bluuebunny/crossref_metadata_2025 huggingface dataset:
Changes in the
agr
Locus Affect Enteritis Caused by Methicillin-Resistant
Staphylococcus aureus
Authors: Sugiyama Yoichi , Okii Kazuya , Murakami Yoshiaki , Yokoyama Takashi , Takesue Yoshio , Ohge Hiroki , Sueda Taijiro , Hiyama Eiso
Published: March 2009

We studied the characteristics of methicillin (meticillin)-resistant
Staphylococcus aureus
(MRSA) strains that caused enteritis. In a previous report, we demonstrated that both phenotypic and genotypic changes were associated with MRSA enteritis; and we hypothesized that the accessory gene regulator (
agr
), which is a global regulator of staphylococcal virulence and upregulates several exoproteins, is the key factor associated with the development of MRSA enteritis. In this study, we examined 12 MRSA isolates associated with enteritis from stool samples and 17 MRSA isolates not associated with enteritis that had the following characteristics: the strains associated with enteritis had the same genotype (genotype A), as detected by pulsed-field gel electrophoresis, or the strains were isolated from stools. The differences between strains that caused enteritis and those that did not cause enteritis strains were examined by quantitative reverse transcription-PCR to assess RNAII,
agrA
, RNAIII, and
tst
expression and by sequencing of the
agr
locus. The levels of expression of
agrA
, RNAIII, and
tst
were higher by the MRSA isolates associated with enteritis than by the MRSA isolates not associated with enteritis, whether or not they were of the same genotype. The levels of expression of RNAII by almost all the clinical isolates were similar. Sequencing of the
agr
locus showed that all MRSA isolates that caused enteritis have
agr
mutations, whereas the MRSA isolates that did not cause enteritis, with three exceptions, did not. Many of the isolates associated with enteritis had the same mutation, especially at the C-terminal end of
agrA
. These results suggest a trend in which mutations in the
agr
locus modify the expression of
agrA
and RNAIII and the production of toxin, all of which may increase the virulence and influence the occurrence of MRSA enteritis.
Brain Abscess Caused by
Tsukamurella tyrosinosolvens
in an Immunocompetent Patient
Authors: Sheng Wang-Huei , Huang Yu-Tsung , Chang Shan-Chwen , Hsueh Po-Ren
Published: March 2009

We describe a previously healthy patient with chronic otitis media complicated with cerebellar abscess caused by
Tsukamurella tyrosinosolvens
. The organism was identified based on conventional biochemical identification methods, PCR-restriction fragment length polymorphism analysis of the
hsp65
gene, and 16S rRNA gene sequencing. The patient was treated successfully with debridements and prolonged antibiotic therapy.
Rapid Molecular Genotyping and Clonal Complex Assignment of
Staphylococcus aureus
Isolates by PCR Coupled to Electrospray Ionization-Mass Spectrometry
Authors: Hall Thomas A. , Sampath Rangarajan , Blyn Lawrence B. , Ranken Raymond , Ivy Cristina , Melton Rachael , Matthews Heather , White Neill , Li Feng , Harpin Vanessa , Ecker David J. , McDougal Linda K. , Limbago Brandi , Ross Tracy , Wolk Donna M. , Wysocki Vicki , Carroll Karen C.
Published: March 2009

We describe a high-throughput assay using PCR coupled to electrospray ionization-mass spectrometry (PCR/ESI-MS) to determine the genotypes of
Staphylococcus aureus
isolates. The primer sets used in the PCR/ESI-MS assay were designed to amplify the same genes analyzed in multilocus sequence typing (MLST). The method was used to identify the clonal complex and USA type of each isolate and is suitable for use in a clinical or public-health setting. The method was validated using a panel of diverse isolates from the Centers for Disease Control and Prevention that were previously characterized by MLST and pulsed-field gel electrophoresis (PFGE). Clinical isolates from two geographically distinct hospitals were characterized, and the clustering results were in agreement with those for repetitive-element PCR and PFGE. The PCR/ESI-MS method enables genotyping of over 180 samples of
S. aureus
per day in an automated fashion.
Pseudomonas aeruginosa
Acute Prostatitis and Urosepsis after Sexual Relations in a Hot Tub
Authors: Dulabon Lori M. , LaSpina Mark , Riddell Scott W. , Kiska Deanna L. , Cynamon Michael
Published: March 2009

We report a case of a previously healthy 38-year-old male with acute prostatitis and concurrent
Pseudomonas aeruginosa
urosepsis. Pulsed-field gel electrophoresis analysis confirmed that the source of the organism was the patient's newly purchased hot tub, which was filled with water from a stream.
DDX3 DEAD-Box RNA Helicase Inhibits Hepatitis B Virus Reverse Transcription by Incorporation into Nucleocapsids
Authors: Wang Haifeng , Kim Seahee , Ryu Wang-Shick
Published: March 2009

Viruses utilize host factors in many steps of their life cycles. Yet, little is known about host factors that contribute to the life cycle of hepatitis B virus (HBV), which replicates its genome by reverse transcription. To identify host factors that contribute to viral reverse transcription, we sought to identify cellular proteins that interact with HBV polymerase (Pol) by using affinity purification coupled with mass spectrometry. One of the HBV Pol-interacting host factors identified was DDX3 DEAD-box RNA helicase, which unwinds RNA in an ATPase-dependent manner. Recently, it was shown that DDX3 is essential for both human immunodeficiency virus and hepatitis C virus infection. In contrast, we found that the ectopic expression of DDX3 led to significantly reduced viral DNA synthesis. The DDX3-mediated inhibition of viral DNA synthesis did not affect RNA encapsidation, a step prior to reverse transcription, and indicated that DDX3 inhibits HBV reverse transcription. Mutational analysis revealed that mutant DDX3 with an inactive ATPase motif, but not that with an inactive RNA helicase motif, failed to inhibit viral DNA synthesis. Our interpretation is that DDX3 inhibits viral DNA synthesis at a step following ATP hydrolysis but prior to RNA unwinding. Finally, OptiPrep density gradient analysis revealed that DDX3 was incorporated into nucleocapsids, suggesting that DDX3 inhibits viral reverse transcription following nucleocapsid assembly. Thus, DDX3 represents a novel host restriction factor that limits HBV infection.
Type I and Type II Interferons Inhibit the Translation of Murine Norovirus Proteins
Authors: Changotra Harish , Jia Yali , Moore Tara N. , Liu Guangliang , Kahan Shannon M. , Sosnovtsev Stanislav V. , Karst Stephanie M.
Published: March 2009

Human noroviruses are responsible for more than 95% of nonbacterial epidemic gastroenteritis worldwide. Both onset and resolution of disease symptoms are rapid, suggesting that components of the innate immune response are critical in norovirus control. While the study of the human noroviruses has been hampered by the lack of small animal and tissue culture systems, our recent discovery of a murine norovirus (MNV) and its in vitro propagation have allowed us to begin addressing norovirus replication strategies and immune responses to norovirus infection. We have previously demonstrated that interferon responses are critical to control MNV-1 infection in vivo and to directly inhibit viral replication in vitro. We now extend these studies to define the molecular basis for interferon-mediated inhibition. Viral replication intermediates were not detected in permissive cells pretreated with type I interferon after either infection or transfection of virion-associated RNA, demonstrating a very early block to virion production that is after virus entry and uncoating. A similar absence of viral replication intermediates was observed in infected primary macrophages and dendritic cells pretreated with type I IFN. This was not due to degradation of incoming genomes in interferon-pretreated cells since similar levels of genomes were present in untreated and pretreated cells through 6 h of infection, and these genomes retained their integrity. Surprisingly, this block to the translation of viral proteins was not dependent on the well-characterized interferon-induced antiviral molecule PKR. Similar results were observed in cells pretreated with type II interferon, except that the inhibition of viral translation was dependent on PKR. Thus, both type I and type II interferon signaling inhibit norovirus translation in permissive myeloid cells, but they display distinct dependence on PKR for this inhibition.
Biochemical Characterization of Arterivirus Nonstructural Protein 11 Reveals the Nidovirus-Wide Conservation of a Replicative Endoribonuclease
Authors: Nedialkova Danny D. , Ulferts Rachel , van den Born Erwin , Lauber Chris , Gorbalenya Alexander E. , Ziebuhr John , Snijder Eric J.
Published: March 2009

Nidoviruses (arteriviruses, coronaviruses, and roniviruses) are a phylogenetically compact but diverse group of positive-strand RNA viruses that includes important human and animal pathogens. Nidovirus RNA synthesis is mediated by a cytoplasmic membrane-associated replication/transcription complex that includes up to 16 viral nonstructural proteins (nsps), which carry common enzymatic activities, like the viral RNA polymerase, but also unusual and poorly understood RNA-processing functions. Of these, a conserved endoribonuclease (NendoU) is a major genetic marker that is unique to nidoviruses. NendoU activity was previously verified in vitro for the coronavirus nsp15, but not for any of its distantly related orthologs from other nidovirus lineages, like the arterivirus nsp11. Here, we show that the bacterially expressed nsp11 proteins of two arteriviruses, equine arteritis virus and porcine respiratory and reproductive syndrome virus, possess pyrimidine-specific endoribonuclease activity. RNA cleavage was independent of divalent cations in vitro and was greatly reduced by replacement of residues previously implicated in catalysis. Comparative characterization of the NendoU activity in arteriviruses and severe acute respiratory syndrome coronavirus revealed common and distinct features of their substrate requirements and reaction mechanism. Our data provide the first biochemical evidence of endoribonuclease activity associated with arterivirus nsp11 and support the conclusion that this remarkable RNA-processing enzyme, whose substrate in the infected cell remains to be identified, distinguishes nidoviruses from all other RNA viruses.
Modified state parameter for characterizing static liquefaction of sand with fines
Authors: Bobei D. C. , Lo S. R. , Wanatowski D. , Gnanendran C. T. , Rahman M. M.
Published: March 2009

An experimental study was carried out to investigate the static liquefaction behaviour of sand with a small amount of plastic and nonplastic fines. Five series of tests were conducted in drained and undrained conditions. The drained test results indicate not only that the failure line coincides with the critical state, but also that the development of volumetric strain during shearing was not sensitive to the initial confining pressure. In both isotropically and anisotropically consolidated undrained tests, a so-called “reverse behaviour” was consistently observed. The results were also interpreted in the critical state framework. The critical and steady state (CS/SS) data were found to trace along the same curve in e–log( p′) space, irrespective of the stress history and effective stress paths. A comparison between the isotropic consolidation line (ICL) and critical state (CS) curve showed that a small amount of fines can significantly change the shape and position of the ICL relative to the CS curve. Furthermore, the soil behaviour manifested in both drained and undrained shearing led to the development of a modified state parameter.
The Structural and Functional Diversity of Metabolite-Binding Riboswitches
Authors: Roth Adam , Breaker Ronald R.
Published: March 2009

The cellular concentrations of certain metabolites are assiduously monitored to achieve appropriate levels of gene expression. Although proteins have long been known to act as sensors in this capacity, metabolite-binding RNAs, or riboswitches, also play an important role. More than 20 distinct classes of riboswitches have been identified to date, and insights to the molecular recognition strategies of a significant subset of these have been provided by detailed structural studies. This diverse set of metabolite-sensing RNAs is found to exploit a variety of distinct mechanisms to regulate genes that are fundamental to metabolism.
Amantadine inhibits platelet-activating factor induced clathrin-mediated endocytosis in human neutrophils
Authors: Eckels Phillip C. , Banerjee Anirban , Moore Ernest E. , McLaughlin Nathan J. D. , Gries Lynn M. , Kelher Marguerite R. , England Kelly M. , Gamboni-Robertson Fabia , Khan Samina Y. , Silliman Christopher C.
Published: March 2009

Receptor signaling is integral for adhesion, emigration, phagocytosis, and reactive oxygen species production in polymorphonuclear neutrophils (PMNs). Priming is an important part of PMN emigration, but it can also lead to PMN-mediated organ injury in the host. Platelet-activating factor (PAF) primes PMNs through activation of a specific G protein-coupled receptor. We hypothesize that PAF priming of PMNs requires clathrin-mediated endocytosis (CME) of the PAF receptor (PAFr), and, therefore, amantadine, known to inhibit CME, significantly antagonizes PAF signaling. PMNs were isolated by standard techniques to >98% purity and tested for viability. Amantadine (1 mM) significantly inhibited the PAF-mediated changes in the cellular distribution of clathrin and the physical colocalization [fluorescence resonance energy transfer positive (FRET+)] of early endosome antigen-1 and Rab5a, known components of CME and similar to hypertonic saline, a known inhibitor of CME. Furthermore, amantadine had no effect on the PAF-induced cytosolic calcium flux; however, phosphorylation of p38 MAPK was significantly decreased. Amantadine inhibited PAF-mediated changes in PMN physiology, including priming of the NADPH oxidase and shape change with lesser inhibition of increases in CD11b surface expression and elastase release. Furthermore, rimantadine, an amantadine analog, was a more potent inhibitor of PAF priming of the N-formyl-methionyl-leucyl-phenylalanine-activated oxidase. PAF priming of PMNs requires clathrin-mediated endocytosis that is inhibited when PMNs are pretreated with either amantadine or rimantadine. Thus, amantadine and rimantadine have the potential to ameliorate PMN-mediated tissue damage in humans.
Loss of function of Sco1 and its interaction with cytochrome c oxidase
Authors: Stiburek Lukas , Vesela Katerina , Hansikova Hana , Hulkova Helena , Zeman Jiri
Published: March 2009

Sco1 and Sco2 are mitochondrial copper-binding proteins involved in the biogenesis of the CuA site in the cytochrome c oxidase (CcO) subunit Cox2 and in the maintenance of cellular copper homeostasis. Human Surf1 is a CcO assembly factor with an important but poorly characterized role in CcO biogenesis. Here, we analyzed the impact on CcO assembly and tissue copper levels of a G132S mutation in the juxtamembrane region of SCO1 metallochaperone associated with early onset hypertrophic cardiomyopathy, encephalopathy, hypotonia, and hepatopathy, assessed the total copper content of various SURF1 and SCO2-deficient tissues, and investigated the possible physical association between CcO and Sco1. The steady-state level of mutant Sco1 was severely decreased in the muscle mitochondria of the SCO1 patient, indicating compromised stability and thus loss of function of the protein. Unlike the wild-type variant, residual mutant Sco1 appeared to migrate exclusively in the monomeric form on blue native gels. Both the activity and content of CcO were reduced in the patient's muscle to ∼10–20% of control values. SCO1-deficient mitochondria showed accumulation of two Cox2 subcomplexes, suggesting that Sco1 is very likely responsible for a different posttranslational aspect of Cox2 maturation than Sco2. Intriguingly, the various SURF1-deficient samples analyzed showed a tissue-specific copper deficiency similar to that of SCO-deficient samples, suggesting a role for Surf1 in copper homeostasis regulation. Finally, both blue native immunoblot analysis and coimmunoprecipitation revealed that a fraction of Sco1 physically associates with the CcO complex in human muscle mitochondria, suggesting a possible direct relationship between CcO and the regulation of cellular copper homeostasis.
Cell proliferation of human ovarian cancer is regulated by the opioid growth factor-opioid growth factor receptor axis
Authors: Donahue Renee N. , McLaughlin Patricia J. , Zagon Ian S.
Published: March 2009

Ovarian cancer is the leading cause of death from gynecological malignancies. Understanding the biology of these tumors, as well as treatment modalities, has been challenging. The opioid growth factor (OGF; [Met5]-enkephalin) and the OGF receptor (OGFr) form an endogenous growth-regulating pathway in homeostasis and neoplasia. In this investigation, we examined the relationship of the OGF-OGFr axis to ovarian cancer, and defined its presence, function, and mechanisms. Using OVCAR-3 and SKOV-3 ovarian cancer cell lines, we found that OGF and OGFr were present and functional. Exogenous OGF was observed to have a dose-dependent, serum-independent, reversible, and receptor-mediated inhibitory action on cell proliferation that was dependent on RNA and protein synthesis. The repressive effect of OGF on cell proliferation also was observed in SW626, CAOV-3, and HEY ovarian cancer cell lines. Endogenous OGF was found to be constitutively produced and tonically active on cell replicative activities, with neutralization of this peptide accelerating cell proliferation. Silencing of OGFr using siRNA technology stimulated cell replication, documenting its integral role. The mechanism of OGF-OGFr action on DNA synthesis was related to the cyclin-dependent kinase inhibitory pathway because knockdown of p16 or p21 in OVCAR-3 cells, and p21 in SKOV-3 cells, eliminated OGF's inhibitory effect on growth. These data are the first to report that the OGF-OGFr system is a native biological regulator of cell proliferation in human ovarian cancer. This information will be important in designing treatment strategies for this deadly disease.
Cold stimulates the behavioral response to hypoxia in newborn mice
Authors: Bollen Bieke , Bouslama Myriam , Matrot Boris , Rotrou Yann , Vardon Guy , Lofaso Frédéric , Van den Bergh Omer , D'Hooge Rudi , Gallego Jorge
Published: March 2009

In newborns, hypoxia elicits increased ventilation, arousal followed by defensive movements, and cries. Cold is known to affect the ventilatory response to hypoxia, but whether it affects the arousal response remains unknown. The aim of the present study was to assess the effects of cold on the ventilatory and arousal responses to hypoxia in newborn mice. We designed an original platform measuring noninvasively and simultaneously the breathing pattern by whole body plethysmography, body temperature by infrared thermography, as well as motor and ultrasonic vocal (USV) responses. Six-day-old mice were exposed twice to 10% O2for 3 min at either cold temperature (26°C) or thermoneutrality (33°C). At 33°C, hypoxia elicited a marked increase in ventilation followed by a small ventilatory decline, small motor response, and almost no USVs. Body temperature was not influenced by hypoxia, and oxygen consumption (V̇o2) displayed minimal changes. At 26°C, hypoxia elicited a slight increase in ventilation with a large ventilatory decline and a large drop of V̇o2. This response was accompanied by marked USV and motor responses. Hypoxia elicited a small decrease in temperature after the return to normoxia, thus precluding any causal influence on the motor and USV responses to hypoxia. In conclusion, cold stimulated arousal and stress responses to hypoxia, while depressing hypoxic hyperpnea. Arousal is an important defense mechanism against sleep-disordered breathing. The dissociation between ventilatory and behavioral responses to hypoxia suggests that deficits in the arousal response associated with sleep breathing disorders cannot be attributed to a depressed hypoxic response.
Effects of aerobic and anaerobic metabolic inhibitors on avian intrapulmonary chemoreceptors
Authors: Pilarski Jason Q. , Solomon Irene C. , Kilgore Delbert L. , Hempleman Steven C.
Published: March 2009

Birds have rapidly responding respiratory chemoreceptors [intrapulmonary chemoreceptors (IPC)] that provide vagal sensory feedback about breathing pattern. IPC are exquisitely sensitive to CO2but are unaffected by hypoxia. IPC continue to respond to CO2during hypoxic and even anoxic conditions, suggesting that they may generate ATP needed for signal transduction anaerobically. To assess IPC energy metabolism, single-cell action potential discharge and acid-base status were recorded from 26 pentobarbital-anesthetized Anas platyrhynchos before and after intravenous infusion of the glycolytic blocker iodoacetate (10–70 mg/kg), mitochondrial blocker rotenone (2 mg/kg), and/or mitochondrial uncoupler 2,4-dinitrophenol (5–15 mg/kg). After 5 min exposure at the highest dosages, iodoacetate inhibited IPC discharge 65% (15.9 ± 0.3 s−1to 5.5 ± 0.3 s−1, P < 0.05), rotenone inhibited discharge 80% (12.9 ± 0.5 s−1to 2.6 ± 0.6 s−1, P < 0.05), and 2,4-dinitrophenol inhibited discharge 19% (14.0 ± 0.3 s−1to 11.3 ± 0.3 s−1, P < 0.05). These results suggest that IPC utilize glucose, require an intact glycolytic pathway, and metabolize the products of glycolysis to CO2and H2O by mitochondrial respiration. The small but significant effect of 2,4-dinitrophenol suggests that ATP production by glycolysis may be sufficient to meet IPC energy demands if NADH can be oxidized to NAD experimentally by uncoupling mitochondria, or physiologically by transient lactate production. A model for IPC spike frequency adaptation is proposed, whereby the rapid onset of phasic IPC discharge requires ATP from anaerobic glycolysis, using lactate as the electron acceptor, and the roll-off in IPC discharge reflects transient acidosis due to intracellular lactic acid accumulation.
On the Consequences of Resampling Microwave Radiometer Observations for Use in Retrieval Algorithms
Authors: Rapp Anita D. , Lebsock M. , Kummerow C.
Published: March 2009

How to deal with the different spatial resolutions of multifrequency satellite microwave radiometer measurements is a common problem in retrievals of cloud properties and rainfall. Data convolution and deconvolution is a common approach to resampling the measurements to a single resolution. Tropical Rainfall Measuring Mission (TRMM) Microwave Imager (TMI) measurements are resampled to the resolution of the 19-GHz field of view for use in a multifrequency optimal estimation retrieval algorithm of cloud liquid water path, total precipitable water, and wind speed. Resampling the TMI measurements is found to have a strong influence on retrievals of cloud liquid water path and a slight influence on wind speed. Beam-filling effects in the resampled brightness temperatures are shown to be responsible for the large differences between the retrievals using the TMI native resolution and resampled brightness temperatures. Synthetic retrievals are performed to test the sensitivity of the retrieved parameters to beam-filling effects in the resampling of each of the different channels. Beam-filling effects due to the convolution of the 85-GHz channels are shown to be the largest contributor to differences in retrieved cloud liquid water path. Differences in retrieved wind speeds are found to be a combination of effects from deconvolving the 10-GHz brightness temperatures and compensation effects due to the lower liquid water path being retrieved by the high-frequency channels. The influence of beam-filling effects on daily and monthly averages of cloud liquid water path is also explored. Results show that space–time averaging of cloud liquid water path cannot fully compensate for the beam-filling effects and should be considered when using cloud liquid water path data for validation or in climate studies.
An Observing System Experiment for Typhoon Conson (2004) Using a Singular Vector Method and DOTSTAR Data
Authors: Yamaguchi Munehiko , Iriguchi Takeshi , Nakazawa Tetsuo , Wu Chun-Chieh
Published: March 2009

An Observing System Experiment (OSE) has been performed to investigate the effectiveness of dropwindsonde observations and a sensitivity analysis technique on a typhoon track forecast. Using dropwindsonde observations for Typhoon Conson at 1200 UTC 8 June 2004, which are derived from Dropwindsonde Observation for Typhoon Surveillance near the Taiwan Region (DOTSTAR), four numerical experiments are conducted, which are different only in terms of the number of dropwindsonde observations used in a data assimilation system: (i) no observation is assimilated; (ii) all observations are assimilated; (iii) observations within a sensitive region as revealed by a singular vector method at the Japan Meteorological Agency (JMA) are assimilated; and (iv) observations outside the sensitive region are assimilated. In the comparison of the four track forecasts, Conson’s northeastward movement is expressed in the second and third simulations while in the first and fourth experiments Conson stays at almost the same position as its initial position. Through the OSE, it is found that DOTSTAR observations had a positive impact on the track forecast for Conson, and that observations within the sensitive region are enough to predict the northeastward movement of Conson, indicating that the JMA singular vector method would be useful for the sampling strategy of targeted observations like DOTSTAR.
A High-Resolution Simulation of Asymmetries in Severe Southern Hemisphere Tropical Cyclone Larry (2006)
Authors: Ramsay Hamish A. , Leslie Lance M. , Kepert Jeffrey D.
Published: March 2009

Advances in observations, theory, and modeling have revealed that inner-core asymmetries are a common feature of tropical cyclones (TCs). In this study, the inner-core asymmetries of a severe Southern Hemisphere tropical cyclone, TC Larry (2006), are investigated using the fifth-generation Pennsylvania State University–National Center for Atmospheric Research Mesoscale Model (MM5) and the Kepert–Wang boundary layer model. The MM5-simulated TC exhibited significant asymmetries in the inner-core region, including rainfall distribution, surface convergence, and low-level vertical motion. The near-core environment was characterized by very low environmental vertical shear and consequently the TC vortex had almost no vertical tilt. It was found that, prior to landfall, the rainfall asymmetry was very pronounced with precipitation maxima consistently to the right of the westward direction of motion. Persistent maxima in low-level convergence and vertical motion formed ahead of the translating TC, resulting in deep convection and associated hydrometeor maxima at about 500 hPa. The asymmetry in frictional convergence was mainly due to the storm motion at the eyewall, but was dominated by the proximity to land at larger radii. The displacement of about 30°–120° of azimuth between the surface and midlevel hydrometeor maxima is explained by the rapid cyclonic advection of hydrometeors by the tangential winds in the TC core. These results for TC Larry support earlier studies that show that frictional convergence in the boundary layer can play a significant role in determining the asymmetrical structures, particularly when the environmental vertical shear is weak or absent.
The Changing Regional Subcultures of the American States and the Utility of a New Cultural Measure
Authors: Lieske Joel
Published: March 2009

This study analyzes changes in the regional subcultures of the United States using 2000 census and religious survey data. The results suggest a remarkable degree of continuity with those the authors identified in an earlier study. In addition, they demonstrate that a new multidimensional measure of state culture does a much better job in predicting social and political behavior than other frequently used indicators. Finally, they show how their new measure of state culture significantly reduces and often eliminates the problem of spatial autocorrelation in many state-level indicators that cannot be explained by differences in economic development and racial—ethnic diversity.
Reciprocal responsiveness to interleukin-12 and interferon-α specifies human CD8+ effector versus central memory T-cell fates
Authors: Ramos Hilario J. , Davis Ann M. , Cole Alexander G. , Schatzle John D. , Forman James , Farrar J. David
Published: March 2009

Multiple innate signals regulate the genesis of effector and memory CD8+ T cells. In this study, we demonstrate that the innate cytokines interleukin (IL)–12 and interferon (IFN)–α/β regulate distinct aspects of effector and memory human CD8+ T-cell differentiation. IL-12 exclusively promoted the development of IFN-γ– and tumor necrosis factor (TNF)–α–secreting T effector memory (TEM) cells, whereas IFN-α drove the development of T central memory (TCM) cells. The development of TEM and TCM was linked to cell division. In rapidly dividing cells, IL-12 programmed TEM through induction of the IL-12 receptor β2. In contrast, IFN-α regulated TCM development by slowing the progression of cell division in a subpopulation of cells that selectively expressed elevated IFN-α/β receptor-2. The strength of signal delivered through T-cell receptor (TCR) engagement regulated the responsiveness of cells to IL-12 and IFN-α. In the presence of both IL-12 and IFN-α, these cytokine signals were amplified as the strength of the TCR signal was increased, promoting the simultaneous development of both TCM and TEM. Together, our results support a novel model in which IL-12 and IFN-α act in a nonredundant manner to regulate the colinear generation of both effector and memory cells.
Tumor necrosis factor-alpha induced expression of matrix metalloproteinase-9 through p21-activated Kinase-1
Authors: Zhou Ling , Yan Chunli , Gieling Roben G , Kida Yujiro , Garner Warren , Li Wei , Han Yuan-Ping
Published: March 2009

Background

Expressed in embryonic development, matrix metalloprotein-9 (MMP-9) is absent in most of developed adult tissues, but recurs in inflammation during tissue injury, wound healing, tumor formation and metastasis. Expression of MMP-9 is tightly controlled by extracellular cues including pro-inflammatory cytokines and extracellular matrix (ECM). While the pathologic functions of MMP-9 are evident, the intracellular signaling pathways to control its expression are not fully understood. In this study we investigated mechanism of cytokine induced MMP-9 with particular emphasis on the role of p21-activated-kinase-1 (PAK1) and the down stream signaling.

Results

In response to TNF-alpha or IL-1alpha, PAK1 was promptly activated, as characterized by a sequential phosphorylation, initiated at threonine-212 followed by at threonine-423 in the activation loop of the kinase, in human skin keratinocytes, dermal fibroblasts, and rat hepatic stellate cells. Ectopic expression of PAK1 variants, but not p38 MAP kinase, impaired the TNF-alpha-induced MMP-9 expression, while other MMPs such as MMP-2, -3 and -14 were not affected. Activation of Jun N-terminal kinase (JNK) and NF-kappaB has been demonstrated to be essential for MMP-9 expression. Expression of inactive PAK1 variants impaired JNK but not NF-kappaB activation, which consequently suppressed the 5'-promoter activities of the MMP-9 gene. After the cytokine-induced phosphorylation, both ectopically expressed and endogenous PAK1 proteins were promptly accumulated even in the condition of suppressing protein synthesis, suggesting the PAK1 protein is stabilized upon TNF-alpha stimulation. Stabilization of PAK1 protein by TNF-alpha treatment is independent of the kinase catalytic activity and p21 GTPase binding capacities. In contrast to epithelial cells, mesenchymal cells require 3-dimensional type-I collagen in response to TNF-alpha to massively express MMP-9. The collagen effect is mediated, in part, by boost JNK activation in a way to cooperate the cytokine signaling.

Conclusion

We identified a novel mechanism for MMP-9 expression in response to injury signals, which is mediated by PAK1 activation and stabilization leading JNK activation.
Role of sgcR3 in positive regulation of enediyne antibiotic C-1027 production of Streptomyces globisporus C-1027
Authors: Wang Lifei , Hu Yunfeng , Zhang Yanjuan , Wang Songmei , Cui Zhihui , Bao Yi , Jiang Wei , Hong Bin
Published: March 2009

Background

C-1027, produced by Streptomyces globisporus C-1027, is one of the most potent antitumoral agents. The biosynthetic gene cluster of C-1027, previously cloned and sequenced, contains at least three putative regulatory genes, i.e. sgcR1, sgcR2 and sgcR3. The predicted gene products of these genes share sequence similarities to StrR, regulators of AraC/XylS family and TylR. The purpose of this study was to investigate the role of sgcR3 in C-1027 biosynthesis.

Results

Overexpression of sgcR3 in S. globisporus C-1027 resulted in a 30–40% increase in C-1027 production. Consistent with this, disruption of sgcR3 abolished C-1027 production. Complementation of the sgcR3-disrupted strain R3KO with intact sgcR3 gene could restore C-1027 production. The results from real time RT-PCR analysis in R3KO mutant and wild type strain indicated that not only transcripts of biosynthetic structural genes such as sgcA1 and sgcC4, but also putative regulatory genes, sgcR1 and sgcR2, were significantly decreased in R3KO mutant. The cross-complementation studies showed that sgcR1R2 could functionally complement sgcR3 disruption in trans. Purified N-terminal His10-tagged SgcR3 showed specific DNA-binding activity to the promoter region of sgcR1R2.

Conclusion

The role of SgcR3 has been proved to be a positive regulator of C-1027 biosynthesis in S. globisporus C-1027. SgcR3 occupies a higher level than SgcR1 and SgcR2 in the regulatory hierarchy that controls C-1027 production and activates the transcription of sgcR1 and sgcR2 by binding directly to the promoter region of sgcR1R2.
Viral and murine interleukin-10 are correctly processed and retain their biological activity when produced in tobacco
Authors: Bortesi Luisa , Rossato Marzia , Schuster Flora , Raven Nicole , Stadlmann Johannes , Avesani Linda , Falorni Alberto , Bazzoni Flavia , Bock Ralph , Schillberg Stefan , Pezzotti Mario
Published: March 2009

Background

Interleukin-10 (IL-10) is a potent anti-inflammatory cytokine, with therapeutic applications in several autoimmune and inflammatory diseases. Oral administration of this cytokine alone, or in combination with disease-associated autoantigens could confer protection form the onset of a specific autoimmune disease through the induction of oral tolerance. Transgenic plants are attractive systems for production of therapeutic proteins because of the ability to do large scale-up at low cost, and the low maintenance requirements. They are highly amenable to oral administration and could become effective delivery systems without extensive protein purification. We investigated the ability of tobacco plants to produce high levels of biologically-active viral and murine IL-10.

Results

Three different subcellular targeting strategies were assessed in transient expression experiments, and stable transgenic tobacco plants were generated with the constructs that yielded the highest accumulation levels by targeting the recombinant proteins to the endoplasmic reticulum. The best yields using this strategy in T1plants were 10.8 and 37.0 μg/g fresh leaf weight for viral and murine IL-10, respectively. The recombinant proteins were purified from transgenic leaf material and characterized in terms of theirN-glycan composition, dimerization and biological activity inin vitroassays. Both molecules formed stable dimers, were able to activate the IL-10 signaling pathway and to induce specific anti-inflammatory responses in mouse J774 macrophage cells.

Conclusion

Tobacco plants are able to correctly process viral and murine IL-10 into biologically active dimers, therefore representing a suitable platform for the production for these cytokines. The accumulation levels obtained are high enough to allow delivery of an immunologically relevant dose of IL-10 in a reasonable amount of leaf material, without extensive purification. This study paves the way to performing feeding studies in mouse models of autoimmune diseases, that will allow the evaluation the immunomodulatory properties and effectiveness of the viral IL-10 in inducing oral tolerance compared to the murine protein.
Peroxisome proliferator-activated receptor-γ protects ERBB2-positive breast cancer cells from palmitate toxicity
Authors: Kourtidis Antonis , Srinivasaiah Rekha , Carkner Richard D , Brosnan M Julia , Conklin Douglas S
Published: March 2009

Introduction

Accumulation of fatty acids and neutral lipids in nonadipose tissues is cytotoxic. We recently showed that ERBB2-positive breast cancer cells produce significantly high amounts of fats, because of overexpression of the peroxisome proliferator-activated receptor (PPAR)γ-binding protein and the nuclear receptor NR1D1 (nuclear receptor subfamily 1, group D, member 1; Rev-erbα). These genes upregulate de novo fatty acid synthesis, which is a critical pathway for the energy production and survival of these cells. NR1D1 and PPARγ-binding protein are functionally related to PPARγ, a well established positive regulator of adipogenesis and lipid storage.

Methods

The effects of GW9662 and exogenously added palmitate on breast cells (BT474, MDA-MB-361, MCF-7, and human mammary epithelial cells) in monolayer culture were assessed. Mass spectrometric quantitation of fatty acids and fluorescence-based high content microscopy assays of cell growth, apoptosis, triglyceride storage and reactive oxygen species production were used.

Results

ERBB2-positive breast cancer cells are more sensitive to inhibition of PPARγ activity by the antagonist GW9662. PPARγ inhibition results in increased levels of total fats in the cells, mostly because of increased amounts of palmitic and stearic unsaturated acids. Administration of exogenous palmitate is lethal to ERBB2-positive but not to ERBB2-negative cells. GW9662 exacerbates the effects of palmitate addition on BT474 and MDA-MB-361 cells, but it has no significant effect on MCF-7 and human mammary epithelial cells. Palmitate administration results in a fivefold to tenfold greater increase in fat stores in ERBB2-negative cells compared with ERBB2-positive cells, which suggests that the ERBB2-positive cells have maximized their ability to store fats and that additional palmitate is toxic to these cells. Both PPARγ inhibition and palmitate administration result in increased reactive oxygen species production in BT474 cells. The cell death that results from this treatment can be counteracted by the antioxidant N-acetyl cysteine.

Conclusions

Our findings indicate that PPARγ activity enables ERBB2-positive breast cancer cells, which produce high levels of fat, to convert fatty acids to triglycerides, allowing these cells to avert the cell death that results from lipotoxicity. Endogenous palmitate toxicity represents a genetically based property of ERBB2-positive breast cancer that can be exploited for therapeutic intervention.
Common genetic control of haemangioblast and cardiac development in zebrafish
Authors: Peterkin Tessa , Gibson Abigail , Patient Roger
Published: March 2009

Over the past few years it has become clear that over half of the mammalian heart derives from outside the heart field as originally defined. Such a second heart field, however, has not been described in zebrafish, which could explain its smaller, two-chambered heart. Instead, zebrafish have a population of haemangioblasts, which is absent in mammalian embryos, raising the possibility that these cells represent the evolutionary ancestor of the second heart field. Here, we show for the first time that the genetic programmes of these anterior haemangioblasts and the adjacent heart field are co-regulated,by transcription factors previously associated with heart but not blood or endothelial development. We demonstrate that gata4, gata5 and gata6 are essential for anterior haemangioblast specification, and for subsequent myelopoiesis, acting as early as cloche and upstream of scl. The requirement for gata4, gata5 and gata6in myeloid, endothelial and cardiac specification is in the mesoderm, but these factors also control, from within the endoderm and the yolk syncytial layer, the migration of the cardiac precursors as they differentiate. This genetic link between the blood/endothelial and cardiac programmes supports the notion that this haemangioblast population in zebrafish is an evolutionary antecedent of the second heart field, and has implications for the differentiation of haemangioblasts and cardiomyocytes from pluripotent cells,and for the origins of stem cells in the adult heart.
Electron-Capture Dissociation and Collision-Induced Dissociation of Lanthanide Metal–Ligand Complexes and Lanthanide Metal–Ligand Complexes Bound to Phosphopeptides
Authors: Mosely Jackie A. , Murray Benjamin S. , Parker David
Published: March 2009

Collision-induced dissociation (CID) and electron-capture dissociation (ECD) of lanthanide metal(III)–ligand complexes and lanthanide metal(III)–ligand–phosphopeptide complexes have been investigated using a Fourier transform-ion cyclotron resonance mass spectrometer (FT-ICR MS). Ternary adducts were formed for [LnL3+ + solvent anion(s) n–](3– n)+ [Ln = europium, terbium and ytterbium, L = heptadentate ligand, solvent anion = acetate or trifluoromethane-sulphonate (triflate)]. Results show that ECD provides much more diagnostically useful information than CID. ECD was found to systematically cleave N–C bonds in the “arms” of the ligand, similar to the N–Cα cleavage of peptides, generating two fragmentation sites per arm of the ligand. The most intense and informative fragment ions were obtained from the terbium complex and it is believed that this is a consequence of terbium's greater reduction potential: the greater the reduction potential, the greater the ligand fragmentation; the lower the reduction potential, the more likely the molecule is to relinquish the solvent anion. The choice of solvent is also shown to be important. In general, the complexes studied fragment easily to lose CH3CO2H; however, particularly for ECD, the complexes retain the triflate anion causing the ligand to fragment instead, thus providing much more useful information. The triflate anion can be displaced by a phosphopeptide to create a lanthanide metal–ligand–phosphopeptide adduct. ECD is able to sequence the phosphopeptide, locating the site of phosphorylation bound to [LnL]3+ and also confirm the identity of the ligand. Small differences in the fragmentation of the lanthanide metal–ligand–phosphopeptide adducts follow the trend Eu < Tb < Yb suggesting that charge density may now be a more significant factor than metal ion reduction potential. ECD analysis of the triflate salts of the terbium complexes is most informative in developing a method to optimise structural information that can be gained from this group of molecules by mass spectrometry.
Laser Desorption/Ablation Plumes from Capillary-Like Restricted Volumes
Authors: Knochenmuss Richard
Published: March 2009

Laser desorption/ionization from structured surfaces has been the object of recently renewed interest. Conditions in the plume of material ablated from such surfaces may differ from those of a sample which is ablated in bulk. Since recombination and secondary ion–molecule reactions in the plume play a major role in determining the types and quantities of ions observed at the detector, these differences are analytically relevant. Desorption/ionization substrates with channels of high aspect ratio are modeled as capillary nozzles, from which free jets are emitted. A previously developed matrix-assisted laser desorption/ionization ablation/ionization model is adapted for these jets. More primary ions reach the detector when ablated from a capillary orifice, but fewer analye ions are created in secondary reactions. These differences in ion yield can persist for arrays of capillaries on the surface, depending on the ratio of their diameter to spacing.
Determination of the Binding Energies in Aromatic Clusters: Resonance-Enhanced Multi-Photon Ionization and Mass Analyzed Threshold Ionization Investigation of the Dichlorobenzene–Argon Complexes
Authors: Gaber Angela , Riese Mikko , Witte Frank , Grotemeyer Jürgen
Published: March 2009

Resonance-enhanced multi-photon ionization (REMPI) and mass-analyzed threshold ionization (MATI) spectroscopic investigations were applied to the van der Waals complexes of the three dichlorobenzene isomers with argon. From the REMPI spectra it is concluded that the argon atom is shifted towards the chlorine atoms during excitation for the ortho and the meta isomers while it stays in the middle of the ring for the para isomer. From the MATI spectra it was possible to determine the binding energies in the ion ground state to 617 cm−1 ± 15 cm−1, 529 cm−1 ± 125 cm−1 and 581 cm−1 ± 76 cm−1 for the para, the meta and the ortho isomer, respectively. Together with theoretical calculations binding energies in the neutral ground state were determined to be 426 cm−1 ± 16 cm−1 for all the three isomers.
A Model for Trade-Up and Change in Considered Brands
Authors: Allenby Greg M. , Garratt Mark J. , Rossi Peter E.
Published: March 2009

A common theme in marketing literature is the acquisition and retention of customers as they trade up from inexpensive introductory offerings to those of higher quality. We develop a nonhomothetic choice model to accommodate effects of advertising, professional recommendation, and other factors that facilitate the description and management of trade-up. Our model allows advertising to affect the relative superiority or inferiority of products. This allows for a wide variety of trade-up patterns beyond those obtained from a standard random utility formulation of the logit model. Our nonhomothetic model allows for advertising to affect more than just brand intercepts (perceived quality), but also the rate at which consumers are willing to trade up to higher-quality brands. Advertising effects are measured using a randomized treatment and evaluated by considering their direct implications for firm pricing and profits.
Creep Crack Growth of Alloy 800H in Controlled-Impurity Helium
Authors: Foulds JR
Published: March 2009

The creep crack growth behavior of Alloy 800H has been characterized at temperatures of 677C (1250F) and 732C (1350F) in controlled-impurity helium. Cold-worked material in various recrystallized conditions, expected to be encountered in elevated-temperature service, was tested in center-cracked tension specimen form under constant load. The tests were conducted under dominantly steady-state conditions where the estimated transition time to steady-state creep for each test is a small fraction of the total test time. The results indicate that the crack-tip field parameter, C*, as was experimentally measured, is a good descriptive parameter for the steady-state creep crack growth rate of this material. In fact, the crack growth rate, da/dt, versus C* data from all five tests spanning three different material conditions, two temperatures, and three orders of magnitude of crack growth rates appeared to consolidate into a single straight line on a logarithmic plot. The metallographic results show that crack extension occurs dominantly by the intergranular creep camage mechanisms anticipated in service.