and light‐dark tests. Maternal aggression or male‐female encounters were not affected which suggests that male hormones are involved in the expression of suppressed aggressiveness. However, testosterone levels and brain monoamines can be excluded and found to be similar between GluR‐A deficient and wild‐type littermates. The reduced AMPA receptor levels caused by the lack of the GluR‐A subunit, and measured by a 30% reduction in hippocampal [3H]‐S‐AMPA binding, seem to be the reason for suppressed male aggressiveness. When we analyzed mice with reduced number of functional AMPA receptors mediated by the genomic introduced GluR‐A(Q582R) channel mutation, we observed again male‐specific suppressed aggression, providing additional evidence for GluR‐A subunit‐containing AMPA receptor involvement in aggression.
Sociability and preference for social novelty in five inbred strains: an approach to assess autistic‐like behavior in mice
Authors: Moy S. S. , Nadler J. J. , Perez A. , Barbaro R. P. , Johns J. M. , Magnuson T. R. , Piven J. , Crawley J. N.
Published: September 2004

Deficits in social interaction are important early markers for autism and related neurodevelopmental disorders with strong genetic components. Standardized behavioral assays that measure the preference of mice for initiating social interactions with novel conspecifics would be of great value for mutant mouse models of autism. We developed a new procedure to assess sociability and the preference for social novelty in mice. To quantitate sociability, each mouse was scored on measures of exploration in a central habituated area, a side chamber containing an unfamiliar conspecific (stranger 1) in a wire cage, or an empty side chamber. In a secondary test, preference for social novelty was quantitated by presenting the test mouse with a choice between the first, now‐familiar, conspecific (stranger 1) in one side chamber, and a second unfamiliar mouse (stranger 2) in the other side chamber. Parameters scored included time spent in each chamber and number of entries into the chambers. Five inbred strains of mice were tested, C57BL/6J, DBA/2J, FVB/NJ, A/J and B6129PF2/J hybrids. Four strains showed significant levels of sociability (spend‐ ing more time in the chamber containing stranger 1 than in the empty chamber) and a preference for social novelty (spending more time in the chamber containing stranger 2 than in the chamber containing the now‐familiar stranger 1). These social preferences were observed in both male and female mice, and in juveniles and adults. The exception was A/J, a strain that demonstrated a preference for the central chamber. Results are discussed in terms of potential applications of the new methods, and the proper controls for the interpretation of social behavior data, including assays for health, relevant sensory abilities and motor functions. This new standardized procedure to quantitate sociability and preference for social novelty in mice provides a method to assess tendencies for social avoidance in mouse models of autism.
The
Sinorhizobium meliloti
ABC Transporter Cho Is Highly Specific for Choline and Expressed in Bacteroids from
Medicago sativa
Nodules
Authors: Dupont Laurence , Garcia Isabelle , Poggi Marie-Christine , Alloing Geneviève , Mandon Karine , Le Rudulier Daniel
Published: September 2004

In
Sinorhizobium meliloti
, choline is the direct precursor of phosphatidylcholine, a major lipid membrane component in the
Rhizobiaceae
family, and glycine betaine, an important osmoprotectant. Moreover, choline is an efficient energy source which supports growth. Using a PCR strategy, we identified three chromosomal genes (
choXWV
) which encode components of an ABC transporter: ChoX (binding protein), ChoW (permease), and ChoV (ATPase). Whereas the best homology scores were obtained with components of betaine ProU-like systems, Cho is not involved in betaine transport. Site-directed mutagenesis of
choX
strongly reduced (60 to 75%) the choline uptake activity, and purification of ChoX, together with analysis of the ligand-binding specificity, showed that ChoX binds choline with a high affinity (
K

D

, 2.7 μM) and acetylcholine with a low affinity (
K

D

, 145 μM) but binds none of the betaines. Uptake competition experiments also revealed that ectoine, various betaines, and choline derivatives were not effective competitors for Cho-mediated choline transport. Thus, Cho is a highly specific high-affinity choline transporter. Choline transport activity and ChoX expression were induced by choline but not by salt stress. Western blotting experiments with antibodies raised against ChoX demonstrated the presence of ChoX in bacteroids isolated from nitrogen-fixing nodules obtained from
Medicago sativa
roots. The
choX
mutation did not have an effect on growth under standard conditions, and neither Nod nor Fix phenotypes were impaired in the mutant, suggesting that the remaining choline uptake system(s) still present in the mutant strain can compensate for the lack of Cho transporter.
Escherichia coli
Glutamate- and Arginine-Dependent Acid Resistance Systems Increase Internal pH and Reverse Transmembrane Potential
Authors: Richard Hope , Foster John W.
Published: September 2004

Due to the acidic nature of the stomach, enteric organisms must withstand extreme acid stress for colonization and pathogenesis.
Escherichia coli
contains several acid resistance systems that protect cells to pH 2. One acid resistance system, acid resistance system 2 (AR2), requires extracellular glutamate, while another (AR3) requires extracellular arginine. Little is known about how these systems protect cells from acid stress. AR2 and AR3 are thought to consume intracellular protons through amino acid decarboxylation. Antiport mechanisms then exchange decarboxylation products for new amino acid substrates. This form of proton consumption could maintain an internal pH (pH
i
) conducive to cell survival. The model was tested by estimating the pH
i
and transmembrane potential (ΔΨ) of cells acid stressed at pH 2.5. During acid challenge, glutamate- and arginine-dependent systems elevated pH
i
from 3.6 to 4.2 and 4.7, respectively. However, when pH
i
was manipulated to 4.0 in the presence or absence of glutamate, only cultures challenged in the presence of glutamate survived, indicating that a physiological parameter aside from pH
i
was also important. Measurements of ΔΨ indicated that amino acid-dependent acid resistance systems help convert membrane potential from an inside negative to inside positive charge, an established acidophile strategy used to survive extreme acidic environments. Thus, reversing ΔΨ may be a more important acid resistance strategy than maintaining a specific pH
i
value.
Two Arginine Repressors Regulate Arginine Biosynthesis inLactobacillus plantarum
Authors: Nicoloff Hervé , Arsène-Ploetze Florence , Malandain Cédric , Kleerebezem Michiel , Bringel Françoise
Published: September 2004

The repression of thecarABoperon encoding carbamoyl phosphate synthase leads toLactobacillus plantarumFB331 growth inhibition in the presence of arginine. This phenotype was used in a positive screening to select spontaneous mutants deregulated in the arginine biosynthesis pathway. Fourteen mutants were genetically characterized for constitutive arginine production. Mutations were located either in one of the arginine repressor genes (argR1orargR2) present inL. plantarumor in a putative ARG operator in the intergenic region of the bipolarcarAB-argCJBDFoperons involved in arginine biosynthesis. Although the presence of two ArgR regulators is commonly found in gram-positive bacteria, only single arginine repressors have so far been well studied inEscherichia coliorBacillus subtilis. InL. plantarum, arginine repression was abolished when ArgR1 or ArgR2 was mutated in the DNA binding domain, or in the oligomerization domain or when an A123D mutation occurred in ArgR1. A123, equivalent to the conserved residue A124 inE. coliArgR involved in arginine binding, was different in the wild-type ArgR2. Thus, corepressor binding sites may be different in ArgR1 and ArgR2, which have only 35% identical residues. Other mutants harbored wild-typeargRgenes, and 20 mutants have lost their ability to grow in normal air without carbon dioxide enrichment; this revealed a link between arginine biosynthesis and a still-unknown CO2-dependent metabolic pathway. In many gram-positive bacteria, the expression and interaction of different ArgR-like proteins may imply a complex regulatory network in response to environmental stimuli.
Structural Determinants Required To Target Penicillin-Binding Protein 3 to the Septum ofEscherichia coli
Authors: Piette André , Fraipont Claudine , den Blaauwen Tanneke , Aarsman Mirjam E. G. , Pastoret Soumya , Nguyen-Distèche Martine
Published: September 2004

InEscherichia coli, cell division is mediated by the concerted action of about 12 proteins that assemble at the division site to presumably form a complex called the divisome. Among these essential division proteins, the multimodular class B penicillin-binding protein 3 (PBP3), which is specifically involved in septal peptidoglycan synthesis, consists of a short intracellular M1-R23 peptide fused to a F24-L39 membrane anchor that is linked via a G40-S70 peptide to an R71-I236 noncatalytic module itself linked to a D237-V577 catalytic penicillin-binding module. On the basis of localization analyses of PBP3 mutants fused to green fluorescent protein by fluorescence microscopy, it appears that the first 56 amino acid residues of PBP3 containing the membrane anchor and the G40-E56 peptide contain the structural determinants required to target the protein to the cell division site and that none of the putative protein interaction sites present in the noncatalytic module are essential for the positioning of the protein to the division site. Based on the effects of increasing production of FtsQ or FtsW on the division of cells expressing PBP3 mutants, it is suggested that these proteins could interact. We postulate that FtsQ could play a role in regulating the assembly of these division proteins at the division site and the activity of the peptidoglycan assembly machineries within the divisome.
An SOS Response Induced by High Pressure inEscherichia coli
Authors: Aertsen Abram , Van Houdt Rob , Vanoirbeek Kristof , Michiels Chris W.
Published: September 2004

Although pressure is an important environmental parameter in microbial niches such as the deep sea and is furthermore used in food preservation to inactivate microorganisms, the fundamental understanding of its effects on bacteria remains fragmentary. Our group recently initiated differential fluorescence induction screening to search for pressure-inducedEscherichia colipromoters and has already reported induction of the heat shock regulon. Here the screening was continued, and we report for the first time that pressure induces a bona fide SOS response inE. coli, characterized by the RecA and LexA-dependent expression ofuvrA,recA, andsulA. Moreover, it was shown that pressure is capable of triggering lambda prophage induction inE. colilysogens. The remnant lambdoid e14 element, however, could not be induced by pressure, as opposed to UV irradiation, indicating subtle differences between the pressure-induced and the classical SOS response. Furthermore, the pressure-induced SOS response seems not to be initiated by DNA damage, sinceΔrecAandlexA1(Ind−) mutants, which are intrinsically hypersensitive to DNA damage, were not sensitized or were only very slightly sensitized for pressure-mediated killing and since pressure treatment was not found to be mutagenic. In light of these findings, the current knowledge of pressure-mediated effects on bacteria is discussed.
RsbV-Independent Induction of the SigB-Dependent General Stress Regulon ofBacillus subtilisduring Growth at High Temperature
Authors: Holtmann Gudrun , Brigulla Matthias , Steil Leif , Schütz Alexandra , Barnekow Karsta , Völker Uwe , Bremer Erhard
Published: September 2004

General stress proteins protectBacillus subtiliscells against a variety of environmental insults. This adaptive response is particularly important for nongrowing cells, to which it confers a multiple, nonspecific, and preemptive stress resistance. Induction of the general stress response relies on the alternative transcription factor, SigB, whose activity is controlled by a partner switching mechanism that also involves the anti-sigma factor, RsbW, and the antagonist protein, RsbV. Recently, the SigB regulon has been shown to be continuously induced and functionally important in cells actively growing at low temperature. With the exception of this chill induction, all SigB-activating stimuli identified so far trigger a transient expression of the SigB regulon that depends on RsbV. Through a proteome analysis and Northern blot and gene fusion experiments, we now show that the SigB regulon is continuously induced in cells growing actively at 51°C, close to the upper growth limit ofB. subtilis. This heat induction of SigB-dependent genes requires the environmental stress-responsive phosphatase RsbU, but not the metabolic stress-responsive phosphatase RsbP. RsbU dependence of SigB activation by heat is overcome in mutants that lack RsbV. In addition, loss of RsbV alone or in combination with RsbU triggers a hyperactivation of the general stress regulon exclusively at high temperatures detrimental for cell growth. These new facets of heat induction of the SigB regulon indicate that the current view of the complex genetic and biochemical regulation of SigB activity is still incomplete and that SigB perceives signals independent of the RsbV-mediated signal transduction pathways under heat stress conditions.
The Small Noncoding DsrA RNA Is an Acid Resistance Regulator inEscherichia coli
Authors: Lease Richard A. , Smith Dorie , McDonough Kathleen , Belfort Marlene
Published: September 2004

DsrA RNA is a small (87-nucleotide) regulatory RNA ofEscherichia colithat acts by RNA-RNA interactions to control translation and turnover of specific mRNAs. Two targets of DsrA regulation are RpoS, the stationary-phase and stress response sigma factor (σs), and H-NS, a histone-like nucleoid protein and global transcription repressor. Genes regulated globally by RpoS and H-NS include stress response proteins and virulence factors for pathogenicE. coli. Here, by using transcription profiling via DNA arrays, we have identified genes induced by DsrA. Steady-state levels of mRNAs from many genes increased with DsrA overproduction, including multiple acid resistance genes ofE. coli. Quantitative primer extension analysis verified the induction of individual acid resistance genes in thehdeAB,gadAX, andgadBCoperons.E. coliK-12 strains, as well as pathogenicE. coliO157:H7, exhibited compromised acid resistance indsrAmutants. Conversely, overproduction of DsrA from a plasmid rendered the acid-sensitivedsrAmutant extremely acid resistant. Thus, DsrA RNA plays a regulatory role in acid resistance. Whether DsrA targets acid resistance genes directly by base pairing or indirectly via perturbation of RpoS and/or H-NS is not known, but in either event, our results suggest that DsrA RNA may enhance the virulence of pathogenicE. coli.
New Protein-Protein Interactions Identified for the Regulatory and Structural Components and Substrates of the Type III Secretion System of the PhytopathogenXanthomonas axonopodisPathovar citri
Authors: Alegria Marcos C. , Docena Cassia , Khater Leticia , Ramos Carlos H. I. , da Silva Ana C. R. , Farah Chuck S.
Published: September 2004

We have initiated a project to identify protein-protein interactions involved in the pathogenicity of the bacterial plant pathogenXanthomonas axonopodispv. citri. Using a yeast two-hybrid system based on Gal4 DNA-binding and activation domains, we have focused on identifying interactions involving subunits, regulators, and substrates of the type III secretion system coded by thehrp(for hypersensitive response and pathogenicity),hrc(forhrpconserved), andhpa(forhrpassociated) genes. We have identified several previously uncharacterized interactions involving (i) HrpG, a two-component system response regulator responsible for the expression ofX. axonopodispv. citrihrpoperons, and XAC0095, a previously uncharacterized protein encountered only inXanthomonasspp.; (ii) HpaA, a protein secreted by the type III secretion system, HpaB, and the C-terminal domain of HrcV; (iii) HrpB1, HrpD6, and HrpW; and (iv) HrpB2 and HrcU. Homotropic interactions were also identified for the ATPase HrcN. These newly identified protein-protein interactions increase our understanding of the functional integration of phytopathogen-specific type III secretion system components and suggest new hypotheses regarding the molecular mechanisms underlyingXanthomonaspathogenicity.
Novel Xylose Dehydrogenase in the Halophilic Archaeon
Haloarcula marismortui
Authors: Johnsen Ulrike , Schönheit Peter
Published: September 2004

During growth of the halophilic archaeon
Haloarcula marismortui
on
d
-xylose, a specific
d
-xylose dehydrogenase was induced. The enzyme was purified to homogeneity. It constitutes a homotetramer of about 175 kDa and catalyzed the oxidation of xylose with both NADP
+
and NAD
+
as cosubstrates with 10-fold higher affinity for NADP
+
. In addition to
d
-xylose,
d
-ribose was oxidized at similar kinetic constants, whereas
d
-glucose was used with about 70-fold lower catalytic efficiency (
k
cat
/

K
m

). With the N-terminal amino acid sequence of the subunit, an open reading frame (ORF)—coding for a 39.9-kDA protein—was identified in the partially sequenced genome of
H. marismortui
. The function of the ORF as the gene designated
xdh
and coding for xylose dehydrogenase was proven by its functional overexpression in
Escherichia coli
. The recombinant enzyme was reactivated from inclusion bodies following solubilization in urea and refolding in the presence of salts, reduced and oxidized glutathione, and substrates. Xylose dehydrogenase showed the highest sequence similarity to glucose-fructose oxidoreductase from
Zymomonas mobilis
and other putative bacterial and archaeal oxidoreductases. Activities of xylose isomerase and xylulose kinase, the initial reactions of xylose catabolism of most bacteria, could not be detected in xylose-grown cells of
H. marismortui
, and the genes that encode them,
xylA
and
xylB
, were not found in the genome of
H. marismortui
. Thus, we propose that this first characterized archaeal xylose dehydrogenase catalyzes the initial step in xylose degradation by
H. marismortui
.
The hFbpABC Transporter from
Haemophilus influenzae
Functions as a Binding-Protein-Dependent ABC Transporter with High Specificity and Affinity for Ferric Iron
Authors: Anderson Damon S. , Adhikari Pratima , Nowalk Andrew J. , Chen Cheng Y. , Mietzner Timothy A.
Published: September 2004

Pathogenic
Haemophilus influenzae
,
Neisseria
spp. (
Neisseria gonorrhoeae
and
N. meningitidis
),
Serratia marcescens
, and other gram-negative bacteria utilize a periplasm-to-cytosol FbpABC iron transporter. In this study, we investigated the
H. influenzae
FbpABC transporter in a siderophore-deficient
Escherichia coli
background to assess biochemical aspects of FbpABC transporter function. Using a radiolabeled Fe
3+
transport assay, we established an apparent

K
m

= 0.9 μM and
V
max
= 1.8 pmol/10
7
cells/min for FbpABC-mediated transport. Complementation experiments showed that hFbpABC is dependent on the FbpA binding protein for transport. The ATPase inhibitor sodium orthovanadate demonstrated dose-dependent inhibition of FbpABC transport, while the protonmotive-force-inhibitor carbonyl cyanide
m
-chlorophenyl hydrazone had no effect. Metal competition experiments demonstrated that the transporter has high specificity for Fe
3+
and selectivity for trivalent metals, including Ga
3+
and Al
3+
, over divalent metals. Metal sensitivity experiments showed that several divalent metals, including copper, nickel, and zinc, exhibited general toxicity towards
E. coli
. Significantly, gallium-induced toxicity was specific only to
E. coli
expressing FbpABC. A single-amino-acid mutation in the gene encoding the periplasmic binding protein, FbpA(Y196I), resulted in a greatly diminished iron binding affinity

K
d

= 5.2 × 10
−4
M
−1
, ∼14 orders of magnitude weaker than that of the wild-type protein. Surprisingly, the mutant transporter [FbpA(Y196I)BC] exhibited substantial transport activity, ∼35% of wild-type transport, with

K
m

= 1.2 μM and
V
max
= 0.5 pmol/10
7
cells/min. We conclude that the FbpABC complexes possess basic characteristics representative of the family of bacterial binding protein-dependent ABC transporters. However, the specificity and high-affinity binding characteristics suggest that the FbpABC transporters function as specialized transporters satisfying the strict chemical requirements of ferric iron (Fe
3+
) binding and membrane transport.
Mutational Analysis of
Xanthomonas
Harpin HpaG Identifies a Key Functional Region That Elicits the Hypersensitive Response in Nonhost Plants
Authors: Kim Jung-Gun , Jeon Eunkyung , Oh Jonghee , Moon Jae Sun , Hwang Ingyu
Published: September 2004

HpaG is a type III-secreted elicitor protein of
Xanthomonas axonopodis
pv. glycines. We have determined the critical amino acid residues important for hypersensitive response (HR) elicitation by random and site-directed mutagenesis of HpaG and its homolog XopA. A plasmid clone carrying
hpaG
was mutagenized by site-directed mutagenesis, hydroxylamine mutagenesis, and error-prone PCR. A total of 52 mutants were obtained, including 51 single missense mutants and 1 double missense mutant. The HR elicitation activity was abolished in the two missense mutants [HpaG(L50P) and HpaG(L43P/L50P)]. Seven single missense mutants showed reduced activity, and the HR elicitation activity of the rest of the mutants was similar to that of wild-type HpaG. Mutational and deletion analyses narrowed the region essential for elicitor activity to the 23-amino-acid peptide (H
2
N-NQGISEKQLDQLLTQLIMALLQQ-COOH). A synthetic peptide of this sequence possessed HR elicitor activity at the same concentration as the HpaG protein. This region has 78 and 74% homology with 23- and 27-amino-acid regions of the HrpW harpin domains, respectively, from
Pseudomonas
and
Erwinia
spp. The secondary structure of the peptide is predicted to be an α-helix, as is the HrpW region that is homologous to HpaG. The predicted α-helix of HpaG is probably critical for the elicitation of the HR in tobacco plants. In addition, mutagenesis of a
xopA
gene yielded two gain-of-function mutants: XopA(F48L) and XopA(F48L/M52L). These results indicate that the 12 amino acid residues between L39 and L50 of HpaG have critical roles in HR elicitation in tobacco plants.
Complementation of an
Escherichia coli
DnaK Defect by Hsc70-DnaK Chimeric Proteins
Authors: Suppini Jean-Philippe , Amor Mouna , Alix Jean-Hervé , Ladjimi Moncef M.
Published: September 2004

Escherichia coli
DnaK and rat Hsc70 are members of the highly conserved 70-kDa heat shock protein (Hsp70) family that show strong sequence and structure similarities and comparable functional properties in terms of interactions with peptides and unfolded proteins and cooperation with cochaperones. We show here that, while the DnaK protein is, as expected, able to complement an
E. coli dnaK
mutant strain for growth at high temperatures and λ phage propagation, Hsc70 protein is not. However, an Hsc70 in which the peptide-binding domain has been replaced by that of DnaK is able to complement this strain for both phenotypes, suggesting that the peptide-binding domain of DnaK is essential to fulfill the specific functions of this protein necessary for growth at high temperatures and for λ phage replication. The implications of these findings on the functional specificities of the Hsp70s and the role of protein-protein interactions in the DnaK chaperone system are discussed.
Transcriptional Repressor CopR: Use of SELEX To Study the
copR
Operator Indicates that Evolution Was Directed at Maximal Binding Affinity
Authors: Freede Peggy , Brantl Sabine
Published: September 2004

CopR is one of the two copy number control elements of the streptococcal plasmid pIP501. It represses transcription of the
repR
mRNA encoding the essential replication initiator protein about 10- to 20-fold by binding to its operator region upstream of the
repR
promoter pII. CopR binds at two consecutive sites in the major groove of the DNA that share the consensus motif 5′-CGTG. Previously, the minimal operator was narrowed down to 17 bp, and equilibrium dissociation constants for DNA binding and dimerization were determined to be 0.4 nM and 1.4 μM, respectively. In this work, we used a SELEX procedure to study
copR
operator sequences of different lengths in combination with electrophoretic mobility shift assays of mutated
copR
operators as well as copy number determinations to assess the sequence requirements for CopR binding. The results suggest that in vivo evolution was directed at maximal binding affinity. Three simultaneous nucleotide exchanges outside the bases directly contacted by CopR only slightly affected CopR binding in vitro or copy numbers in vivo. Furthermore, the optimal spacer sequence was found to comprise 7 bp, to be AT rich, and to need an A/T and a T at the 3′ positions, whereas broad variations in the sequences flanking the minimal 17-bp operator were well tolerated.
A Novel Mutation within the CentralListeria monocytogenesRegulator PrfA That Results in Constitutive Expression of Virulence Gene Products
Authors: Wong Kendy K. Y. , Freitag Nancy E.
Published: September 2004

The PrfA protein ofListeria monocytogenesfunctions as a key regulatory factor for the coordinated expression of many virulence genes during bacterial infection of host cells. PrfA activity is controlled by multiple regulatory mechanisms, including an apparent requirement for either the presence of a cofactor or some form of posttranslational modification that regulates the activation of PrfA. In this study, we describe the identification and characterization of a novel PrfA mutation that results in constitutive activation of the PrfA protein. The PrfA L140F mutation was found to confer high-level expression of PrfA-regulated genes and to be functionally dominant over the wild-type allele. The presence of the PrfA L140F mutation resulted in the aggregation ofL. monocytogenesin broth culture and, unlike previously describedprfAmutations, appeared to be slightly toxic to the bacteria. High-level PrfA-dependent gene expression showed no additional increase inL. monocytogenesstrains containing an additional copy ofprfAL140F despite a >4-fold increase in PrfA protein levels. In contrast, the introduction of multiple copies of the wild-typeprfAallele toL. monocytogenesresulted in a corresponding increase in PrfA-dependent gene expression, although overall expression levels remained far below those observed for PrfA L140F strains. These results suggest a hierarchy of PrfA regulation, such that the relative levels of PrfA protein present within the cell correlate with the levels of PrfA-dependent gene expression when the protein is not in its fully activated state; however, saturating levels of the protein are then quickly reached when PrfA is converted to its active form. Regulation of the PrfA activation status must be an important facet ofL. monocytogenessurvival, as mutations that result in constitutive PrfA activation may have deleterious consequences for bacterial physiology.
Amino Acid Contacts between Sigma 70 Domain 4 and the Transcription Activators RhaS and RhaR
Authors: Wickstrum Jason R. , Egan Susan M.
Published: September 2004

The RhaS and RhaR proteins are transcription activators that respond to the availability of
l
-rhamnose and activate transcription of the operons in the
Escherichia coli
l
-rhamnose catabolic regulon. RhaR activates transcription of
rhaSR
, and RhaS activates transcription of the operon that encodes the
l
-rhamnose catabolic enzymes,
rhaBAD
, as well as the operon that encodes the
l
-rhamnose transport protein,
rhaT
. RhaS is 30% identical to RhaR at the amino acid level, and both are members of the AraC/XylS family of transcription activators. The RhaS and RhaR binding sites overlap the −35 hexamers of the promoters they regulate, suggesting they may contact the σ
70
subunit of RNA polymerase as part of their mechanisms of transcription activation. In support of this hypothesis, our lab previously identified an interaction between RhaS residue D241 and σ
70
residue R599. In the present study, we first identified two positively charged amino acids in σ
70
, K593 and R599, and three negatively charged amino acids in RhaR, D276, E284, and D285, that were important for RhaR-mediated transcription activation of the
rhaSR
operon. Using a genetic loss-of-contact approach we have obtained evidence for a specific contact between RhaR D276 and σ
70
R599. Finally, previous results from our lab separately showed that RhaS D250A and σ
70
K593A were defective at the
rhaBAD
promoter. Our genetic loss-of-contact analysis of these residues indicates that they identify a second site of contact between RhaS and σ
70
.
Transcription by
Methanothermobacter thermautotrophicus
RNA Polymerase In Vitro Releases Archaeal Transcription Factor B but Not TATA-Box Binding Protein from the Template DNA
Authors: Xie Yunwei , Reeve John N.
Published: September 2004

Transcription initiation in
Archaea
requires the assembly of a preinitiation complex containing the TATA- box binding protein (TBP), transcription factor B (TFB), and RNA polymerase (RNAP). The results reported establish the fate of
Methanothermobacter thermautotrophicus
TBP and TFB following transcription initiation by
M. thermautotrophicus
RNAP in vitro. TFB is released after initiation, during extension of the transcript from 4 to 24 nucleotides, but TBP remains bound to the template DNA. Regulation of archaeal transcription initiation by a repressor competition with TBP for TATA-box region binding must accommodate this observation.
mRNA Secondary Structure Modulates Translation of Tat-Dependent Formate Dehydrogenase N
Authors: Punginelli Claire , Ize Bérengère , Stanley Nicola R. , Stewart Valley , Sawers Gary , Berks Ben C. , Palmer Tracy
Published: September 2004

Formate dehydrogenase N (FDH-N) of
Escherichia coli
is a membrane-bound enzyme comprising FdnG, FdnH, and FdnI subunits organized in an (αβγ)
3
configuration. The FdnG subunit carries a Tat-dependent signal peptide, which localizes the protein complex to the periplasmic side of the membrane. We noted that substitution of the first arginine (R
5
) in the twin arginine signal sequence of FdnG for a variety of other amino acids resulted in a dramatic (up to 60-fold) increase in the levels of protein synthesized. Bioinformatic analysis suggested that the mRNA specifying the first 17 codons of
fdnG
forms a stable stem-loop structure. A detailed mutational analysis has demonstrated the importance of this mRNA stem-loop in modulating FDH-N translation.
Bacterial Thymidylate Synthase with Intein, Group II Intron, and Distinctive ThyX Motifs
Authors: Liu Xiang-Qin , Yang Jing
Published: September 2004

The ThyX class of thymidylate synthases was previously characterized by a common ThyX motif, RHRX7S. We report bacterial ThyX sequences having distinctive ThyX motifs, suggesting a more general ThyX motif, R/THRX7-8S. One ThyX sequence has an intein in its ThyX motif that was shown to do protein splicing and a group II intron in its gene, suggesting a hot spot for these self-splicing mobile elements.
Recombinant Cyclophilins Lack Nuclease Activity
Authors: Manteca Angel , Sanchez Jesus
Published: September 2004

Several single-domain prokaryotic and eukaryotic cyclophilins have been identified as also being unspecific nucleases with a role in DNA degradation during the lytic processes that accompany bacterial cell death and eukaryotic apoptosis. Evidence is provided here that the supposed nuclease activity of human and bacterial recombinant cyclophilins is due to contamination of the proteins by the host
Escherichia coli
endonuclease and is not an intrinsic property of these proteins.
Biofilm Mode of Growth of
Streptococcus intermedius
Favored by a Competence-Stimulating Signaling Peptide
Authors: Petersen Fernanda C. , Pecharki Daniele , Scheie Anne A.
Published: September 2004

Gram-positive and gram-negative bacteria use quorum sensing to coordinate population behavior. In several streptococci, quorum sensing mediated by competence-stimulating peptides (CSP) is associated with development of competence for transformation. We show here that a synthetic CSP favored the biofilm mode of growth of
Streptococcus intermedius
without affecting the rate of culture growth.
Extensive Genomic Polymorphism within
Mycobacterium avium
Authors: Semret Makeda , Zhai Gary , Mostowy Serge , Cleto Cynthia , Alexander David , Cangelosi Gerard , Cousins Debby , Collins Desmond M. , van Soolingen Dick , Behr Marcel A.
Published: September 2004

We have initiated comparative genomic analysis of
Mycobacterium avium
subspecies by DNA microarray, uncovering 14 large sequence polymorphisms (LSPs) comprising over 700 kb that distinguish
M. avium
subsp.
avium
from
M. avium
subsp.
paratuberculosis
. Genes predicted to encode metabolic pathways were overrepresented in the LSPs, and analysis revealed a polymorphism within the mycobactin biosynthesis operon that potentially explains the in vitro mycobactin dependence of
M. avium
subsp.
paratuberculosis
.
STRING-LIKE SOLUTIONS OF THE GAUGED NONLINEAR SIGMA MODEL
Authors: VANAIE HAMID REZA , RIAZI NEMATOLLAH
Published: September 2004

We show that the least energy conditions in the gauged nonlinear sigma model with Chern–Simons term lead to exact soliton-like solutions which look like domain walls. In fact, they are string-like solutions on the 2D plane. We will derive and discuss the corresponding solutions, and compute their energy and charge per unit length. The spin of the solutions is shown to vanish.
ESTIMATIVE OF INELASTICITY COEFFICIENT FROM EMULSION CHAMBER DATA
Authors: PORTELLA H. M. , de OLIVEIRA L. C. S. , LIMA C. E. C.
Published: September 2004

Atmospheric diffusion of high energy cosmic rays is studied analytically and the obtained integral electromagnetic fluxes are compared with the data measured by emulsion chamber detectors at mountain altitudes. We find a good consistency between them when the average nucleon inelasticity coefficient is varying between 0.50 and 0.65.
PENTAQUARKS
Authors: ZHU SHI-LIN
Published: September 2004

Since LEPS collaboration reported the first evidence of Θ+ pentaquark in early 2003, eleven other experimental groups have confirmed this exotic state while many other groups did not see any signal. If this state is further established by future high statistical experiments, its discovery shall be one of the most important events in hadron physics for the past three decades. This exotic baryon with such a low mass and so narrow a width imposes a big challenge to hadron theorists. Up to now, there have appeared more than two hundred theoretical papers trying to interpret this charming state. I will review some important theoretical developments on pentaquarks based on my biased personal views.
THE EULERIAN STRETCH OF A NETWORK TOPOLOGY AND THE ENDING GUARANTEE OF A CONVERGENCE ROUTING
Authors: BARTH DOMINIQUE , BERTHOME PASCAL , COHEN JOHANNE
Published: September 2004

In this paper, we focus on convergence packet routing techniques in an all-optical network, obtained from an Eulerian routing in the digraph modeling the target network. Given an Eulerian circuit [Formula: see text] in a digraph G, we deal with the maximal number [Formula: see text] of arcs that a packet has to follow on [Formula: see text] from its origin to its destination (we talk about the ending guarantee of the routing). We consider the Eulerian diameter of G as defined by [Formula: see text], where Eul(G) is the set of all the Eulerian circuits in G. After giving a preliminary result about the complexity of finding ℰ(G) for any digraph G, we give some lower and upper bounds of this parameter. The main part of the paper is devoted to the description of a combinatorial design of various network topologies having good Eulerian diameters.
EFFICIENT AND SECURE AUTONOMOUS SYSTEM BASED TRACEBACK
Authors: DURRESI ARJAN , PARUCHURI VAMSI , BAROLLI LEONARD , KANNAN RAJGOPAL , IYENGAR S. S.
Published: September 2004

The design of the IP protocol makes it difficult to reliably identify the originator of an IP packet, causing the defense against Distributed Denial of Service attacks to become one of the hardest problems on the Internet today. Previous solutions for this problem try to traceback to the exact origin of the attack by requiring every router's participation. For many reasons this requirement is impractical and the victim results with an approximate location of the attacker. Reconstruction of the whole path is also very difficult due to the sheer size of the Internet. This paper presents lightweight schemes for tracing back to the attack-originating AS instead to the exact origin itself. Once the attack-originating AS is determined, all further routers in the path to the attacker are within that AS and under the control of a single entity; which can presumably monitor local traffic in a more direct way than a generalized, Internet scale, packet marking scheme can. We furthermore, provide a scheme to prevent compromised routers from forging markings.
MODERATE CONCURRENCY CONTROL IN DISTRIBUTED OBJECT SYSTEMS
Authors: WATANABE KENICHI , SUGIYAMA YOUSUKE , ENOKIDO TOMOYA , TAKIZAWA MAKOTO
Published: September 2004

Objects are concurrently manipulated only through methods issued by multiple transactions in object-based systems. We first extend traditional read and write lock modes to methods on objects. We newly introduce availability and exclusion types of conflicting relations among methods. Then, we define a partially ordered relation on lock modes showing which modes are stronger than others. We newly propose a moderate concurrency control algorithm. Before manipulating an object through a method, the object is locked in a weaker mode than an intrinsic mode of the method. Then, the lock mode is escalated to the method mode. The weaker the initial mode is, the more concurrency is obtained but the more frequently deadlock occurs.
RATIONAL RADIAL DISTORTION MODELS OF CAMERA LENSES WITH ANALYTICAL SOLUTION FOR DISTORTION CORRECTION
Authors: MA LILI , CHEN YANGQUAN , MOORE KEVIN L.
Published: September 2004

The common approach to radial distortion is by the means of polynomial approximation, which introduces distortion-specific parameters into the camera model and requires estimation of these distortion parameters. The task of estimating radial distortion is to find a radial distortion model that allows easy undistortion as well as satisfactory accuracy. This paper presents a new class of rational radial distortion models with easy analytical undistortion formulae. Experimental results are presented to show that with this class of rational radial distortion models, satisfactory and comparable accuracy can be achieved.
THE PROCESS AND MODEL OF INFORMATION ACQUISITION
Authors: MEI TAO , WANG XIAOHUA , ZHANG JINGJING
Published: September 2004

As the first link in the information chain, Information Acquisition has become one of the most important branches in information science and technology. A basic information acquisition process is a multi-layer one that consists of the material layer for signal transduction, the data layer for sensor data fusion and the perception layer for information abstraction. While the material layer can be described as a continuous channel in information transmission, the data layer is a multiple discrete channel in the communication system. To analyze the perception layer, the μ-space theory in high dimensional geometric topology and the Γ-space theory based on statistical mechanics are introduced. A general model of information acquisition, which explains the working principle of intelligent sensors with self-adaptive function, is established according to the acquisition process and bionic principle.
PSYCHOPHYSICS, SOFT-HAPTICS AND INFORMATION ACQUISITION FOR INTERNET-BASED ROBOTIC TELEOPERATION
Authors: MENG MAX Q.-H. , LIU POLLEY R. , SHEK WILLIAM L. C. , LIU PETER X.
Published: September 2004

This article discusses the basic ideas and the practical applications of information acquisition in relation to psychophysics theory as implemented via the Soft-Haptics concept for Internet-based robotic teleoperation systems. In specific, a study on using acoustic display for navigation applications in Internet-based robot teleoperation is presented via psychophysics tests and experimental studies of the Internet-based teleoperated robotic systems to illustrate the ideas and concepts of this paper.
Magnetic resonance spectroscopy to measure hepatic triglyceride content: prevalence of hepatic steatosis in the general population
Authors: Szczepaniak Lidia S. , Nurenberg Pamela , Leonard David , Browning Jeffrey D. , Reingold Jason S. , Grundy Scott , Hobbs Helen H. , Dobbins Robert L.
Published: September 2004

Despite the increasing prevalence of nonalcoholic fatty liver disease (NAFLD), the criteria used to diagnose the disorder remain poorly defined. Localized proton magnetic resonance spectroscopy (MRS) accurately measures hepatic triglyceride content (HTGC) but has been used only in small research studies. Here, MRS was used to analyze the distribution of HTGC in 2,349 participants from the Dallas Heart Study (DHS). The reproducibility of the procedure was validated by showing that duplicate HTGC measurements were high correlated ( r = 0.99, P < 0.001) and that the coefficient of variation between measurements was low (8.5%). Intake of a high-fat meal did not significantly affect the measurements, and values were similar when measurements were made from the right and left hepatic lobes. To determine the “upper limit of normal” for HTGC, the distribution of HTGC was examined in the 345 subjects from the DHS who had no identifiable risk factors for hepatic steatosis (nonobese, nondiabetic subjects with minimal alcohol consumption, normal liver function tests, and no known liver disease). The 95th percentile of HTGC in these subjects was 5.56%, which corresponds to a hepatic triglyceride level of 55.6 mg/g. With this value as a cutoff, the prevalence of hepatic steatosis in Dallas County was estimated to be 33.6%. Thus MRS provides a sensitive, quantitative, noninvasive method to measure HTGC and, when applied to a large urban US population, revealed a strikingly high prevalence of hepatic steatosis.
Evidence against a role for insulin-signaling proteins PI 3-kinase and Akt in insulin resistance in human skeletal muscle induced by short-term GH infusion
Authors: Jessen Niels , Djurhuus Christian B. , Jørgensen Jens O. L. , Jensen Lasse S. , Møller Niels , Lund Sten , Schmitz Ole
Published: September 2004

Prolonged growth hormone (GH) excess is known to be associated with insulin resistance, but the underlying mechanisms remain unknown. The aim of this study was to assess the impact of GH on insulin-stimulated glucose metabolism and insulin signaling in human skeletal muscle. In a cross-over design, eight healthy male subjects (age 26.0 ± 0.8 yr and body mass index 24.1 ± 0.5 kg/m2) were infused for 360 min with either GH (Norditropin, 45 ng·kg−1·min−1) or saline. During the final 180 min of the infusion, a hyperinsulinemic euglycemic clamp was performed (insulin infusion rate: 1.2 mU·kg−1·min−1). Muscle biopsies from vastus lateralis were taken before GH/saline administration and after 60 min of hyperinsulinemia. GLUT4 content and insulin signaling, as assessed by insulin receptor substrate (IRS)-1-associated phosphatidylinositol 3-kinase and Akt activity were determined. GH levels increased to a mean (±SE) level of 20.0 ± 2.3 vs. 0.5 ± 0.2 μg/l after saline infusion ( P < 0.01). During GH infusion, the glucose infusion rate during hyperinsulinemia was reduced by 38% ( P < 0.01). In both conditions, free fatty acids were markedly suppressed during hyperinsulinemia. Despite skeletal muscle insulin resistance, insulin still induced a similar ∼3-fold rise in IRS-1-associated PI 3-kinase activity (269 ± 105 and 311 ± 71% compared with baseline, GH vs. saline). GH infusion did not change Akt protein expression, and insulin caused an ∼13-fold increase in Akt activity (1,309 ± 327 and 1,287 ± 173%) after both GH and saline infusion. No difference in total GLUT4 content was noted (114.7 ± 7.4 and 107.6 ± 16.7 arbitrary units, GH vs. saline, compared with baseline). In conclusion, insulin resistance in skeletal muscle induced by short-term GH administration is not associated with detectable changes in the upstream insulin-signaling cascade or reduction in total GLUT4. Yet unknown mechanisms in insulin signaling downstream of Akt may be responsible.