bluuebunny/crossref_metadata_2025_split huggingface dataset:
Possible Nonconductive Role of
Geobacter sulfurreducens
Pilus Nanowires in Biofilm Formation
Authors: Reguera Gemma , Pollina Rachael B. , Nicoll Julie S. , Lovley Derek R. 
Published: December 2006

ABSTRACT

Geobacter sulfurreducens
required expression of electrically conductive pili to form biofilms on Fe(III) oxide surfaces, but pili were also essential for biofilm development on plain glass when fumarate was the sole electron acceptor. Furthermore, pili were needed for cell aggregation in agglutination studies. These results suggest that the pili of
G. sulfurreducens
also have a structural role in biofilm formation.
Identification of Mycobacterial σ Factor Binding Sites by Chromatin Immunoprecipitation Assays
Authors: Rodrigue Sébastien , Brodeur Joëlle , Jacques Pierre-Étienne , Gervais Alain L. , Brzezinski Ryszard , Gaudreau Luc 
Published: December 2006

ABSTRACT

Mycobacterium tuberculosis
and
Mycobacterium bovis
are responsible for infections that cause a substantial amount of death, suffering, and loss around the world. Still, relatively little is known about the mechanisms of gene expression in these bacteria. Here, we used genome-wide location assays to identify direct target genes for mycobacterial σ factors. Chromatin immunoprecipitation assays were performed with
M. bovis
BCG for Myc-tagged proteins expressed using an anhydrotetracycline-inducible promoter, and enriched DNA fragments were hybridized to a microarray representing intergenic regions from the
M. tuberculosis
H37Rv genome. Several putative target genes were validated by quantitative PCR. The corresponding transcriptional start sites were identified for σ
F
, σ
C
, and σ
K
, and consensus promoter sequences are proposed. Our conclusions were supported by the results of in vitro transcription assays. We also examined the role of each holoenzyme in the expression of σ factor genes. Our results revealed that many σ factors are expressed from autoregulated promoters.
Localization of the Naturally Occurring Plasmid ColE1 at the Cell Pole
Authors: Yao Shiyin , Helinski Donald R. , Toukdarian Aresa 
Published: December 2006

ABSTRACT

The naturally occurring plasmid ColE1 was found to localize as a cluster in one or both of the cell poles of
Escherichia coli
. In addition to the polar localization of ColE1 in most cells, movement of the plasmid to the midcell position was observed in time-lapse studies. ColE1 could be displaced from its polar location by the p15A replicon, pBAD33, but not by plasmid RK2. The displacement of ColE1 by pBAD33 resulted in an almost random positioning of ColE1 foci in the cell and also in a loss of segregational stability, as evidenced by the large number of cells carrying pBAD33 with no visible ColE1 focus and as confirmed by ColE1 stability studies. The addition of the active partitioning systems of the F plasmid (
sopABC
) or RK2 (O
B1
incC korB
) resulted in movement of the ColE1 replicon from the cell pole to within the nucleoid region. This repositioning did not result in destabilization but did result in an increase in the number of plasmid foci, most likely due to partial declustering. These results are consistent with the importance of
par
regions to the localization of plasmids to specific regions of the cell and demonstrate both localization and dynamic movement for a naturally occurring plasmid that does not encode a replication initiation protein or a partitioning system that is required for plasmid stability.
Transcription from the P3Promoter of theBacillus subtilis spxGene Is Induced in Response to Disulfide Stress
Authors: Leelakriangsak Montira , Zuber Peter 
Published: December 2006

ABSTRACTThespxgene ofBacillus subtilisencodes a global regulator that controls transcription initiation in response to oxidative stress by interaction with RNA polymerase (RNAP). It is located in a dicistronic operon with theyjbCgene. Thespxgene DNA complements anspxnull mutation with respect to disulfide stress resistance, suggesting thatspxis transcribed from a promoter located in the intergenic region ofyjbCandspx. Transcription of theyjbC-spxoperon has been reported to be driven by four promoters, three (P1, P2, and PB) residing upstream ofyjbCand one (PM) located in the intergenic region betweenyjbCandspx. Primer extension analysis uncovered a second intergenic promoter, P3, from which transcription is elevated in cells treated with the thiol-specific oxidant diamide. P3is utilized by the σAform of RNA polymerase in vitro without the involvement of a transcriptional activator. Transcriptional induction from P3did not require an Spx-RNAP interaction and was observed in a deletion mutant lacking DNA upstream of position −40 of the P3promoter start site. Deletion mutants with endpoints 3′ to the P3transcriptional start site (positions +5, +15, and +30) showed near-constitutive transcription at the induced level, indicating the presence of a negative control element downstream of the P3promoter sequence. Point mutations characterized bybgaBfusion expression and primer extension analyses uncovered evidence for a secondcis-acting site in the P3promoter sequence itself. The data indicate thatspxtranscription is under negative transcriptional control that is reversed when disulfide stress is encountered.
Dual Negative Control ofspxTranscription Initiation from the P3Promoter by Repressors PerR and YodB inBacillus subtilis
Authors: Leelakriangsak Montira , Kobayashi Kazuo , Zuber Peter 
Published: December 2006

ABSTRACTThespxgene encodes an RNA polymerase-binding protein that exerts negative and positive transcriptional control in response to oxidative stress inBacillus subtilis. It resides in theyjbC-spxoperon and is transcribed from at least five promoters located in theyjbCregulatory region or in theyjbC-spxintergenic region. Induction ofspxtranscription in response to treatment with the thiol-specific oxidant diamide is the result of transcription initiation at the P3promoter located upstream of thespxcoding sequence. Previous studies conducted elsewhere and analyses of transcription factor mutants using transformation array technology have uncovered two transcriptional repressors, PerR and YodB, that target thecis-acting negative control elements of the P3promoter. Expression of anspx-bgaBfusion carrying the P3promoter is elevated in ayodBorperRmutant, and an additive increase in expression was observed in ayodB perRdouble mutant. Primer extension analysis ofspxRNA shows the same additive increase in P3transcript levels inyodB perRmutant cells. Purified YodB and PerR repressspxtranscription in vitro when wild-typespxP3promoter DNA was used as a template. Point mutations at positions within the P3promoter relieved YodB-dependent repression, while a point mutation at position +24 reduced PerR repression. DNase I footprinting analysis showed that YodB protects a region that includes the P3−10 and −35 regions, while PerR binds to a region downstream of the P3transcriptional start site. The binding of both repressors is impaired by the treatment of footprinting reactions with diamide or hydrogen peroxide. The study has uncovered a mechanism of dual negative control that relates to the oxidative stress response of gram-positive bacteria.
Only One of Five
groEL
Genes Is Required for Viability and Successful Symbiosis in
Sinorhizobium meliloti
Authors: Bittner Alycia N. , Foltz Amanda , Oke Valerie 
Published: December 2006

ABSTRACT

Many bacterial species contain multiple copies of the genes that encode the chaperone GroEL and its cochaperone, GroES, including all of the fully sequenced root-nodulating bacteria that interact symbiotically with legumes to generate fixed nitrogen. In particular, in
Sinorhizobium meliloti
there are four
groESL
operons and one
groEL
gene. To uncover functional redundancies of these genes during growth and symbiosis, we attempted to construct strains containing all combinations of
groEL
mutations. Although a double
groEL1 groEL2
mutant cannot be constructed, we demonstrate that the quadruple
groEL1 groESL3 groEL4 groESL5
and
groEL2 groESL3 groEL4 groESL5
mutants are viable. Therefore, like
E. coli
and other species,
S. meliloti
requires only one
groEL
gene for viability, and either
groEL1
or
groEL2
will suffice. The
groEL1 groESL5
double mutant is more severely affected for growth at both 30°C and 40°C than the single mutants, suggesting overlapping functions in stress response. During symbiosis the quadruple
groEL2 groESL3 groEL4 groESL5
mutant acts like the wild type, but the quadruple
groEL1 groESL3 groEL4 groESL5
mutant acts like the
groEL1
single mutant, which cannot fully induce
nod
gene expression and forms ineffective nodules. Therefore, the only
groEL
gene required for symbiosis is
groEL1
. However, we show that the other
groE
genes are expressed in the nodule at lower levels, suggesting minor roles during symbiosis. Combining our data with other data, we conclude that
groESL1
encodes the housekeeping GroEL/GroES chaperone and that
groESL5
is specialized for stress response.
Role of SpoVA Proteins in Release of Dipicolinic Acid during Germination of
Bacillus subtilis
Spores Triggered by Dodecylamine or Lysozyme
Authors: Vepachedu Venkata Ramana , Setlow Peter 
Published: December 2006

ABSTRACT

The release of dipicolinic acid (DPA) during the germination of
Bacillus subtilis
spores by the cationic surfactant dodecylamine exhibited a pH optimum of ∼9 and a temperature optimum of 60°C. DPA release during dodecylamine germination of
B. subtilis
spores with fourfold-elevated levels of the SpoVA proteins that have been suggested to be involved in the release of DPA during nutrient germination was about fourfold faster than DPA release during dodecylamine germination of wild-type spores and was inhibited by HgCl
2
. Spores carrying temperature-sensitive mutants in the
spoVA
operon were also temperature sensitive in DPA release during dodecylamine germination as well as in lysozyme germination of decoated spores. In addition to DPA, dodecylamine triggered the release of amounts of Ca
2+
almost equivalent to those of DPA, and at least one other abundant spore small molecule, glutamic acid, was released in parallel with Ca
2+
and DPA. These data indicate that (i) dodecylamine triggers spore germination by opening a channel in the inner membrane for Ca
2+
-DPA and other small molecules, (ii) this channel is composed at least in part of proteins, and (iii) SpoVA proteins are involved in the release of Ca
2+
-DPA and other small molecules during spore germination, perhaps by being a part of a channel in the spore's inner membrane.
Identification of Genes Involved in Swarming Motility Using aPseudomonas aeruginosaPAO1 Mini-Tn5-luxMutant Library
Authors: Overhage Joerg , Lewenza Shawn , Marr Alexandra K. , Hancock Robert E. W. 
Published: December 2006

ABSTRACTDuring a screening of a mini-Tn5-luxCDABEtransposon mutant library ofPseudomonas aeruginosaPAO1 for alterations in swarming motility, 36 mutants were identified with Tn5insertions in genes for the synthesis or function of flagellin and type IV pilus, in genes for the Xcp-related type II secretion system, and in regulatory, metabolic, chemosensory, and hypothetical genes with unknown functions. These mutants were differentially affected in swimming and twitching motility but in most cases had only a minor additional motility defect. Our data provide evidence that swarming is a more complex type of motility, since it is influenced by a large number of different genes inP. aeruginosa. Conversely, many of the swarming-negative mutants also showed an impairment in biofilm formation, indicating a strong relationship between these types of growth states.
Evidence that RpoS (σ
S
) in
Borrelia burgdorferi
Is Controlled Directly by RpoN (σ
54
/σ
N
)
Authors: Smith Alexandra H. , Blevins Jon S. , Bachlani Gulnaz N. , Yang Xiaofeng F. , Norgard Michael V. 
Published: December 2006

ABSTRACT

The alternative sigma factor (RpoN-RpoS) pathway controls the expression of key virulence factors in
Borrelia burgdorferi
. However, evidence to support whether RpoN controls
rpoS
directly or, perhaps, indirectly via a transactivator has been lacking. Herein we provide biochemical and genetic evidence that RpoN directly controls
rpoS
in
B. burgdorferi
.
Biochemical and Genetic Analysis of the γ-Resorcylate (2,6-Dihydroxybenzoate) Catabolic Pathway in
Rhizobium
sp. Strain MTP-10005: Identification and Functional Analysis of Its Gene Cluster
Authors: Yoshida Masahiro , Oikawa Tadao , Obata Hitoshi , Abe Katsumasa , Mihara Hisaaki , Esaki Nobuyoshi 
Published: December 2006

ABSTRACT

We identified a gene cluster that is involved in the γ-resorcylate (2,6-dihydroxybenzoate) catabolism of the aerobic bacterium
Rhizobium
sp. strain MTP-10005. The cluster consists of the
graRDAFCBEK
genes, and
graA
,
graB
,
graC
, and
graD
were heterologously expressed in
Escherichia coli
. Enzymological studies showed that
graD
,
graA
,
graC
, and
graB
encode the reductase (GraD) and oxygenase (GraA) components of a resorcinol hydroxylase (EC 1.14.13.x), a maleylacetate reductase (GraC) (EC 1.3.1.32), and a hydroxyquinol 1,2-dioxygenase (GraB) (EC 1.13.11.37). Bioinformatic analyses suggested that
graE
,
graR
, and
graK
encode a protein with an unknown function (GraE), a MarR-type transcriptional regulator (GraR), and a benzoate transporter (GraK). Quantitative reverse transcription-PCR of
graF
, which encodes γ-resorcylate decarboxylase, revealed that the maximum relative mRNA expression level ([5.93 ± 0.82] × 10
−4
) of
graF
was detected in the total RNA of the cells after one hour of cultivation when γ-resorcylate was used as the sole carbon source. Reverse transcription-PCR of
graDAFCBE
showed that these genes are transcribed as a single mRNA and that the transcription of the gene cluster is induced by γ-resorcylate. These results suggested that the
graDAFCBE
genes are responsible as an operon for the growth of
Rhizobium
sp. strain MTP-10005 on γ-resorcylate and are probably regulated by GraR at the transcriptional level. This is the first report of the γ-resorcylate catabolic pathway in an aerobic bacterium.
HER2 evaluation using the novel rabbit monoclonal antibody SP3 and CISH in tissue microarrays of invasive breast carcinomas
Authors: Ricardo Sara Alexandra Vinhas , Milanezi Fernanda , Carvalho Sílvia Teresa , Leitão Dina Raquel Aguilera , Schmitt Fernando Carlos Lander 
Published: December 2006

Background:Laboratory methods for HER2 assessment currently include immunohistochemical (IHC) methods (measuring protein overexpression) and fluorescence in situ hybridisation (FISH) (measuring gene amplification). The measure of HER2 protein by IHC is usually assessed by the mouse monoclonal antibody CB11, and polyclonal antibodies (Herceptest) directed against the internal portion of the receptor. Recently, chromogenic in situ hybridisation (CISH), in which HER2 is detected by a peroxidase reaction and the gene amplification can be determined by regular bright-field microscopy, has emerged as an alternative to FISH.Aims:To evaluate the status of HER2 in tissue microarrays (TMAs) of invasive breast cancer using the novel rabbit monoclonal antibody SP3 directed against the external portion of HER2, and correlate the results with CB11 and CISH.Methods:IHC was performed with two antibodies (CB11 and SP3) and CISH for HER2 in 10 TMA blocks with 190 formalin-fixed paraffin-embedded cases of invasive breast carcinomas.Results:The correlation between SP3 and CB11 was significant (p<0.001) with an agreement rate of 86.9%. When the staining pattern of the two antibodies was compared, the majority of SP3 immunostainings were assessed more easily, with a strong complete membrane staining pattern without non-specific cytoplasmic staining. There was a good correlation between SP3 and CISH (p<0.001). 23/24 SP3 3+ cases showed gene amplification, 97.3% of the cases without gene amplification were SP3 negative, and 6/7 SP3 2+ were amplified.Conclusion:The high level of agreement between SP3, a monoclonal antibody that recognises the extracellular domain of the HER2 receptor, and CB11 and CISH, shows that this novel antibody is a reliable candidate to evaluate the expression of HER2 in breast cancer.
Expression of EphA2 and EphrinA-1 in vulvar carcinomas and its relation to prognosis
Authors: Holm R , Knopp S , Suo Z , Tropè C , Nesland J M 
Published: December 2006

Aims:To examine the expression of EphA2 and EphrinA-1 in vulvar squamous cell carcinomas and investigate their prognostic relevance.Methods:Tumours from 224 patients with vulvar squamous cell carcinomas were investigated for expression of EphA2 and EphrinA-1 using single and double immunostaining methods.Results:High expression (strong/moderate staining intensity) of EphA2 and EphrinA-1 was observed in 114 (51%) and 126 (56%) vulvar carcinomas, respectively. In the three cases tested using the double immunostaining method, colocalisation of EphA2 and EphrinA-1 proteins was identified in the same neoplastic cells. High EphA2 expression was significantly correlated to high expression of EphrinA-1 (p<0.01) and cyclin A (p<0.01), large tumour size (p = 0.03), deep invasion (p<0.01) and higher FIGO stage (p = 0.05). A correlation between high EphrinA-1 expression and high levels of cyclin A (p<0.01) and p21 (p<0.01), deep invasion (p<0.01) and higher FIGO stage (p = 0.01) was also seen. In univariate analysis, high expression of EphrinA-1 was associated with poor survival (p = 0.03). However, in the multivariate analysis neither EphrinA-1 nor EphA2 were significantly correlated to survival.Conclusions:EphA2 and EphrinA-1 were overexpressed in 51% and 56% of the vulvar squamous cell carcinomas, respectively, and high levels of EphA2 and EphrinA-1 proteins were associated with deep tumour invasion and high FIGO stage. However, EphA2 and EphrinA-1 were not independently associated with clinical outcome in vulvar carcinomas.
Urine cytology screening for polyoma virus infection following renal transplantation: the Oxford experience
Authors: Thamboo Thomas P , Jeffery Katie J M , Friend Peter J , Turner Gareth D H , Roberts Ian S D 
Published: December 2006

Objective:To review the first year of a monthly urine cytology screening service, introduced to identify renal transplant patients at risk of polyoma virus nephropathy (PVN), at an early, potentially treatable, stage.Methods and results:Monthly urine samples (n = 392) were received from 97/108 transplant recipients in 2005. Of 56 patients with follow-up >6 months, 20% and 9% had significant (>10 decoy cells/cytospin) and non-significant positive cytology, respectively. The first positive urine samples occurred most commonly in the second and third month post-transplantation and patients with significantly positive samples had higher 3-month and 6-month serum creatinine levels than patients with negative urine cytology (p<0.01). Four patients with positive urine cytology had a subsequent positive plasma BK virus PCR; 3/97 patients had biopsy-proven PVN, all in the third month, 1–6 weeks after first positive urine samples.Conclusions:Significant PV viruria is common following renal transplantation with onset usually within the first 3 months. Viruria is associated with worse graft function at 3 and 6 months. The time between urine positivity and clinical PVN is short. More frequent early urine screening would be required to achieve clinical benefit.
MYCamplification in breast cancer: a chromogenic in situ hybridisation study
Authors: Rodriguez-Pinilla S Maria , Jones Robin L , Lambros Maryou B K , Arriola Edurne , Savage Kay , James Michelle , Pinder Sarah E , Reis-Filho Jorge S 
Published: December 2006

Aims:To analyse the correlation betweenMYCamplification and various clinicopathological features and outcome in a cohort of 245 patients with invasive breast carcinoma treated with surgery followed by anthracycline-based chemotherapy. Given the high prevalence ofMYCamplification in tumours ofBRCA1mutation carriers and the similarities between these and sporadic “basal-like” carcinomas, the prevalence ofMYCamplification in “basal-like” breast carcinomas was investigated.Methods:MYCgene copy number was assessed on tissue microarrays containing duplicate cores of 245 invasive breast carcinomas by means of chromogenic in situ hybridisation using SpotLight C-MYC amplification probe and chromosome 8 centromeric probe (CEP8). Signals were evaluated at 400× magnification; 30 morphologically unequivocal neoplastic cells in each core were counted for the presence of the gene and CEP8 probes.Results:Amplification was defined as aMYC:CEP8 ratio >2. Signals for bothMYCand CEP8 were assessable in 196/245 (80%) tumours.MYCamplification was found in 19/196 cases (9.7%) and was not associated with tumour size, histological grade, positivity for oestrogen receptor, progesterone receptor, HER2, epidermal growth factor, cytokeratins 14, 5/6 and 17, MIB1 or p53. Only 4% of basal-like carcinomas showedMYCamplification, compared to 8.75% and 10.7% of luminal and HER2 tumours respectively. On univariate analysis,MYCamplification displayed a significant association with shorter metastasis-free and overall survival and proved to be an independent prognostic factor on multivariate survival analysis.Conclusion:MYCamplification is not associated with “basal-like” phenotype and proved to be an independent prognostic factor for breast cancer patients treated with anthracycline-based chemotherapy.
Endothelin-1 expression correlates with atypical histological features in mammary phyllodes tumours
Authors: Tse Gary M , Chaiwun Benjaporn , Lau Kin-Mang , Scolyer Richard , Lee C Soon , Karim Rooshdiya Z , Putti Thomas C , Law Bonita K , Lui Philip C , Tan Puay Hoon 
Published: December 2006

Background and Aims:Endothelin-1 expression is increased in infiltrating duct carcinoma and is associated with larger tumour size, higher histological grade and lymphovascular permeation. This has not been evaluated in phyllodes tumours, which are uncommon fibroepithelial lesions with potential for local recurrences or distant metastasis. While the grading of phyllodes tumours depends on a combination of histological parameters, prediction of their behaviour remains difficult.Method:A large series of 461 phyllodes tumours (291 benign, 115 borderline malignant and 55 frankly malignant) were evaluated for endothelin-1 expression in both the epithelial cells and stromal cells by immunohistochemistry; results were correlated with the tumour grade.Results:For benign phyllodes tumours, the epithelial staining of endothelin was negative, weak, moderate and strong in 6%, 26%, 15% and 53% of cases respectively; results were 4%, 18%, 19% and 59% respectively for borderline and 6%, 18%, 6% and 70% respectively for frankly malignant tumours. For the stromal staining, the negative, weak, moderate and strong staining was 32%, 19%, 18% and 31% respectively for benign phyllodes, 24%, 13%, 10% and 53% respectively for borderline and 8%, 16%, 17% and 59% respectively for frankly malignant tumours. There was correlation between epithelial and stromal staining, and the stromal staining correlated with histological features of stromal cellularity, stromal cell nuclear pleomorphism, margin status and stromal overgrowth.Conclusion:These observations suggest a close relationship between the epithelial and stromal elements in phyllodes tumours; endothelin may play a significant role in the malignant progression of phyllodes tumours.