bluuebunny/crossref_metadata_2025_split huggingface dataset:
AccD6, a Member of the Fas II Locus, Is a Functional Carboxyltransferase Subunit of the Acyl-Coenzyme A Carboxylase in
Mycobacterium tuberculosis
Authors: Daniel Jaiyanth , Oh Tae-Jin , Lee Chang-Muk , Kolattukudy Pappachan E. 
Published: November 2006

ABSTRACT

The
Mycobacterium tuberculosis
acyl-coenzyme A (CoA) carboxylases provide the building blocks for de novo fatty acid biosynthesis by fatty acid synthase I (FAS I) and for the elongation of FAS I end products by the FAS II complex to produce meromycolic acids. The
M. tuberculosis
genome contains three biotin carboxylase subunits (AccA1 to -3) and six carboxyltransferase subunits (AccD1 to -6), with
accD6
located in a genetic locus that contains members of the FAS II complex. We found by quantitative real-time PCR analysis that the transcripts of
accA3
,
accD4
,
accD5
, and
accD6
are expressed at high levels during the exponential growth phases of
M. tuberculosis
in vitro. Microarray analysis of
M. tuberculosis
transcripts indicated that the transcripts for
accA3
,
accD4
,
accD5
,
accD6
, and
accE
were repressed during later growth stages. AccD4 and AccD5 have been previously studied, but there are no reports on the function of AccD6. We expressed AccA3 (α
3
) and AccD6 (β
6
) in
E. coli
and purified them by affinity chromatography. We report here that reconstitution of the α
3
-β
6
complex yielded an active acyl-CoA carboxylase. Kinetic characterization of this carboxylase showed that it preferentially carboxylated acetyl-CoA (1.1 nmol/mg/min) over propionyl-CoA (0.36 nmol/mg/min). The activity of the α
3
-β
6
complex was inhibited by the ε subunit. The α
3
-β
6
carboxylase was inhibited significantly by dimethyl itaconate, C75, haloxyfop, cerulenin, and 1,2-cyclohexanedione. Our results suggest that the β
6
subunit could play an important role in mycolic acid biosynthesis by providing malonyl-CoA to the FAS II complex.
The
Streptococcus mutans vicX
Gene Product Modulates
gtfB
/
C
Expression, Biofilm Formation, Genetic Competence, and Oxidative Stress Tolerance
Authors: Senadheera M. Dilani , Lee Andrew W. C. , Hung David C. I. , Spatafora Grace A. , Goodman Steven D. , Cvitkovitch Dennis G. 
Published: November 2006

ABSTRACT

Streptococcus mutans
is considered one of the primary etiologic agents of dental caries. Previously, we characterized the VicRK two-component signal transduction system, which regulates multiple virulence factors of
S. mutans
. In this study, we focused on the
vicX
gene of the
vicRKX
tricistronic operon. To characterize
vicX
, we constructed a nonpolar deletion mutation in the
vicX
coding region in
S. mutans
UA159. The growth kinetics of the mutant (designated SmuvicX) showed that the doubling time was longer and that there was considerable sensitivity to paraquat-induced oxidative stress. Supplementing a culture of the wild-type UA159 strain with paraquat significantly increased the expression of
vicX
(
P
< 0.05, as determined by analysis of variance [ANOVA]), confirming the role of this gene in oxidative stress tolerance in
S. mutans
. Examination of mutant biofilms revealed architecturally altered cell clusters that were seemingly denser than the wild-type cell clusters. Interestingly,
vicX
-deficient cells grown in a glucose-supplemented medium exhibited significantly increased glucosyltransferase B/C (
gtfB
/
C
) expression compared with the expression in the wild type (
P
< 0.05, as determined by ANOVA). Moreover, a sucrose-dependent adhesion assay performed using an
S. mutans
GS5-derived
vicX
null mutant demonstrated that the adhesiveness of this mutant was enhanced compared with that of the parent strain and isogenic mutants of the parent strain lacking
gtfB
and/or
gtfC
. Also, disruption of
vicX
reduced the genetic transformability of the mutant approximately 10-fold compared with that of the parent strain (
P
< 0.05, as determined by ANOVA). Collectively, these findings provide insight into important phenotypes controlled by the
vicX
gene product that can impact
S. mutans
pathogenicity.
Genetic Diversity among Botulinum Neurotoxin-Producing Clostridial Strains
Authors: Hill K. K. , Smith T. J. , Helma C. H. , Ticknor L. O. , Foley B. T. , Svensson R. T. , Brown J. L. , Johnson E. A. , Smith L. A. , Okinaka R. T. , Jackson P. J. , Marks J. D. 
Published: November 2006

ABSTRACT

Clostridium botulinum
is a taxonomic designation for many diverse anaerobic spore-forming rod-shaped bacteria that have the common property of producing botulinum neurotoxins (BoNTs). The BoNTs are exoneurotoxins that can cause severe paralysis and death in humans and other animal species. A collection of 174
C. botulinum
strains was examined by amplified fragment length polymorphism (AFLP) analysis and by sequencing of the 16S rRNA gene and BoNT genes to examine the genetic diversity within this species. This collection contained representatives of each of the seven different serotypes of botulinum neurotoxins (BoNT/A to BoNT/G). Analysis of the16S rRNA gene sequences confirmed previous identifications of at least four distinct genomic backgrounds (groups I to IV), each of which has independently acquired one or more BoNT genes through horizontal gene transfer. AFLP analysis provided higher resolution and could be used to further subdivide the four groups into subgroups. Sequencing of the BoNT genes from multiple strains of serotypes A, B, and E confirmed significant sequence variation within each serotype. Four distinct lineages within each of the BoNT A and B serotypes and five distinct lineages of serotype E strains were identified. The nucleotide sequences of the seven toxin genes of the serotypes were compared and showed various degrees of interrelatedness and recombination, as was previously noted for the nontoxic nonhemagglutinin gene, which is linked to the BoNT gene. These analyses contribute to the understanding of the evolution and phylogeny within this species and assist in the development of improved diagnostics and therapeutics for the treatment of botulism.
Biological Relevance of Colony Morphology and Phenotypic Switching by
Burkholderia pseudomallei
Authors: Chantratita Narisara , Wuthiekanun Vanaporn , Boonbumrung Khaemaporn , Tiyawisutsri Rachaneeporn , Vesaratchavest Mongkol , Limmathurotsakul Direk , Chierakul Wirongrong , Wongratanacheewin Surasakdi , Pukritiyakamee Sasithorn , White Nicholas J. , Day Nicholas P. J. , Peacock Sharon J. 
Published: November 2006

ABSTRACT

Melioidosis is a notoriously protracted illness and is difficult to cure. We hypothesize that the causative organism,
Burkholderia pseudomallei
, undergoes a process of adaptation involving altered expression of surface determinants which facilitates persistence in vivo and that this is reflected by changes in colony morphology. A colony morphotyping scheme and typing algorithm were developed using clinical
B. pseudomallei
isolates. Morphotypes were divided into seven types (denoted I to VII). Type I gave rise to other morphotypes (most commonly type II or III) by a process of switching in response to environmental stress, including starvation, iron limitation, and growth at 42°C. Switching was associated with complex shifts in phenotype, one of which (type I to type II) was associated with a marked increase in production of factors putatively associated with in vivo concealment. Isogenic types II and III, derived from type I, were examined using several experimental models. Switching between isogenic morphotypes occurred in a mouse model, where type II appeared to become adapted for persistence in a low-virulence state. Isogenic type II demonstrated a significant increase in intracellular replication fitness compared with parental type I after uptake by epithelial cells in vitro. Isogenic type III demonstrated a higher replication fitness following uptake by macrophages in vitro, which was associated with a switch to type II. Mixed
B. pseudomallei
morphologies were common in individual clinical specimens and were significantly more frequent in samples of blood, pus, and respiratory secretions than in urine and surface swabs. These findings have major implications for therapeutics and vaccine development.
Characterization of OpdH, a
Pseudomonas aeruginosa
Porin Involved in the Uptake of Tricarboxylates
Authors: Tamber Sandeep , Maier Elke , Benz Roland , Hancock Robert E. W. 
Published: November 2006

ABSTRACT

The
Pseudomonas aeruginosa
outer membrane is intrinsically impermeable to many classes of antibiotics, due in part to its relative lack of general uptake pathways. Instead, this organism relies on a large number of substrate-specific uptake porins. Included in this group are the 19 members of the OprD family, which are involved in the uptake of a diverse array of metabolites. One of these porins, OpdH, has been implicated in the uptake of
cis
-aconitate. Here we demonstrate that this porin may also enable
P. aeruginosa
to take up other tricarboxylates. Isocitrate and citrate strongly and specifically induced the
opdH
gene via a mechanism involving derepression by the putative two-component regulatory system PA0756-PA0757. Planar bilayer analysis of purified OpdH demonstrated that it was a channel-forming protein with a large single-channel conductance (230 pS in 1 M KCl; 10-fold higher than that of OprD); however, we were unable to demonstrate the presence of a tricarboxylate binding site within the channel. Thus, these data suggest that the requirement for OpdH for efficient growth on tricarboxylates was likely due to the specific expression of this large-channel porin under particular growth conditions.
Essential Bacterial Functions Encoded by Gene Pairs
Authors: Thomaides Helena B. , Davison Ella J. , Burston Lisa , Johnson Hazel , Brown David R. , Hunt Alison C. , Errington Jeffery , Czaplewski Lloyd 
Published: November 2006

ABSTRACTTo address the need for new antibacterials, a number of bacterial genomes have been systematically disrupted to identify essential genes. Such programs have focused on the disruption of single genes and may have missed functions encoded by gene pairs or multiple genes. In this work, we hypothesized that we could predict the identity of pairs of proteins within one organism that have the same function. We identified 135 putative protein pairs inBacillus subtilisand attempted to disrupt the genes forming these, singly and then in pairs. The single gene disruptions revealed new genes that could not be disrupted individually and other genes required for growth in minimal medium or for sporulation. The pairwise disruptions revealed seven pairs of proteins that are likely to have the same function, as the presence of one protein can compensate for the absence of the other. Six of these pairs are essential for bacterial viability and in four cases show a pattern of species conservation appropriate for potential antibacterial development. This work highlights the importance of combinatorial studies in understanding gene duplication and identifying functional redundancy.
Role of the β1 Subunit in the Function and Stability of the 20S Proteasome in the Hyperthermophilic ArchaeonPyrococcus furiosus
Authors: Madding Lara S. , Michel Joshua K. , Shockley Keith R. , Conners Shannon B. , Epting Kevin L. , Johnson Matthew R. , Kelly Robert M. 
Published: November 2006

ABSTRACTThe hyperthermophilic archaeonPyrococcus furiosusgenome encodes three proteasome component proteins: one α protein (PF1571) and two β proteins (β1-PF1404 and β2-PF0159), as well as an ATPase (PF0115), referred to as proteasome-activating nucleotidase. Transcriptional analysis of theP. furiosusdynamic heat shock response (shift from 90 to 105°C) showed that the β1 gene was up-regulated over twofold within 5 minutes, suggesting a specific role during thermal stress. Consistent with transcriptional data, two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that incorporation of the β1 protein relative to β2 into the 20S proteasome (core particle [CP]) increased with increasing temperature for both native and recombinant versions. For the recombinant enzyme, the β2/β1 ratio varied linearly with temperature from 3.8, when assembled at 80°C, to 0.9 at 105°C. The recombinant α+β1+β2 CP assembled at 105°C was more thermostable than either the α+β1+β2 version assembled at 90°C or the α+β2 version assembled at either 90°C or 105°C, based on melting temperature and the biocatalytic inactivation rate at 115°C. The recombinant CP assembled at 105°C was also found to have different catalytic rates and specificity for peptide hydrolysis, compared to the 90°C assembly (measured at 95°C). Combination of the α and β1 proteins neither yielded a large proteasome complex nor demonstrated any significant activity. These results indicate that the β1 subunit in theP. furiosus20S proteasome plays a thermostabilizing role and influences biocatalytic properties, suggesting that β subunit composition is a factor in archaeal proteasome function during thermal stress, when polypeptide turnover is essential to cell survival.
Characterization ofFrancisella tularensisOuter Membrane Proteins
Authors: Huntley Jason F. , Conley Patrick G. , Hagman Kayla E. , Norgard Michael V. 
Published: November 2006

ABSTRACTFrancisella tularensisis a gram-negative coccobacillus that is capable of causing severe, fatal disease in a number of mammalian species, including humans. Little is known about the proteins that are surface exposed on the outer membrane (OM) ofF. tularensis, yet identification of such proteins is potentially fundamental to understanding the initial infection process, intracellular survival, virulence, immune evasion and, ultimately, vaccine development. To facilitate the identification of putativeF. tularensisouter membrane proteins (OMPs), the genomes of both the type A strain (Schu S4) and type B strain (LVS) were subjected to six bioinformatic analyses for OMP signatures. Compilation of the bioinformatic predictions highlighted 16 putative OMPs, which were cloned and expressed for the generation of polyclonal antisera. Total membranes were extracted from both Schu S4 and LVS by spheroplasting and osmotic lysis, followed by sucrose density gradient centrifugation, which separated OMs from cytoplasmic (inner) membrane and other cellular compartments. Validation of OM separation and enrichment was confirmed by probing sucrose gradient fractions with antibodies to putative OMPs and inner membrane proteins.F. tularensisOMs typically migrated in sucrose gradients between densities of 1.17 and 1.20 g/ml, which differed from densities typically observed for other gram-negative bacteria (1.21 to 1.24 g/ml). Finally, the identities of immunogenic proteins were determined by separation on two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and mass spectrometric analysis. This is the first report of a direct method forF. tularensisOM isolation that, in combination with computational predictions, offers a more comprehensive approach for the characterization ofF. tularensisOMPs.
Physical and Functional Interactions between
Escherichia coli
MutY Glycosylase and Mismatch Repair Protein MutS
Authors: Bai Haibo , Lu A-Lien 
Published: November 2006

ABSTRACT

Escherichia coli
MutY and MutS increase replication fidelity by removing adenines that were misincorporated opposite 7,8-dihydro-8-oxo-deoxyguanines (8-oxoG), G, or C. MutY DNA glycosylase removes adenines from these mismatches through a short-patch base excision repair pathway and thus prevents G:C-to-T:A and A:T-to-G:C mutations. MutS binds to the mismatches and initiates the long-patch mismatch repair on daughter DNA strands. We have previously reported that the human MutY homolog (hMYH) physically and functionally interacts with the human MutS homolog, hMutSα (Y. Gu et al., J. Biol. Chem. 277:11135-11142, 2002). Here, we show that a similar relationship between MutY and MutS exists in
E. coli
. The interaction of MutY and MutS involves the Fe-S domain of MutY and the ATPase domain of MutS. MutS, in eightfold molar excess over MutY, can enhance the binding activity of MutY with an A/8-oxoG mismatch by eightfold. The MutY expression level and activity in
mutS
mutant strains are sixfold and twofold greater, respectively, than those for the wild-type cells. The frequency of A:T-to-G:C mutations is reduced by two- to threefold in a
mutS mutY
mutant compared to a
mutS
mutant. Our results suggest that MutY base excision repair and mismatch repair defend against the mutagenic effect of 8-oxoG lesions in a cooperative manner.
Catalase (KatA) and Alkyl Hydroperoxide Reductase (AhpC) Have Compensatory Roles in Peroxide Stress Resistance and Are Required for Survival, Persistence, and Nasal Colonization inStaphylococcus aureus
Authors: Cosgrove Kate , Coutts Graham , Jonsson Ing-Marie , Tarkowski Andrej , Kokai-Kun John F. , Mond James J. , Foster Simon J. 
Published: November 2006

ABSTRACTOxidative-stress resistance inStaphylococcus aureusis linked to metal ion homeostasis via several interacting regulators. In particular, PerR controls the expression of a regulon of genes, many of which encode antioxidants. Two PerR regulon members,ahpC(alkylhydroperoxide reductase) andkatA(catalase), show compensatory regulation, with independent and linked functions. AnahpCmutation leads to increased H2O2resistance due to greaterkatAexpression via relief of PerR repression. Moreover, AhpC provides residual catalase activity present in akatAmutant. Mutation of bothkatAandahpCleads to a severe growth defect under aerobic conditions in defined media (attributable to lack of catalase activity). This results in the inability to scavenge exogenous or endogenously produced H2O2, resulting in accumulation of H2O2in the medium. This leads to DNA damage, the likely cause of the growth defect. Surprisingly, thekatA ahpCmutant is not attenuated in two independent models of infection, which implies reduced oxygen availability during infection. In contrast, both AhpC and KatA are required for environmental persistence (desiccation) and nasal colonization. Thus, oxidative-stress resistance is an important factor in the ability ofS. aureusto persist in the hospital environment and so contribute to the spread of human disease.
In vitro and In vivo Activity of SKI-606, a Novel Src-Abl Inhibitor, against Imatinib-Resistant Bcr-Abl+ Neoplastic Cells
Authors: Puttini Miriam , Coluccia Addolorata Maria Luce , Boschelli Frank , Cleris Loredana , Marchesi Edoardo , Donella-Deana Arianna , Ahmed Shaheen , Redaelli Sara , Piazza Rocco , Magistroni Vera , Andreoni Federica , Scapozza Leonardo , Formelli Franca , Gambacorti-Passerini Carlo 
Published: November 2006

Abstract
Resistance to imatinib represents an important scientific and clinical issue in chronic myelogenous leukemia. In the present study, the effects of the novel inhibitor SKI-606 on various models of resistance to imatinib were studied. SKI-606 proved to be an active inhibitor of Bcr-Abl in several chronic myelogenous leukemia cell lines and transfectants, with IC50 values in the low nanomolar range, 1 to 2 logs lower than those obtained with imatinib. Cells expressing activated forms of KIT or platelet-derived growth factor receptor (PDGFR), two additional targets of imatinib, were unaffected by SKI-606, whereas activity was found against PIM2. SKI-606 retained activity in cells where resistance to imatinib was caused by BCR-ABL gene amplification and in three of four Bcr-Abl point mutants tested. In vivo experiments confirmed SKI-606 activity in models where resistance was not caused by mutations as well as in cells carrying the Y253F, E255K, and D276G mutations. Modeling considerations attribute the superior activity of SKI-606 to its ability to bind a conformation of Bcr-Abl different from imatinib. (Cancer Res 2006; 66(23): 11314-22)
Macrophages Regulate the Angiogenic Switch in a Mouse Model of Breast Cancer
Authors: Lin Elaine Y. , Li Jiu-Feng , Gnatovskiy Leoid , Deng Yan , Zhu Liyin , Grzesik Dustin A. , Qian Hong , Xue Xiao-nan , Pollard Jeffrey W. 
Published: November 2006

AbstractThe development of a tumor vasculature or access to the host vasculature is a crucial step for the survival and metastasis of malignant tumors. Although therapeutic strategies attempting to inhibit this step during tumor development are being developed, the biological regulation of this process is still largely unknown. Using a transgenic mouse susceptible to mammary cancer, PyMT mice, we have characterized the development of the vasculature in mammary tumors during their progression to malignancy. We show that the onset of the angiogenic switch, identified as the formation of a high-density vessel network, is closely associated with the transition to malignancy. More importantly, both the angiogenic switch and the progression to malignancy are regulated by infiltrated macrophages in the primary mammary tumors. Inhibition of the macrophage infiltration into the tumor delayed the angiogenic switch and malignant transition whereas genetic restoration of the macrophage population specifically in these tumors rescued the vessel phenotype. Furthermore, premature induction of macrophage infiltration into premalignant lesions promoted an early onset of the angiogenic switch independent of tumor progression. Taken together, this study shows that tumor-associated macrophages play a key role in promoting tumor angiogenesis, an essential step in the tumor progression to malignancy. (Cancer Res 2006; 66(23): 11238-46)
Marked Genomic Differences Characterize Primary and Secondary Glioblastoma Subtypes and Identify Two Distinct Molecular and Clinical Secondary Glioblastoma Entities
Authors: Maher Elizabeth A. , Brennan Cameron , Wen Patrick Y. , Durso Laura , Ligon Keith L. , Richardson Aaron , Khatry Deepak , Feng Bin , Sinha Raktim , Louis David N. , Quackenbush John , Black Peter McL. , Chin Lynda , DePinho Ronald A. 
Published: November 2006

Abstract
Glioblastoma is classified into two subtypes on the basis of clinical history: “primary glioblastoma” arising de novo without detectable antecedent disease and “secondary glioblastoma” evolving from a low-grade astrocytoma. Despite their distinctive clinical courses, they arrive at an indistinguishable clinical and pathologic end point highlighted by widespread invasion and resistance to therapy and, as such, are managed clinically as if they are one disease entity. Because the life history of a cancer cell is often reflected in the pattern of genomic alterations, we sought to determine whether primary and secondary glioblastomas evolve through similar or different molecular pathogenetic routes. Clinically annotated primary and secondary glioblastoma samples were subjected to high-resolution copy number analysis using oligonucleotide-based array comparative genomic hybridization. Unsupervised classification using genomic nonnegative matrix factorization methods identified three distinct genomic subclasses. Whereas one corresponded to clinically defined primary glioblastomas, the remaining two stratified secondary glioblastoma into two genetically distinct cohorts. Thus, this global genomic analysis showed wide-scale differences between primary and secondary glioblastomas that were previously unappreciated, and has shown for the first time that secondary glioblastoma is heterogeneous in its molecular pathogenesis. Consistent with these findings, analysis of regional recurrent copy number alterations revealed many more events unique to these subclasses than shared. The pathobiological significance of these shared and subtype-specific copy number alterations is reinforced by their frequent occurrence, resident genes with clear links to cancer, recurrence in diverse cancer types, and apparent association with clinical outcome. We conclude that glioblastoma is composed of at least three distinct molecular subtypes, including novel subgroups of secondary glioblastoma, which may benefit from different therapeutic strategies. (Cancer Res 2006; 66(23): 11502-13)