bluuebunny/crossref_metadata_2025 huggingface dataset:
Regulation of the Intracellular Free Iron Pool by Dpr Provides Oxygen Tolerance to
Streptococcus mutans
Authors: Yamamoto Yuji , Fukui Kôichi , Koujin Naoko , Ohya Hiroaki , Kimura Kazuhiko , Kamio Yoshiyuki
Published: September 2004

Dpr is an iron-binding protein required for oxygen tolerance in
Streptococcus mutans
. We previously proposed that Dpr could confer oxygen tolerance to the bacterium by sequestering intracellular free iron ions that catalyze generation of highly toxic radicals (Y. Yamamoto, M. Higuchi, L. B. Poole, and Y. Kamio, J. Bacteriol. 182:3740-3747, 2000; Y. Yamamoto, L. B. Poole, R. R. Hantgan, and Y. Kamio, J. Bacteriol. 184:2931-2939, 2002). Here, we examined the intracellular free iron status of wild-type (WT) and
dpr
mutant strains of
S. mutans
, before and after exposure to air, by using electron spin resonance spectrometry. Under anaerobic conditions, free iron ion concentrations of WT and
dpr
strains were 225.9 ± 2.6 and 333.0 ± 61.3 μM, respectively. Exposure of WT cells to air for 1 h induced Dpr expression and reduced intracellular free iron ion concentrations to 22.5 ± 5.3 μM; under these conditions,
dpr
mutant cells maintained intracellular iron concentration at 230.3 ± 28.8 μM. A decrease in cell viability and genomic DNA degradation was observed in the
dpr
mutant exposed to air. These data indicate that regulation of the intracellular free iron pool by Dpr is required for oxygen tolerance in
S. mutans
.
DegU-P Represses Expression of the Motilityfla-cheOperon inBacillus subtilis
Authors: Amati Giuseppe , Bisicchia Paola , Galizzi Alessandro
Published: September 2004

Bacillus subtilisimplements several adaptive strategies to cope with nutrient limitation experienced at the end of exponential growth. The DegS-DegU two-component system is part of the network involved in the regulation of postexponential responses, such as competence development, the production of exoenzymes, and motility. ThedegU32(Hy) mutation extends the half-life of the phosphorylated form of DegU (DegU-P); this in turn increases the production of alkaline protease, levan-sucrase, and other exoenzymes and inhibits motility and the production of flagella. The expression of the flagellum-specific sigma factor SigD, of the flagellin genehag, and of thefla-cheoperon is strongly reduced in adegU32(Hy) genetic background. To investigate the mechanism of action of DegU-P on motility, we isolated mutants ofdegU32(Hy) that completely suppressed the motility deficiency. The mutations were genetically mapped and characterized by PCR and sequencing. Most of the mutations were found to delete a transcriptional termination signal upstream of the main flagellar operon,fla-che, thus allowing transcriptional readthrough from thecodoperon. Two additional mutations improved the σA-dependent promoter sequence of thefla-cheoperon. Using an electrophoretic mobility shift assay, we have demonstrated that purified DegU binds specifically to the PApromoter region of thefla-cheoperon. The data suggest that DegU represses transcription of thefla-cheoperon, and they indicate a central role of the operon in regulating the synthesis and assembly of flagella.
Involvement of a Plasmid-Encoded Type IV Secretion System in the Plant Tissue Watersoaking Phenotype of
Burkholderia cenocepacia
Authors: Engledow Amanda S. , Medrano Enrique G. , Mahenthiralingam Eshwar , LiPuma John J. , Gonzalez Carlos F.
Published: September 2004

Burkholderia cenocepacia
strain K56-2, a representative of the
Burkholderia cepacia
complex, is part of the epidemic and clinically problematic ET12 lineage. The strain produced plant tissue watersoaking (ptw) on onion tissue, which is a plant disease-associated trait. Using plasposon mutagenesis, mutants in the ptw phenotype were generated. The translated sequence of a disrupted gene (
ptwD4
) from a ptw-negative mutant showed homology to VirD4-like proteins. Analysis of the region proximal to the transfer gene homolog identified a gene cluster located on the 92-kb resident plasmid that showed homology to type IV secretion systems. The role of
ptwD4
,
ptwC
,
ptwB4
, and
ptwB10
in the expression of ptw activity was determined by conducting site-directed mutagenesis. The ptw phenotype was not expressed by K56-2 derivatives with a disruption in
ptwD4
,
ptwB4
, or
ptwB10
but was observed in a derivative with a disruption in
ptwC
. Complementation of ptw-negative K56-2 derivatives in
trans
resulted in complete restoration of the ptw phenotype. In addition, analysis of culture supernatants revealed that the putative ptw effector(s) was a secreted, heat-stable protein(s) that caused plasmolysis of plant protoplasts. A second chromosomally encoded type IV secretion system with complete homology to the VirB-VirD system was identified in K56-2. Site-directed mutagenesis of key secretory genes in the VirB-VirD system did not affect expression of the ptw phenotype. Our findings indicate that in strain K56-2, the plasmid-encoded Ptw type IV secretion system is responsible for the secretion of a plant cytotoxic protein(s).
Denitrification Genes RegulateBrucellaVirulence in Mice
Authors: Baek Seung-Hun , Rajashekara Gireesh , Splitter Gary A. , Shapleigh James P.
Published: September 2004

Brucellais the causative agent of the zoonotic disease brucellosis, which is endemic in many parts of the world. Genome sequencing ofB. suisandB. melitensisrevealed that both are complete denitrifiers. To learn more about the role of denitrification in these animal pathogens, a study of the role of denitrification in the closely relatedB. neotomaewas undertaken. In contrast toB. suisandB. melitensis, it was found thatB. neotomaeis a partial denitrifier that can reduce nitrate to nitrite but no further. Examination of theB. neotomaegenome showed that a deletion in the denitrification gene cluster resulted in complete loss ofnirVand the partial deletion ofnirKandnnrA. Even though thenoroperon is intact, anorC-lacZpromoter fusion was not expressed inB. neotomae. However, thenorC-lacZfusion was expressed in the related denitrifierAgrobacterium tumefaciens, suggesting that the lack of expression inB. neotomaeis due to inactivation of NnrA. AnarK-lacZpromoter fusion was found to exhibit nitrate-dependent expression consistent with the partial denitrifier phenotype. Complementation of the deleted region inB. neotomaeby usingnirK,nirV, andnnrAfromB. melitensisrestored the ability ofB. neotomaeto reduce nitrite. There was a significant difference in the death of IRF-1−/−mice when infected withB. neotomaecontainingnirK, nirV, andnnrAand those infected with wild-typeB. neotomae. The wild-type strain killed all the infected mice, whereas most of the mice infected withB. neotomaecontainingnirK,nirV, andnnrAsurvived.
Sinorhizobium melilotiExoR and ExoS Proteins Regulate both Succinoglycan and Flagellum Production
Authors: Yao Shi-Yi , Luo Li , Har Katherine J. , Becker Anke , Rüberg Silvia , Yu Guan-Qiao , Zhu Jia-Bi , Cheng Hai-Ping
Published: September 2004

The production of theSinorhizobium melilotiexopolysaccharide, succinoglycan, is required for the formation of infection threads inside root hairs, a critical step during the nodulation of alfalfa (Medicago sativa) byS. meliloti. Two bacterial mutations,exoR95::Tn5andexoS96::Tn5, resulted in the overproduction of succinoglycan and a reduction in symbiosis. Systematic analyses of the symbiotic phenotypes of the two mutants demonstrated their reduced efficiency of root hair colonization. In addition, both theexoR95andexoS96mutations caused a marked reduction in the biosynthesis of flagella and consequent loss of ability of the cells to swarm and swim. Succinoglycan overproduction did not appear to be the cause of the suppression of flagellum biosynthesis. Further analysis indicated that both theexoR95andexoS96mutations affected the expression of the flagellum biosynthesis genes. These findings suggest that both the ExoR protein and the ExoS/ChvI two-component regulatory system are involved in the regulation of both succinoglycan and flagellum biosynthesis. These findings provide new avenues of understanding of the physiological changesS. meliloticells go through during the early stages of symbiosis and of the signal transduction pathways that mediate such changes.
Targeted Mutagenesis of the
Mycobacterium smegmatis mca
Gene, Encoding a Mycothiol-Dependent Detoxification Protein
Authors: Rawat Mamta , Uppal Mandeep , Newton Gerald , Steffek Micah , Fahey Robert C. , Av-Gay Yossef
Published: September 2004

Mycothiol (MSH), a functional analogue of glutathione (GSH) that is found exclusively in actinomycetes, reacts with electrophiles and toxins to form MSH-toxin conjugates. Mycothiol
S
-conjugate amidase (Mca) then catalyzes the hydrolysis of an amide bond in the
S
conjugates, producing a mercapturic acid of the toxin, which is excreted from the bacterium, and glucosaminyl inositol, which is recycled back to MSH. In this study, we have generated and characterized an allelic exchange mutant of the
mca
gene of
Mycobacterium smegmatis
. The
mca
mutant accumulates the
S
conjugates of the thiol-specific alkylating agent monobromobimane and the antibiotic rifamycin S. Introduction of
M. tuberculosis mca
epichromosomally or introduction of
M. smegmatis mca
integratively resulted in complementation of Mca activity and reduced levels of
S
conjugates. The mutation in
mca
renders the mutant strain more susceptible to electrophilic toxins, such as
N
-ethylmalemide, iodoacetamide, and chlorodinitrobenzene, and to several oxidants, such as menadione and plumbagin. Additionally we have shown that the
mca
mutant is also more susceptible to the antituberculous antibiotic streptomycin. Mutants disrupted in genes belonging to MSH biosynthesis are also more susceptible to streptomycin, providing further evidence that Mca detoxifies streptomycin in the mycobacterial cell in an MSH-dependent manner.
Among Multiple Phosphomannomutase Gene Orthologues, Only One Gene Encodes a Protein with Phosphoglucomutase and Phosphomannomutase Activities in
Thermococcus kodakaraensis
Authors: Rashid Naeem , Kanai Tamotsu , Atomi Haruyuki , Imanaka Tadayuki
Published: September 2004

Four orthologous genes (TK1108, TK1404, TK1777, and TK2185) that can be annotated as phosphomannomutase (PMM) genes (COG1109) have been identified in the genome of the hyperthermophilic archaeon
Thermococcus kodakaraensis
KOD1. We previously found that TK1777 actually encodes a phosphopentomutase. In order to determine which of the remaining three orthologues encodes a phosphoglucomutase (PGM), we examined the PGM activity in
T. kodakaraensis
cells and identified the gene responsible for this activity. Heterologous gene expression and purification and characterization of the recombinant protein indicated that TK1108 encoded a protein with high levels of PGM activity (690 U mg
−1
), along with high levels of PMM activity (401 U mg
−1
). Similar analyses of the remaining two orthologues revealed that their protein products exhibited neither PGM nor PMM activity. PGM activity and transcription of TK1108 in
T. kodakaraensis
were found to be higher in cells grown on starch than in cells grown on pyruvate. Our results clearly indicate that, among the four PMM gene orthologues in
T. kodakaraensis
, only one gene, TK1108, actually encodes a protein with PGM and PMM activities.
The
Bacteroides fragilis
Pathogenicity Island Is Contained in a Putative Novel Conjugative Transposon
Authors: Franco Augusto A.
Published: September 2004

The genetic element flanking the
Bacteroides fragilis
pathogenicity island (BfPAI) in enterotoxigenic
B. fragilis
(ETBF) strain 86-5443-2-2 and a related genetic element in NCTC 9343 were characterized. The results suggested that these genetic elements are members of a new family of conjugative transposons (CTns) not described previously. These putative CTns, designated CTn86 and CTn9343 for ETBF 86-5443-2-2 and NCTC 9343, respectively, differ from previously described
Bacteroides
species CTns in a number of ways. These new transposons do not carry
tetQ
, and the excision from the chromosome to form a circular intermediate is not regulated by tetracycline; they are predicted to differ in their mechanism of transposition; and their sequences have very limited similarity with CTnDOT or other described CTns. CTn9343 is 64,229 bp in length, contains 61 potential open reading frames, and both ends contain IS
21
transposases. Colony blot hybridization, PCR, and sequence analysis indicated that CTn86 has the same structure as CTn9343 except that CTn86 lacks a ∼7-kb region containing truncated integrase (
int2
) and
rteA
genes and it contains the BfPAI integrated between the
mob
region and the
bfmC
gene. If these putative CTns were to be demonstrated to be transmissible, this would suggest that the
bft
gene can be transferred from ETBF to nontoxigenic
B. fragilis
strains by a mechanism similar to that for the spread of antibiotic resistance genes.
Biosynthesis of Chloro-β-Hydroxytyrosine, a Nonproteinogenic Amino Acid of the Peptidic Backbone of Glycopeptide Antibiotics
Authors: Puk Oliver , Bischoff Daniel , Kittel Claudia , Pelzer Stefan , Weist Stefan , Stegmann Efthimia , Süssmuth Roderich D. , Wohlleben Wolfgang
Published: September 2004

The role of the putative P450 monooxygenase OxyD and the chlorination time point in the biosynthesis of the glycopeptide antibiotic balhimycin produced by
Amycolatopsis balhimycina
were analyzed. The
oxyD
gene is located directly downstream of the
bhp
(perhydrolase) and
bpsD
(nonribosomal peptide synthetase D) genes, which are involved in the synthesis of the balhimycin building block β-hydroxytyrosine (β-HT). Reverse transcriptase experiments revealed that
bhp
,
bpsD
, and
oxyD
form an operon.
oxyD
was inactivated by an in-frame deletion, and the resulting mutant was unable to produce an active compound. Balhimycin production could be restored (i) by complementation with an
oxyD
gene, (ii) in cross-feeding studies using
A. balhimycina
JR1 (a null mutant with a block in the biosynthesis pathway of the building blocks hydroxy- and dihydroxyphenylglycine) as an excretor of the missing precursor, and (iii) by supplementation of β-HT in the growth medium. These data demonstrated an essential role of OxyD in the formation pathway of this amino acid. Liquid chromatography-electrospray ionization-mass spectrometry analysis indicated the biosynthesis of completely chlorinated balhimycin by the
oxyD
mutant when culture filtrates were supplemented with nonchlorinated β-HT. In contrast, supplementation with 3-chloro-β-HT did not restore balhimycin production. These results indicated that the chlorination time point was later than the stage of free β-HT, most likely during heptapeptide synthesis.
Regulation of Tetralin Biodegradation and Identification of Genes Essential for Expression of
thn
Operons
Authors: Martínez-Pérez O. , Moreno-Ruiz E. , Floriano B. , Santero E.
Published: September 2004

The tetralin biodegradation genes of
Sphingomonas macrogolitabida
strain TFA are clustered in two closely linked and divergent operons. To analyze expression of both operons under different growth conditions, transcriptional and translational gene fusions of the first genes of each operon to
lacZ
have been constructed in plasmids unable to replicate in
Sphingomonas
and integrated by recombination into the genome of strain TFA. Expression analysis indicated that the transcription of both genes is induced in similar ways by the presence of tetralin. Gene expression in both operons is also subjected to overimposed catabolic repression. Two additional genes named
thnR
and
thnY
have been identified downstream of
thnCA3A4
genes. ThnR is similar to LysR-type regulators, and mutational analysis indicated that ThnR is strictly required for expression of the
thn
operons. Unlike other LysR-type regulators, ThnR does not repress its own synthesis. In fact, ThnR activates its own expression, since
thnR
is cotranscribed with the
thnCA3A4
genes. ThnY is similar to the ferredoxin reductase components of dioxygenase systems and shows the fer2 domain, binding a Cys
4
[2Fe-2S] iron sulfur center, and the FAD-binding domain, common to those reductases. However, it lacks the NAD-binding domain. Intriguingly, ThnY has a regulatory role, since it is also strictly required for expression of the
thn
operons. Given the similarity of ThnY to reductases and the possibility of its being present in the two redox states, it is tempting to speculate that ThnY is a regulatory component connecting expression of the
thn
operons to the physiological status of the cell.
Reversible Phase Variation in the
phnE
Gene, Which Is Required for Phosphonate Metabolism in
Escherichia coli
K-12
Authors: Iqbal Samina , Parker George , Davidson Helen , Moslehi-Rahmani Elham , Robson Robert L.
Published: September 2004

It is known that
Escherichia coli
K-12 is cryptic (Phn
−
) for utilization of methyl phosphonate (MePn) and that Phn
+
variants can be selected for growth on MePn as the sole P source. Variants arise from deletion via a possible slip strand mechanism of one of three direct 8-bp repeat sequences in
phnE
, which restores function to a component of a putative ABC type transporter. Here we show that Phn
+
variants are present at the surprisingly high frequency of >10
−2
in K-12 strains. Amplified-fragment length polymorphism analysis was used to monitor instability in
phnE
in various strains growing under different conditions. This revealed that, once selection for growth on MePn is removed,
Phn
+
revertants reappear and accumulate at high levels through reinsertion of the 8-bp repeat element sequence. It appears that, in K-12,
phnE
contains a high-frequency reversible gene switch, producing phase variation which either allows (“on” form) or blocks (“off” form) MePn utilization. The switch can also block usage of other metabolizable alkyl phosphonates, including the naturally occurring 2-aminoethylphosphonate. All K-12 strains, obtained from collections, appear in the “off” form even when bearing mutations in
mutS
,
mutD
, or
dnaQ
which are known to enhance slip strand events between repetitive sequences. The ability to inactivate the
phnE
gene appears to be unique to K-12 strains since the B strain is naturally Phn
+
and lacks the inactivating 8-bp insertion in
phnE
, as do important pathogenic strains for which genome sequences are known and also strains isolated recently from environmental sources.
Arginine Biosynthesis inThermotoga maritima: Characterization of the Arginine-SensitiveN-Acetyl-l-Glutamate Kinase
Authors: Fernández-Murga M. Leonor , Gil-Ortiz Fernando , Llácer José L. , Rubio Vicente
Published: September 2004

To help clarify the control of arginine synthesis inThermotoga maritima, the putative gene (argB) forN-acetyl-l-glutamate kinase (NAGK) from this microorganism was cloned and overexpressed, and the resulting protein was purified and shown to be a highly thermostable and specific NAGK that is potently and selectively inhibited by arginine. Therefore, NAGK is inT. maritimathe feedback control point of arginine synthesis, a process that in this organism involves acetyl group recycling and appears not to involve classical acetylglutamate synthase. The inhibition of NAGK by arginine was found to be pH independent and to depend sigmoidally on the concentration of arginine, with a Hill coefficient (N) of ∼4, and the 50% inhibitory arginine concentration (I0.5) was shown to increase with temperature, approaching above 65°C the I0.50observed at 37°C with the mesophilic NAGK ofPseudomonas aeruginosa(the best-studied arginine-inhibitable NAGK). At 75°C, the inhibition by arginine ofT. maritimaNAGK was due to a large increase in theKmfor acetylglutamate triggered by the inhibitor, but at 37°C arginine also substantially decreased theVmaxof the enzyme. The NAGKs ofT. maritimaandP. aeruginosabehaved in gel filtration as hexamers, justifying the sigmoidicity and high Hill coefficient of arginine inhibition, and arginine or the substrates failed to disaggregate these enzymes. In contrast,Escherichia coliNAGK is not inhibited by arginine and is dimeric, and thus the hexameric architecture may be an important determinant of arginine sensitivity. Potential thermostability determinants ofT. maritimaNAGK are also discussed.
Characterization of Heme Uptake Cluster Genes in the Fish PathogenVibrio anguillarum
Authors: Mouriño Susana , Osorio Carlos R. , Lemos Manuel L.
Published: September 2004

Vibrio anguillarumcan utilize hemin and hemoglobin as sole iron sources. In previous work we identified HuvA, theV. anguillarumouter membrane heme receptor by complementation of a heme utilization mutant with a cosmid clone (pML1) isolated from a genomic library ofV. anguillarum. In the present study, we describe a gene cluster contained in cosmid pML1, coding for nine potential heme uptake and utilization proteins: HuvA, the heme receptor; HuvZ and HuvX; TonB, ExbB, and ExbD; HuvB, the putative periplasmic binding protein; HuvC, the putative inner membrane permease; and HuvD, the putative ABC transporter ATPase. AV. anguillarumstrain with an in-frame chromosomal deletion of the nine-gene cluster was impaired for growth with heme or hemoglobin as the sole iron source. Single-gene in-frame deletions were constructed, demonstrating that each of thehuvAZBCDgenes are essential for utilization of heme as an iron source inV. anguillarum, whereashuvXis not. When expressed inEscherichia coli hemA(strain EB53), a plasmid carrying the gene for the heme receptor, HuvA, was sufficient to allow the use of heme as the porphyrin source. For utilization of heme as an iron source inE. coli ent(strain 101ESD), thetonB exbBDandhuvBCDgenes were required in addition tohuvA. TheV. anguillarumheme uptake cluster shows some differences in gene arrangement when compared to homologous clusters described for otherVibriospecies.
AglZ Is a Filament-Forming Coiled-Coil Protein Required for Adventurous Gliding Motility ofMyxococcus xanthus
Authors: Yang Ruifeng , Bartle Sarah , Otto Rebecca , Stassinopoulos Angela , Rogers Matthew , Plamann Lynda , Hartzell Patricia
Published: September 2004

TheaglZgene ofMyxococcus xanthuswas identified from a yeast two-hybrid assay in which MglA was used as bait. MglA is a 22-kDa cytoplasmic GTPase required for both adventurous and social gliding motility and sporulation. Genetic studies showed thataglZis part of the A motility system, because disruption or deletion ofaglZabolished movement of isolated cells andaglZ sglKdouble mutants were nonmotile. TheaglZgene encodes a 153-kDa protein that interacts with purified MglA in vitro. The N terminus of AglZ shows similarity to the receiver domain of two-component response regulator proteins, while the C terminus contains heptad repeats characteristic of coiled-coil proteins, such as myosin. Consistent with this motif, expression of AglZ inEscherichia coliresulted in production of striated lattice structures. Similar to the myosin heavy chain, the purified C-terminal coiled-coil domain of AglZ forms filament structures in vitro.
Inactivations of
rsbU
and
sarA
by IS
256
Represent Novel Mechanisms of Biofilm Phenotypic Variation in
Staphylococcus epidermidis
Authors: Conlon Kevin M. , Humphreys Hilary , O'Gara James P.
Published: September 2004

Expression of
ica
operon-mediated biofilm formation in
Staphylococcus epidermidis
RP62A is subject to phase variable regulation. Reversible transposition of IS
256
into
icaADBC
or downregulation of
icaADBC
expression are two important mechanisms of biofilm phenotypic variation. Interestingly, the presence of IS
256
was generally associated with a more rapid rate of phenotypic variation, suggesting that IS
256
insertions outside the
ica
locus may affect
ica
transcription. Consistent with this, we identified variants with diminished
ica
expression, which were associated with IS
256
insertions in the σ
B
activator
rsbU
or
sarA
. Biofilm development and
ica
expression were activated only by ethanol and not NaCl in
rsbU
::IS
256
insertion variants, which were present in ∼11% of all variants. σ
B
activity was impaired in
rsbU
::IS
256
variants, as evidenced by reduced expression of the σ
B
-regulated genes
asp23
,
csb9
, and
rsbV
. Moreover, expression of
sarA
, which is σ
B
regulated, and SarA-regulated RNAIII were also suppressed. A biofilm-forming phenotype was restored to
rsbU
::IS
256
variants only after repeated passage and was not associated with IS
256
excision from
rsbU
. Only one
sarA
::IS
256
insertion mutant was identified among 43 biofilm-negative variants. Both NaCl and ethanol-activated
ica
expression in this
sarA
::IS
256
variant, but only ethanol increased biofilm development. Unlike
rsbU
::IS
256
variants, reversion of the
sarA
::IS
256
variant to a biofilm-positive phenotype was accompanied by precise excision of IS
256
from
sarA
and restoration of normal
ica
expression. These data identify new roles for IS
256
in
ica
and biofilm phenotypic variation and demonstrate the capacity of this element to influence the global regulation of transcription in
S. epidermidis
.
Bacillus subtilisStoA Is a Thiol-Disulfide Oxidoreductase Important for Spore Cortex Synthesis
Authors: Erlendsson Lýður S. , Möller Mirja , Hederstedt Lars
Published: September 2004

Bacillus subtilisis an endospore-forming bacterium. There are indications that protein disulfide linkages occur in spores, but the role of thiol-disulfide chemistry in spore synthesis is not understood. Thiol-disulfide oxidoreductases catalyze formation or breakage of disulfide bonds in proteins. CcdA is the onlyB. subtilisthiol-disulfide oxidoreductase that has previously been shown to play some role in endospore biogenesis. In this work we show that lack of the StoA (YkvV) protein results in spores sensitive to heat, lysozyme, and chloroform. Compared to CcdA deficiency, StoA deficiency results in a 100-fold-stronger negative effect on sporulation efficiency. StoA is a membrane-bound protein with a predicted thioredoxin-like domain probably localized in the intermembrane space of the forespore. Electron microscopy of spores of CcdA- and StoA-deficient strains showed that the spore cortex is absent in both cases. The BdbD protein catalyzes formation of disulfide bonds in proteins on the outer side of the cytoplasmic membrane but is not required for sporulation. Inactivation ofbdbDwas found to suppress the sporulation defect of a strain deficient in StoA. Our results indicate that StoA is a thiol-disulfide oxidoreductase that is involved in breaking disulfide bonds in cortex components or in proteins important for cortex synthesis.
The Paralogous Pairs of Genes Involved in Clavulanic Acid and Clavam Metabolite Biosynthesis Are Differently Regulated in
Streptomyces clavuligerus
Authors: Tahlan Kapil , Anders Cecilia , Jensen Susan E.
Published: September 2004

Carboxyethylarginine synthase, encoded by the paralogous
ceaS1
and
ceaS2
genes, catalyzes the first reaction in the shared biosynthetic pathway leading to clavulanic acid and the other clavam metabolites in
Streptomyces clavuligerus
. The nutritional regulation of
ceaS1
and
ceaS2
expression was analyzed by reverse transcriptase PCR and by the use of the enhanced green fluorescent protein-encoding gene (e
gfp
) as a reporter.
ceaS1
was transcribed in complex soy medium only, whereas
ceaS2
was transcribed in both soy and defined starch-asparagine (SA) media. The transcriptional start points of the two genes were also mapped to a C residue 98 bp upstream of
ceaS1
and a G residue 51 bp upstream of the
ceaS2
start codon by S1 nuclease protection and primer extension analyses. Furthermore, transcriptional mapping of the genes encoding the beta-lactam synthetase (
bls1
) and proclavaminate amidinohydrolase (
pah1
) isoenzymes from the paralogue gene cluster indicated that a single polycistronic transcript of ∼4.9 kb includes
ceaS1
,
bls1
, and
pah1
. The expression of
ceaS1
and
ceaS2
in a mutant strain defective in the regulatory protein CcaR was also examined.
ceaS1
transcription was not affected in the
ccaR
mutant, whereas that of
ceaS2
was greatly reduced compared to the wild-type strain. Overall, our results suggest that different mechanisms are involved in regulating the expression of
ceaS1
and
ceaS2
, and presumably also of other paralogous genes that encode proteins involved in the early stages of clavulanic acid and clavam metabolite biosynthesis.
Temporal Global Changes in Gene Expression during Temperature Transition in
Yersinia pestis
Authors: Motin Vladimir L. , Georgescu Anca M. , Fitch Joseph P. , Gu Pauline P. , Nelson David O. , Mabery Shalini L. , Garnham Janine B. , Sokhansanj Bahrad A. , Ott Linda L. , Coleman Matthew A. , Elliott Jeffrey M. , Kegelmeyer Laura M. , Wyrobek Andrew J. , Slezak Thomas R. , Brubaker Robert R. , Garcia Emilio
Published: September 2004

DNA microarrays encompassing the entire genome of
Yersinia pestis
were used to characterize global regulatory changes during steady-state vegetative growth occurring after shift from 26 to 37°C in the presence and absence of Ca
2+
. Transcriptional profiles revealed that 51, 4, and 13 respective genes and open reading frames (ORFs) on pCD, pPCP, and pMT were thermoinduced and that the majority of these genes carried by pCD were downregulated by Ca
2+
. In contrast, Ca
2+
had little effect on chromosomal genes and ORFs, of which 235 were thermally upregulated and 274 were thermally downregulated. The primary consequence of these regulatory events is profligate catabolism of numerous metabolites available in the mammalian host.
The Secretion Signal of YopN, a Regulatory Protein of the
Yersinia enterocolitica
Type III Secretion Pathway
Authors: Goss John W. , Sorg Joseph A. , Ramamurthi Kumaran S. , Ton-That Hung , Schneewind Olaf
Published: September 2004

The type III secretion signal of
Yersinia enterocolitica
YopN was mapped using a gene fusion approach.
yopN
codons 1 to 12 were identified as critical for signal function. Several synonymous mutations that abolish secretion of hybrid proteins without altering the codon specificity of
yopN
mRNA were identified.
A BASIC CLASS OF TWISTED OPEN WZW STRINGS
Authors: HALPERN M. B. , HELFGOTT C.
Published: September 2004

Recently, Giusto and Halpern reported the open-string description of a certain basic class of untwisted open WZW strings, including their associated noncommutative geometry and open-string KZ equations. In this paper, we combine this development with results from the theory of current-algebraic orbifolds to find the open-string description of a corresponding basic class of twisted open WZW strings, which begin and end on different WZW branes. The basic class of twisted open WZW strings is in 1-to-1 correspondence with the twisted sectors of all closed-string WZW orbifolds, and moreover, the basic class can be decomposed into a large collection of open-string WZW orbifolds. At the classical level, these open-string orbifolds exhibit new twisted noncommutative geometries, and we also find the relevant twisted open-string KZ equations which describe these orbifolds at the quantum level. In a related development, we also formulate the closed-string description (in terms of twisted boundary states) of the general twisted open WZW string.
ANALYTIC INVARIANT CHARGE AND THE LATTICE STATIC QUARK–ANTIQUARK POTENTIAL
Authors: NESTERENKO A. V.
Published: September 2004

A recently developed model for the QCD analytic invariant charge is compared with quenched lattice simulation data on the static quark–antiquark potential. By employing this strong running coupling one is able to obtain the confining quark–antiquark potential in the framework of the one-gluon exchange model. To achieve this objective a technique for evaluating the integrals of a required form is developed. Special attention is paid here to removing the divergences encountered the calculations. All this enables one to examine the asymptotic behavior of the potential at both small and large distances with high accuracy. An explicit expression for the quark–antiquark potential, which interpolates between these asymptotics, and satisfies the concavity condition, is proposed. The derived potential coincides with the perturbative results at small distances, and it is in a good agreement with the lattice data in the nonperturbative physically-relevant region. An estimation of the parameter Λ QCD is obtained for the case of pure gluodynamics. It is found to be consistent with all the previous estimations of Λ QCD in the framework of approach in hand.
EMBEDDING A COMPLETE BINARY TREE INTO A THREE-DIMENSIONAL GRID
Authors: BEIN WOLFGANG W. , LARMORE LAWRENCE L. , SHIELDS CHARLES O. , SUDBOROUGH I. HAL
Published: September 2004

We describe total congestion 1 embeddings of complete binary trees into three dimensional grids with low expansion ratio r. That is, we give a one-to-one embedding of any complete binary tree into a hexahedron shaped grid such that (a) the number of nodes in the grid is at most r times the number of nodes in the tree, and (b) no tree nodes or edges occupy the same grid positions. The first strategy embeds trees into cube shaped 3D grids. That is, 3D grids in which all dimensions are roughly equal in size, and which thus have no limit in the number of layers. The technique uses a recursive scheme, and we obtain an expansion ratio of r=1.09375. We then give strategies which embed trees into flat 3D grid shapes. That is, we map complete binary trees into 3D grids with a fixed, small number of layers k. Using again a recursive scheme, for k=2, we obtain r=1.25. By a rather different technique, which intricately weaves the branches of various subtrees into each other, we are able to obtain very tight embeddings: We have r=1.171875 for embeddings into five layer grids and r=1.09375 for embeddings into seven layer grids.
THE MESH WITH BINARY TREE NETWORKS: AN ENHANCED MESH WITH LOW BUS-LOADING
Authors: DHARMASENA HETTIHE P. , VAIDYANATHAN RAMACHANDRAN
Published: September 2004

The literature abounds with enhanced mesh models and numerous results for them. These models employ buses connecting a large (usually a polynomial) number of processors resulting in high bus loading and a very low bus data rate. In this paper we propose an enhanced mesh model called the mesh with binary-tree (multiple bus) network (MBTN) with low bus loading. We measure the cost of the model in terms of its loading, degree (connections per processor), number of buses and layout area. We also devise an algorithm to perform prefix computations on the MBTN. The MBTN's parameters can be selected to provide a wide range of tradeoffs between its cost and performance. Most importantly, for every major existing enhanced mesh model, there is a choice of parameters that makes the MBTN superior to the existing model.

Key to the MBTN is a novel implementation of a binary-tree algorithm on a multiple bus network (MBN). We propose two constant loading MBNs that can run binary tree algorithms optimally. We also show that any network (such as the MBTN) that is capable of running binary tree algorithms efficiently can also solve linear recurrences efficiently. These results may be of independent interest.
POLYNOMIAL TIME SOLVABILITY OF THE WEIGHTED RING ARC-LOADING PROBLEM WITH INTEGER SPLITTING
Authors: YUAN JINJIANG , ZHOU SANMING
Published: September 2004

In the Weighted Ring Arc-Loading Problem with Integer Splitting, we are given a set of communication requests each associated with a non-negative integer weight. The problem is to find a routing scheme such that the maximum load on arcs of the ring is minimized, subject to that the weight of each request may be split into two integral parts routed in two directions around the ring, where the load of an arc is the sum of parts routed through the arc. A pseudo-polynomial algorithm for this problem is implied by a result in [G. Wilfong and P. Winkler, Ring routing and wavelength translation, Proceedings of the 9th ACM-SIAM Symposium on Discrete Algorithms, San Fancisco, CA, 1998, 333-341]. By refining the rounding technique developed in the same paper, we prove that the problem can be solved in polynomial time.
A NEW APPROACH FOR MULTI-SYSTEM/SENSOR DECISION FUSION BASED ON JOINT MEASURES
Authors: RASHIDI ALI J. , GHASSEMIAN M. HASSAN
Published: September 2004

This article describes the joint measures method as a new powerful method for the development of a high performance multi-sensor data/image fusion scheme at the decision level. The images are received from distributed multiple sensors, which sense the targets in different spectral bands including visible, infrared, thermal and microwave. At first, we study the decision fusion methods, including voting schemes, rank based algorithm, Bayesian inference, and the Dempster-Shafer method. Then, we extract the mathematical properties of multi-sensor local classification results and use them for modeling of the classifier performances by the two new measures, i.e. the plausibility and correctness. Then we establish the plausibility and correctness distribution vectors and matrices for introducing the two improvements of the Dempster-Shafer method, i.e. the DS (CM) and DS (PM) methods. After that we introduce the joint measures decision fusion method based on using these two measures jointly. The Joint Measures Method (JMM) can deal with any decision fusion problem in the case of uncertain local classifiers results as well as clear local classifiers results. Finally, we deploy the new and previous methods for the fusion of the two different sets of multispectral image classification local results and we also compare their reliabilities, the commission errors and the omission errors. The results obviously show that the DS (PM), DS (CM) and JMM methods which use the special properties of the local classifiers and classes, have much better accuracies and reliabilities than other methods. In addition, we show that the reliability of the JMM is at least 3% higher than all other methods.
CONCEPT CELL MODEL FOR KNOWLEDGE REPRESENTATION
Authors: ZHENG JEFFREY Z. J. , ZHENG CHRISTIAN H. H. , KUNII TOSIYASU L.
Published: September 2004

This paper proposes a unified knowledge model establishing a core structure of knowledge representation in natural and artificial intelligence systems. The Concept Cell Model proposes the use of acyclic lattices to model a concept formed from a knowledge network of simpler concepts. Declarative and procedural knowledge are explicitly defined as the time-invariant and time-variant relationship of concepts. Examples of a Restaurant Servicing Concept Cell and an extended Shopping Complex Concept Cell are used to demonstrate the functionality of this model. Major existing theoretic and engineering Ontology knowledge schools are compared under this framework.
ANALYSIS OF HUMAN WALKING TRAJECTORIES FOR SURVEILLANCE
Authors: LEE KA KEUNG , XU YANGSHENG
Published: September 2004

Surveillance of public places has become a worldwide concern in recent years. The ability to identify abnormal human behaviors in real-time is fundamental to the success of intelligent surveillance systems. The recognition of abnormal and suspicious human walking patterns is an important step towards the achievement of this goal. In this research, we develop an intelligent visual surveillance system that can classify normal and abnormal human walking trajectories in outdoor environments by learning from demonstration. The system takes into account both the local and global characteristics of the observed trajectories and is able to identify their normality in real-time. By utilizing support vector learning and a similarity measure based on hidden Markov models, the developed system has produced satisfactory results on real-life data during testing. Moreover, we utilize the approach of longest common subsequence (LCSS) in determining the similarity between different types of walking trajectories. In order to establish the position and speed boundaries required for the similarity measure, we compare the performance of a number of approaches, including fixed boundary values, variable boundary values, learning boundary by support vector regression, and learning boundary by cascade neural networks.
SELF-POWERED WIRELESS TEMPERATURE SENSING USING MEMS-BASED AA-SIZE ENERGY TRANSDUCER
Authors: LEE JOHNNY M. H. , YUEN STEVE C. L. , LUK MIMI H. M. , CHAN GORDON M. H. , LEI KING FONG , LI WEN J. , LEONG PHILIP H. W. , YAM YEUNG
Published: September 2004

This paper presents the design and experimental results of a Micro Power Generator (MPG) which harvests mechanical energy from its environment and converts this energy into useful electrical power. The energy transduction component is mainly a magnet and a resonating spring made using SU-8 molding and MEMS electroplating technologies. We have shown that when the MPG is packaged into an AA battery size container along with a power-management circuit that consists of rectifiers and a capacitor, it is capable of producing ~1.6 V DC when charged for less than 1 min. Our goal is to realize a MPG to function with low input mechanical frequencies while producing enough power for low-power wireless applications.
Mitochondrial metabolism reveals a functional architecture in intact islets of Langerhans from normal and diabeticPsammomys obesus
Authors: Katzman S. M. , Messerli M. A. , Barry D. T. , Grossman A. , Harel T. , Wikstrom J. D. , Corkey B. E. , Smith P. J. S. , Shirihai O. S.
Published: September 2004

The cells within the intact islet of Langerhans function as a metabolic syncytium, secreting insulin in a coordinated and oscillatory manner in response to external fuel. With increased glucose, the oscillatory amplitude is enhanced, leading to the hypothesis that cells within the islet are secreting with greater synchronization. Consequently, non-insulin-dependent diabetes mellitus (NIDDM; type 2 diabetes)-induced irregularities in insulin secretion oscillations may be attributed to decreased intercellular coordination. The purpose of the present study was to determine whether the degree of metabolic coordination within the intact islet was enhanced by increased glucose and compromised by NIDDM. Experiments were performed with isolated islets from normal and diabetic Psammomys obesus. Using confocal microscopy and the mitochondrial potentiometric dye rhodamine 123, we measured mitochondrial membrane potential oscillations in individual cells within intact islets. When mitochondrial membrane potential was averaged from all the cells in a single islet, the resultant waveform demonstrated clear sinusoidal oscillations. Cells within islets were heterogeneous in terms of cellular synchronicity (similarity in phase and period), sinusoidal regularity, and frequency of oscillation. Cells within normal islets oscillated with greater synchronicity compared with cells within diabetic islets. The range of oscillatory frequencies was unchanged by glucose or diabetes. Cells within diabetic (but not normal) islets increased oscillatory regularity in response to glucose. These data support the hypothesis that glucose enhances metabolic coupling in normal islets and that the dampening of oscillatory insulin secretion in NIDDM may result from disrupted metabolic coupling.
Role of CD38 in myometrial Ca2+transients: modulation by progesterone
Authors: Thompson Michael , Barata da Silva Hosana , Zielinska Weronika , White Thomas A. , Bailey Jeffrey P. , Lund Frances E. , Sieck Gary C. , Chini Eduardo N.
Published: September 2004

Oxytocin-induced Ca2+transients play an important role in myometrial contractions. Here, using a knockout model, we found that the enzyme CD38, responsible for the synthesis of the second messenger cyclic ADP-ribose (cADPR), plays an important role in the oxytocin-induced Ca2+transients and contraction. We also observed that CD38 is necessary for TNF-α-increased agonist-stimulated Ca2+transients in human myometrial cells. We provide experimental evidence that the TNF-α effect is mediated by increased expression of the enzyme CD38. First, we observed that TNF-α increased oxytocin-induced Ca2+transients and CD38 expression in human myometrial cells. Moreover, using small interference RNA technology, we observed that TNF-α stimulation of agonist-induced Ca2+transients was abolished by blocking the expression of CD38. In control experiments, we observed that activation of the component of the TNF-α signaling pathway, NF-κB, was not affected by the treatments. Finally, we observed that the effects of TNF-α on CD38 cyclase and oxytocin-induced Ca2+transients are abolished by progesterone. In conclusion, we provide the first experimental evidence that CD38 is important for myometrial Ca2+transients and contraction. Moreover, CD38 is necessary for the TNF-α-mediated augmentation of agonist-induced Ca2+transients in myometrial cells. We propose that the balance between cytokines and placental steroids regulates the expression of CD38 in vivo and cell responsiveness to oxytocin.
Mechanism, temporal patterns, and magnitudes of the metabolic responses to the KATPchannel agonist diazoxide
Authors: Raju Bharathi , Cryer Philip E.
Published: September 2004

To assess the mechanism, temporal patterns, and magnitudes of the metabolic responses to the ATP-dependent potassium channel agonist diazoxide, neuroendocrine and metabolic responses to intravenous diazoxide (saline, 1.0 and 2.0 mg/kg) and oral diazoxide (placebo, 4.0 and 6.0 mg/kg) were assessed in healthy young adults. Intravenous diazoxide produced rapid, but transient, decrements ( P = 0.0023) in plasma insulin (e.g., nadirs of 2.8 ± 0.5 and 1.8 ± 0.3 μU/ml compared with 7.0 ± 1.0 μU/ml after saline at 4.0–7.5 min) and C-peptide ( P = 0.0228) associated with dose-related increments in plasma glucose ( P = 0.0044) and serum nonesterified fatty acids ( P < 0.0001). After oral diazoxide, plasma insulin appeared to decline, as did C-peptide, again associated with dose-related increments in plasma glucose ( P < 0.0001) and serum nonesterified fatty acids ( P = 0.0141). Plasma glucagon, as well as cortisol and growth hormone, was not altered. Plasma epinephrine increased ( P = 0.0215) slightly only after intravenous diazoxide. There were dose-related increments in plasma norepinephrine ( P = 0.0038 and P = 0.0005, respectively), undoubtedly reflecting a compensatory sympathetic neural response to vasodilation produced by diazoxide, but these would not raise plasma glucose or serum nonesterified fatty acid levels. Thus selective suppression of insulin secretion, without stimulation of glucagon secretion, raised plasma glucose and serum nonesterified fatty acid concentrations. These findings define the temporal patterns and magnitudes of the metabolic responses to diazoxide and underscore the primacy of regulated insulin secretion in the physiological regulation of postabsorptive carbohydrate and lipid metabolism.
Time course of the hepatic adaptation to TPN: interaction with glycogen depletion
Authors: Chen Sheng-Song , Torres-Sanchez Carlos J. , Hosein Nadeen , Zhang Yiqun , Lacy D. Brooks , McGuinness Owen P.
Published: September 2004

In response to chronic (5 days) TPN, the liver becomes a major site of glucose disposal, removing ∼45% (4.5 mg·kg−1·min−1) of exogenous glucose. Moreover, ∼70% of glucose is not stored but released as lactate. We aimed to determine in chronically catheterized conscious dogs the time course of adaptation to TPN and the glycogen depletion impact on early time course. After an 18-h ( n = 5) fast, TPN was infused into the inferior vena cava for 8 ( n = 5) or 24 h ( n = 6). A third group, of 42-h-fasted animals ( n = 6), was infused with TPN for 8 h. TPN was infused at a rate designed to match the dog's calculated basal energy and nitrogen requirements. NHGU (−2.3 ± 0.1 to 2.2 ± 0.7 to 3.9 ± 0.6 vs. −1.7 ± 0.3 to 1.1 ± 0.5 to 2.9 ± 0.4 mg·kg−1·min−1, basal to 4 to 8 h, 18 vs. 42 h) and net hepatic lactate release (0.7 ± 0.3 to 0.6 ± 0.1 to 1.4 ± 0.2 vs. −0.6 ± 0.1 to 0.1 ± 0.1 to 0.8 ± 0.1 mg·kg−1·min−1, basal to 4 to 8 h) increased progressively. Net hepatic glycogen repletion and tracer determined that glycogen syntheses were similar. After 24 h of TPN, NHGU (5.4 ± 0.6 mg·kg−1·min−1) and net hepatic lactate release (2.6 ± 0.4 mg·kg−1·min−1) increased further. In summary, 1) most hepatic adaptation to TPN occurs within 24 h after initiation of TPN, and 2) prior glycogen depletion does not augment hepatic adaptation rate.