bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional Characterization of Clostridium difficile Spore Coat Proteins
Authors: Permpoonpattana Patima , Phetcharaburanin Jutarop , Mikelsone Anna , Dembek Marcin , Tan Sisareuth , Brisson Marie-Clémence , La Ragione Roberto , Brisson Alain R. , Fairweather Neil , Hong Huynh A. , Cutting Simon M. 
Published: January 2013

ABSTRACT

Spores of
Clostridium difficile
play a key role in the dissemination of this important human pathogen, and until recently little has been known of their functional characteristics. Genes encoding six spore coat proteins (
cotA
,
cotB
,
cotCB
,
cotD
,
cotE
, and
sodA
) were disrupted by ClosTron insertional mutagenesis. Mutation of one gene,
cotA
, presented a major structural defect in spore assembly, with a clear misassembly of the outermost layers of the spore coat. The CotA protein is most probably subject to posttranslational modification and could play a key role in stabilizing the spore coat. Surprisingly, mutation of the other spore coat genes did not affect the integrity of the spore, although for the
cotD
,
cotE
, and
sodA
mutants, enzyme activity was reduced or abolished. This could imply that these enzymatic proteins are located in the exosporium or alternatively that they are structurally redundant. Of the spore coat proteins predicted to carry enzymatic activity, three were confirmed to be enzymes using both
in vivo
and
in vitro
methods, the latter using recombinant expressed proteins. These were a manganese catalase, encoded by
cotD
, a superoxide dismutase (SOD), encoded by
sodA
, and a bifunctional enzyme with peroxiredoxin and chitinase activity, encoded by
cotE
. These enzymes being exposed on the spore surface would play a role in coat polymerization and detoxification of H
2
O
2
. Two additional proteins, CotF (a tyrosine-rich protein and potential substrate for SodA) and CotG (a putative manganese catalase) were shown to be located at the spore surface.
Effects of H
2
and Formate on Growth Yield and Regulation of Methanogenesis in Methanococcus maripaludis
Authors: Costa Kyle C. , Yoon Sung Ho , Pan Min , Burn June A. , Baliga Nitin S. , Leigh John A. 
Published: January 2013

ABSTRACT

Hydrogenotrophic methanogenic
Archaea
are defined by an H
2
requirement for growth. Despite this requirement, many hydrogenotrophs are also capable of growth with formate as an electron donor for methanogenesis. While certain responses of these organisms to hydrogen availability have been characterized, responses to formate starvation have not been reported. Here we report that during continuous culture of
Methanococcus maripaludis
under defined nutrient conditions, growth yields relative to methane production decreased markedly with either H
2
excess or formate excess. Analysis of the growth yields of several mutants suggests that this phenomenon occurs independently of the storage of intracellular carbon or a transcriptional response to methanogenesis. Using microarray analysis, we found that the expression of genes encoding coenzyme F
420
-dependent steps of methanogenesis, including one of two formate dehydrogenases, increased with H
2
starvation but with formate occurred at high levels regardless of limitation or excess. One gene, encoding H
2
-dependent methylene-tetrahydromethanopterin dehydrogenase, decreased in expression with either H
2
limitation or formate limitation. Expression of genes for the second formate dehydrogenase, molybdenum-dependent formylmethanofuran dehydrogenase, and molybdenum transport increased specifically with formate limitation. Of the two formate dehydrogenases, only the first could support growth on formate in batch culture where formate was in excess.
Novel Modulators Controlling Entry into Sporulation in Bacillus subtilis
Authors: Garti-Levi Sharon , Eswara Ashlee , Smith Yoav , Fujita Masaya , Ben-Yehuda Sigal 
Published: January 2013

ABSTRACT

Upon nutrient deprivation,
Bacillus subtilis
initiates the developmental process of sporulation by integrating environmental and extracellular signals. These signals are channeled into a phosphorelay ultimately activating the key transcriptional regulator of sporulation, Spo0A. Subsequently, phosphorylated Spo0A regulates the expression of genes required for sporulation to initiate. Here we identified a group of genes whose transcription levels are controlled by Spo0A during exponential growth. Among them, three upregulated genes, termed
sivA
,
sivB
(
bslA
), and
sivC
, encode factors found to inhibit Spo0A activation. We furthermore show that the Siv factors operate by reducing the activity of histidine kinases located at the top of the sporulation phosphorelay, thereby decreasing Spo0A phosphorylation. Thus, we demonstrate the existence of modulators, positively controlled by Spo0A, which inhibit inappropriate entry into the costly process of sporulation, when conditions are favorable for exponential growth.
Coordination of FocA and Pyruvate Formate-Lyase Synthesis in Escherichia coli Demonstrates Preferential Translocation of Formate over Other Mixed-Acid Fermentation Products
Authors: Beyer Lydia , Doberenz Claudia , Falke Dörte , Hunger Doreen , Suppmann Bernhard , Sawers R. Gary 
Published: January 2013

ABSTRACT

Enterobacteria such as
Escherichia coli
generate formate, lactate, acetate, and succinate as major acidic fermentation products. Accumulation of these products in the cytoplasm would lead to uncoupling of the membrane potential, and therefore they must be either metabolized rapidly or exported from the cell.
E. coli
has three membrane-localized formate dehydrogenases (FDHs) that oxidize formate. Two of these have their respective active sites facing the periplasm, and the other is in the cytoplasm. The bidirectional FocA channel translocates formate across the membrane delivering substrate to these FDHs. FocA synthesis is tightly coupled to synthesis of pyruvate formate-lyase (PflB), which generates formate. In this study, we analyze the consequences on the fermentation product spectrum of altering FocA levels, uncoupling FocA from PflB synthesis or blocking formate metabolism. Changing the
focA
translation initiation codon from GUG to AUG resulted in a 20-fold increase in FocA during fermentation and an ∼3-fold increase in PflB. Nevertheless, the fermentation product spectrum throughout the growth phase remained similar to that of the wild type. Formate, acetate, and succinate were exported, but only formate was reimported by these cells. Lactate accumulated in the growth medium only in mutants lacking FocA, despite retaining active PflB, or when formate could not be metabolized intracellularly. Together, these results indicate that FocA has a strong preference for formate as a substrate
in vivo
and not other acidic fermentation products. The tight coupling between FocA and PflB synthesis ensures adequate substrate delivery to the appropriate FDH.
Exoprotein Production Correlates with Morphotype Changes of Nonmotile Shewanella oneidensis Mutants
Authors: Shi Miaomiao , Wu Lin , Xia Yu , Chen Haijiang , Luo Qixia , Sun Linlin , Gao Haichun 
Published: January 2013

ABSTRACT

We report a previously undescribed mechanism for the rugose morphotype in
Shewanella oneidensis
, a research model for investigating redox transformations of environmental contaminants. Bacteria may form smooth or rugose colonies on agar plates. In general, conversion from the smooth to rugose colony morphotype is attributed to increased production of exopolysaccharide (EPS). In this work, we discovered that aflagellate
S. oneidensis
mutants grew into rugose colonies, whereas those with nonfunctional flagellar filaments remained smooth. EPS production was not altered in either case, but mutants with the rugose morphotype showed significantly reduced exoprotein secretion. The idea that exoproteins at a reduced level correlate with rugosity gained support from smooth suppressor strains of an aflagellate rugose
fliD
(encoding the capping protein) mutant, which restored the exoprotein level to the levels of the wild-type and mutant strains with a smooth morphotype. Further analyses revealed that SO1072 (a putative GlcNAc-binding protein) was one of the highly upregulated exoproteins in these suppressor strains. Most intriguingly, this study identified a compensatory mechanism of SO1072 to flagellins possibly mediated by bis-(3′-5′)-cyclic dimeric GMP.
The Cpx Stress Response Confers Resistance to Some, but Not All, Bactericidal Antibiotics
Authors: Mahoney Tara F. , Silhavy Thomas J. 
Published: January 2013

ABSTRACT

It has recently been suggested that bactericidal antibiotics, including aminoglycoside antibiotics (AGAs), and toxic small molecules, such as hydroxyurea (HU), kill bacteria the same way, namely, by generating reactive oxygen species (ROS) via a process requiring activation of the Cpx stress response. We suggest an opposite, protective role for Cpx. We have confirmed the initial finding that
cpxA
null mutations confer resistance to HU. However, the two-component sensor CpxA is both a kinase and a phosphatase, and previous work from our lab has shown that removing CpxA can activate the stress response owing to buildup of the phosphorylated response regulator (CpxR∼P) that occurs in the absence of the phosphatase activity. We show that a dominant
cpxA
* mutation that constitutively activates the Cpx stress response confers a high level of resistance to both HU and AGAs in a CpxR-dependent manner. In contrast, inactivating the CpxR response regulator by mutating the phosphorylation site (D51A) or the putative DNA-binding motif (M199A) does not increase resistance to HU or AGAs. Taken together, these results demonstrate that activation of the Cpx stress response can protect cells from HU and AGAs. However, the Cpx response does not increase resistance to all classes of bactericidal antibiotics, as the
cpxA
* mutants are not significantly more resistant to fluoroquinolones or β-lactams than wild-type cells. Thus, it seems unlikely that all bactericidal antibiotics kill by the same mechanism.
Topology and Accessibility of Germination Proteins in the Bacillus subtilis Spore Inner Membrane
Authors: Korza George , Setlow Peter 
Published: January 2013

ABSTRACT

Access to a membrane-impermeant biotinylation reagent as well as protease sensitivity was used to determine germination proteins' topology in the inner membrane (IM) of decoated dormant spores and intact germinated
Bacillus subtilis
spores. The proteins examined were four nutrient germinant receptor (GR) subunits, the GerD protein, essential for normal GR-dependent spore germination, the SpoVAD protein, essential for dipicolinic acid movement across the IM, the SleB cortex-lytic enzyme, and the YpeB protein, essential for SleB assembly in spores, as well as green fluorescent protein (GFP) in the spore core. GerD and SpoVAD as well as GFP in the spore were not biotinylated in decoated dormant spores. However, GR subunits, SleB, and YpeB were biotinylated 4 to 36% in decoated dormant spores, although these levels were not increased by higher biotinylation reagent concentrations or longer reaction times. In contrast, the germination proteins were largely biotinylated in germinated spores, although GFP was not. All of the germination proteins in the germinated spore's IM, but not spore core GFP, were largely sensitive to an exogenous protease. These results, coupled with predicted or experimentally determined structural data, indicate that (i) these germination proteins are at least partially and in some cases completely on the outer surface of the spore's IM and (ii) there is significant reorganization of these germination proteins' structure or environment in the IM during spore germination.
Evidence for the effectiveness of electrophysical modalities for treatment of medial and lateral epicondylitis: a systematic review
Authors: Dingemanse Rudi , Randsdorp Manon , Koes Bart W , Huisstede Bionka M A 
Published: January 2013

BackgroundSeveral treatments are available to treat epicondylitis. Among these are instrumental electrophysical modalities, ranging from ultrasound, extracorporeal shock wave therapy (ESWT), transcutaneous electrical nerve stimulation (TENS) to laser therapy, commonly used to treat epicondylitis.ObjectivesTo present an evidence-based overview of the effectiveness of electrophysical modality treatments for both medial and lateral epicondylitis (LE).MethodsSearches in PubMed, EMBASE, CINAHL and Pedro were performed to identify relevant randomised clinical trials (RCTs) and systematic reviews. Two reviewers independently extracted data and assessed the methodological quality. A best-evidence synthesis was used to summarise the results.ResultsA total of 2 reviews and 20 RCTs were included, all of which concerned LE. Different electrophysical regimes were evaluated: ultrasound, laser, electrotherapy, ESWT, TENS and pulsed electromagnetic field therapy. Moderate evidence was found for the effectiveness of ultrasound versus placebo on mid-term follow-up. Ultrasound plus friction massage showed moderate evidence of effectiveness versus laser therapy on short-term follow-up. On the contrary, moderate evidence was found in favour of laser therapy over plyometric exercises on short-term follow-up. For all other modalities only limited/conflicting evidence for effectiveness or evidence of no difference in effect was found.ConclusionsPotential effectiveness of ultrasound and laser for the management of LE was found. To draw more definite conclusions high-quality RCTs examining different intensities are needed as well as studies focusing on long-term follow-up results.
A Multifunctional Chimeric Chaperone Serves as a Novel Immune Modulator Inducing Therapeutic Antitumor Immunity
Authors: Yu Xiaofei , Guo Chunqing , Yi Huanfa , Qian Jie , Fisher Paul B. , Subjeck John R. , Wang Xiang-Yang 
Published: January 2013

Abstract
Converting the immunosuppressive tumor environment into one that is favorable to the induction of antitumor immunity is indispensable for effective cancer immunotherapy. Here, we strategically incorporate a pathogen (i.e., flagellin)-derived, NF-κB–stimulating “danger” signal into the large stress protein or chaperone Grp170 (HYOU1/ORP150) that was previously shown to facilitate antigen crosspresentation. This engineered chimeric molecule (i.e., Flagrp170) is capable of transporting tumor antigens and concurrently inducing functional activation of dendritic cells (DC). Intratumoral administration of adenoviruses expressing Flagrp170 induces a superior antitumor response against B16 melanoma and its distant lung metastasis compared with unmodified Grp170 and flagellin. The enhanced tumor destruction is accompanied with significantly increased tumor infiltration by CD8+ cells as well as elevation of IFN-γ and interleukin (IL)-12 levels in the tumor sites. In situ Ad.Flagrp170 therapy provokes systemic activation of CTLs that recognize several antigens naturally expressing in melanoma (e.g., gp100/PMEL and TRP2/DCT). The mechanistic studies using CD11c-DTR transgenic mice and Batf3-deficient mice reveal that CD8α+ DCs are required for the improved T-cell crosspriming. Antibody neutralization assays show that IL-12 and IFN-γ are essential for the Flagrp170-elicited antitumor response, which also involves CD8+ T cells and natural killer cells. The therapeutic efficacy of Flagrp170 and its immunostimulating activity are also confirmed in mouse prostate cancer and colon carcinoma. Together, targeting the tumor microenvironment with this chimeric chaperone is highly effective in mobilizing or restoring antitumor immunity, supporting the potential therapeutic use of this novel immunomodulator in the treatment of metastatic diseases. Cancer Res; 73(7); 2093–103. ©2013 AACR.
Genetic Variation in Transforming Growth Factor Beta 1 and Mammographic Density in Singapore Chinese Women
Authors: Lee Eunjung , Van Den Berg David , Hsu Chris , Ursin Giske , Koh Woon-Puay , Yuan Jian-Min , Stram Daniel O. , Yu Mimi C. , Wu Anna H. 
Published: January 2013

Abstract
TGF-β plays a critical role in normal mammary development and morphogenesis. Decreased TGF-β signaling has been associated with increased mammographic density. Percent mammographic density (PMD) adjusted for age and body mass index (BMI) is a strong risk factor and predictor of breast cancer risk. PMD is highly heritable, but few genetic determinants have been identified. We investigated the association between genetic variation in TGFB1 and PMD using a cross-sectional study of 2,038 women who were members of the population-based Singapore Chinese Health Study cohort. We assessed PMD using a computer-assisted method. We used linear regression to examine the association between nine tagging single-nucleotide polymorphisms (SNP) of TGFB1 and PMD and their interaction with parity, adjusting for age, BMI, and dialect group. We calculated P values adjusted for correlated tests (PACT) to account for multiple testing. The strongest association was observed for rs2241716. Adjusted PMD was higher by 1.5% per minor allele (PACT = 0.04). When stratifying by parity, this association was limited to nulliparous women. For nulliparous women, adjusted PMD was higher by 8.6% per minor allele (PACT = 0.003; P for interaction with parity = 0.002). Three additional TGFB1 tagging SNPs, which were in linkage disequilibrium with rs2241716, were statistically significantly associated with adjusted PMD (PACT &lt; 0.05) for nulliparous women. However, none of these three SNPs showed statistically significant association after adjusting for rs2241716. Our data support that TGFB1 genetic variation may be an important genetic determinant of mammographic density measure that predicts breast cancer risk, particularly in nulliparous women. Cancer Res; 73(6); 1876–82. ©2012 AACR.
Characterization of a Novel PERK Kinase Inhibitor with Antitumor and Antiangiogenic Activity
Authors: Atkins Charity , Liu Qi , Minthorn Elisabeth , Zhang Shu-Yun , Figueroa David J. , Moss Katherine , Stanley Thomas B. , Sanders Brent , Goetz Aaron , Gaul Nathan , Choudhry Anthony E. , Alsaid Hasan , Jucker Beat M. , Axten Jeffrey M. , Kumar Rakesh 
Published: January 2013

Abstract
The unfolded protein response (UPR) is a signal transduction pathway that coordinates cellular adaptation to microenvironmental stresses that include hypoxia, nutrient deprivation, and change in redox status. These stress stimuli are common in many tumors and thus targeting components of the UPR signaling is an attractive therapeutic approach. We have identified a first-in-class, small molecule inhibitor of the eukaryotic initiation factor 2-alpha kinase 3 (EIF2AK3) or PERK, one of the three mediators of UPR signaling. GSK2656157 is an ATP-competitive inhibitor of PERK enzyme activity with an IC50 of 0.9 nmol/L. It is highly selective for PERK with IC50 values &gt;100 nmol/L against a panel of 300 kinases. GSK2656157 inhibits PERK activity in cells with an IC50 in the range of 10–30 nmol/L as shown by inhibition of stress-induced PERK autophosphorylation, eIF2α substrate phosphorylation, together with corresponding decreases in ATF4 and CAAT/enhancer binding protein homologous protein (CHOP) in multiple cell lines. Oral administration of GSK2656157 to mice shows a dose- and time-dependent pharmacodynamic response in pancreas as measured by PERK autophosphorylation. Twice daily dosing of GSK2656157 results in dose-dependent inhibition of multiple human tumor xenografts growth in mice. Altered amino acid metabolism, decreased blood vessel density, and vascular perfusion are potential mechanisms for the observed antitumor effect. However, despite its antitumor activity, given the on-target pharmacologic effects of PERK inhibition on pancreatic function, development of any PERK inhibitor in human subjects would need to be cautiously pursued in cancer patients. Cancer Res; 73(6); 1993–2002. ©2012 AACR.
Enhanced Sonographic Imaging to Diagnose Lymph Node Metastasis: Importance of Blood Vessel Volume and Density
Authors: Li Li , Mori Shiro , Kodama Mizuho , Sakamoto Maya , Takahashi Shoki , Kodama Tetsuya 
Published: January 2013

AbstractLymph node size is an important variable in ultrasound diagnosis of lymph node metastasis. However, the size criterion often leads to oversight of tumor-positive lymph nodes within the range of “normal” size, such that more accurate diagnostic criteria for lymph node metastasis are required. In this study, we show how diagnosis of lymph node metastasis can be improved by evaluating changes in blood vessel volume and density using a novel contrast-enhanced high-frequency ultrasound (CE-HFUS) system with Sonazoid. An MRL/MpJ-lpr/lpr (MRL/lpr) mouse model of lymph node metastasis was used in which lymph nodes are similar in size to humans. Metastasis via lymphatic vessels to proper axillary lymph nodes (proper ALN) was induced by injection of tumor cells into the subiliac lymph nodes. Within 21 days of injection, significant increases in blood vessel volume and density, but no increases in the size of the proper ALNs, were observed. The increase in blood vessel density was confirmed with immunohistochemical analysis and was positively related to tumor cell proliferation as measured using bioluminescence imaging. Together, our results showed that alterations in blood vessel volume and density precede alterations in lymph node size in the early stages of lymph node metastasis. Detection of these changes by ultrasonography may offer new criteria for early diagnosis of lymph node metastasis. Cancer Res; 73(7); 2082–92. ©2013 AACR.
Viral Load of High-Risk Human Papillomaviruses as Reliable Clinical Predictor for the Presence of Cervical Lesions
Authors: Schmitt Markus , Depuydt Christophe , Benoy Ina , Bogers Johannes , Antoine Jerome , Pawlita Michael , Arbyn Marc 
Published: January 2013

Abstract
Background: Infections with high-risk human papillomaviruses (Hr-HPV) can cause malignant transformation of the human cervical epithelium. HPV DNA tests generally are very sensitive to detect cervical neoplastic lesions but also identify transient HPV infections. As a consequence, the specificity and positive predictive value are low.
Methods: We analyzed viral load of Hr- and possibly Hr-HPV types more than seven orders of magnitude (on a log10 scale) in 999 consecutive BD-SurePath liquid-based cervical cytology samples from routine cervical screening enriched with atypical squamous cells of undetermined significance (n = 100), low-grade squamous intraepithelial lesions (LSIL; n = 100), and high-grade squamous intraepithelial lesions (HSIL; n = 97) using type-specific multiplex quantitative real-time PCR and the BSGP5+/6+-PCR/MPG assay. In the 36-month follow-up, 79 histologically verified CIN2+ and 797 double-negative cytology cases were identified.
Results: Viral loads in LSIL and HSIL were significantly increased compared with no intraepithelial lesion or malignancy in both the quantitative PCR (qPCR) and BSGP5+/6+-PCR/MPG assay (P &lt; 0.0001). The mean viral loads in LSIL and HSIL were not significantly different. Using a newly determined high viral load cut off for 14 Hr-HPV types, the sensitivity for prevalent CIN3+ remained at 100% for both assays compared with the minimal detection threshold. The specificity (corresponding to double-negative cytology at subsequent screening episodes) increased substantially (qPCR, from 91.1% to 95.7%; BSGP5+/6+-PCR/MPG, from 79.8% to 96.2%).
Conclusions: Compared with DNA positivity alone, high Hr-HPV viral loads could reduce the amount of false positive results detected by the BSGP5+/6+-PCR/MPG and qPCR by 81.4% and 52.1%, respectively.
Impact: Quantitative type-specific HPV DNA assays show high flexibility in defining thresholds that allow optimizing clinical accuracy for cervical cancer precursors. Cancer Epidemiol Biomarkers Prev; 22(3); 406–14. ©2013 AACR.