bluuebunny/crossref_metadata_2025 huggingface dataset:
Distinct promoters regulate tissue‐specific and differential expression of kallikrein 6 in CNS demyelinating disease
Authors: Christophi G. P. , Isackson P. J. , Blaber S. , Blaber M. , Rodriguez M. , Scarisbrick I. A.
Published: December 2004

Kallikrein 6 is a serine protease expressed abundantly in normal adult human and rodent CNS, and therein is regulated by injury. In the case of CNS demyelinating disease, K6 expression in CNS occurs additionally in perivascular and parenchymal inflammatory cells suggesting a role in pathogenesis. Herein we describe two unique transcripts that occur within the human and mouse K6 genes that differ in their 5′‐untranslated regions. These transcripts have identical translation initiation sites in exon 3, are expressed in a tissue‐specific fashion and are differentially regulated in response to CNS injury. While the human and mouse 5′‐transcripts differ in sequence they are identical in genomic organization and tissue‐specific expression. The most 5′‐transcript, designated transcript 1, includes exon 1–7, and was detectable in all CNS regions, but not in any non‐CNS tissues examined (spleen, thymus, liver, kidney, pancreas, submandibular gland and peripheral nerve). In contrast, transcript 2 lacks exon 1, but contains a unique sequence at the 5′‐end of exon 2, designated exon 2A. Transcript 2 was expressed both in CNS and in each peripheral tissue. In a murine model of human CNS demyelinating inflammatory disease induced by Theiler's picornovirus, mouse K6 transcript 1 was up‐regulated in brain and spinal cord at acute and more chronic phases of CNS inflammation and demyelination, while overall transcript 2 expression was not significantly altered. However, in isolated splenocyte cultures, transcript 2 was up‐regulated two‐fold by cellular activation. Tissue‐specific expression patterns and differential regulation in CNS disease indicates that each K6 5′‐transcript is probably regulated by unique promoter elements and may serve as a molecular target to treat inflammatory demyelinating disease.
The response of synaptophysin and microtubule‐associated protein 1 to restraint stress in rat hippocampus and its modulation by venlafaxine
Authors: Xu Haiyun , He Jue , Richardson J. Steven , Li Xin‐Min
Published: December 2004

As part of our continuing study of neural plasticity in rat hippocampus, we examined two structural proteins involved in neuronal plasticity, synaptophysin (SYP) and microtubule‐associated protein 1 (MAP1) for their response to repeated restraint stress and modulation of such response by the antidepressant drug venlafaxine. This drug has the pharmacological action of inhibiting the reuptake of serotonin and norepinephrine in nerve terminals. We subjected the rats to restraint stress for 4 h per day for three days, and then injected the animals intraperitoneally (i.p.) with vehicle or 5 mg/kg/day of venlafaxine for various time periods. In all, eight groups of 10 rats each were used. The expression of these two proteins in hippocampal tissue of the rats was examined by means of western blot and immunohistochemical staining techniques. We found that restraint stress decreased the expression of SYP in the rat hippocampus by 50% (p < 0.01), and increased the expression of MAP1 by 60% (p < 0.01). SYP returned to the pre‐stress levels in three weeks and MAP1 in two weeks. In animals treated with venlafaxine post‐stress, SYP returned to pre‐stress levels after 2 weeks and MAP1 after 1 week. These findings enhance our understanding of the compromise of the hippocampus by stressful assaults, and may be relevant to the action of venlafaxine in the treatment of patients with major depression, a mental disease thought to be related to the mal‐adaptation of subjects to environmental stressors.
Interaction of human and mouse Aβ peptides
Authors: Fung Justin , Frost David , Chakrabartty Avijit , McLaurin JoAnne
Published: December 2004

Transgenic mice over‐expressing mutant human amyloid precursor protein have become an important tool for research on Alzheimer's disease (AD) and, in particular, for therapeutic screening. Many models have reported formation of amyloid plaques with age as is detected in AD. However, the plaques generated in transgenic mice are more soluble than human plaques. Differences in solubility may occur for a number of reasons; one proposal is the presence of murine Aβ peptides within the CNS milieu. Here, we report the interaction of human and murine Aβ peptides, Aβ40 and Aβ42, utilizing a fluorescence assay to monitor formation of mixed pre‐fibrillar aggregates, electron microscopy to examine morphological characteristics and detergent solubility to monitor stability. Our results demonstrate that interspecies Aβ aggregates and fibres are readily formed and are more stable than homogenous human fibres. Furthermore, these results suggest that the presence of endogenous murine Aβ in human APP transgenic mice does not account for the increased solubility of plaques.
Effects of neonatal ventral hippocampal lesion in rats on stress‐induced acetylcholine release in the prefrontal cortex
Authors: Laplante François , Stevenson Carl W. , Gratton Alain , Srivastava Lalit K. , Quirion Rémi
Published: December 2004

Excitotoxic neonatal ventral hippocampus (NVH) lesions in rats result in characteristic post‐pubertal hyper‐responsiveness to stress and cognitive abnormalities analogous to those described in schizophrenia and suggestive of alterations in dopamine (DA) neurotransmission. Converging lines of evidence also point to dysfunctions in the cortical cholinergic system in neuropsychiatric disorders. In previous studies, we observed alterations in dopaminergic modulation of acetylcholine (Ach) release in the prefrontal cortex (PFC) in post‐pubertal NVH‐lesioned rats. These two neurotransmitter systems are involved in the stress response as PFC release of DA and Ach is enhanced in response to some stressful stimuli. As adult NVH‐lesioned rats are behaviorally more reactive to stress, we investigated the effects of NVH lesions on tail‐pinch stress‐induced Ach and DA release in the PFC. Using in vivo microdialysis, we observed that tail‐pinch stress resulted in significantly greater increases in prefrontal cortical Ach release in post‐pubertal NVH‐lesioned rats (220% baseline) compared with sham‐operated controls (135% baseline). Systemic administration of the D1‐like receptor antagonist SCH 23390 (0.5 mg/kg i.p.) or the D2‐like receptor antagonist haloperidol (0.2 mg/kg i.p.), as well as intra‐PFC administration of the D2‐like antagonist sulpiride (100 µm), reduced stress‐induced Ach release in PFC of adult NVH‐lesioned rats. By contrast, intra‐PFC administration of SCH 23390 (100 µm) failed to affect stress‐induced Ach release in PFC of NVH‐lesioned rats. Interestingly, using in vivo voltammetry, stress‐induced stimulation of PFC DA release was found to be attenuated in adult NVH‐lesioned rats. Taken together, these data suggest developmentally specific reorganization of prefrontal cortical cholinergic innervation notably regarding its regulation by DA neurotransmission.
Calcitonin gene‐related peptide regulates gene transcription in primary afferent neurons
Authors: Anderson L. E. , Seybold V. S.
Published: December 2004

Although primary afferent neurons express receptors for calcitonin gene‐related peptide (CGRP), understanding of the cellular effects of these receptors is limited. We determined that CGRP receptors regulate gene transcription in primary afferent neurons through a cyclic AMP (cAMP)‐dependent pathway. CGRP increased cAMP in neonatal dorsal root ganglion (DRG) neurons in a concentration‐dependent manner that was blocked by the receptor antagonist CGRP8‐37. The response to CGRP also occurred in adult DRG cells. In contrast, CGRP did not alter the concentration of free intracellular calcium in neonatal or adult DRG neurons. Immunohistochemical data showed that one downstream effect of the cAMP signaling pathway was phosphorylation of cAMP response element binding (CREB) protein, suggesting that CGRP regulates gene expression. This interpretation was supported by evidence that CGRP increased CRE‐dependent gene transcription in neurons transiently transfected with a CRE‐luciferase DNA reporter construct. The effect of CGRP on gene transcription was inhibited by H89, myristoylated‐protein kinase A inhibitor14−22‐amide and U0126, indicating that protein kinase A and mitogen‐activated protein kinase/extracellular receptor kinase kinase are enzymes that mediate effects of CGRP on gene transcription. Therefore, CGRP receptors may regulate expression of proteins by primary afferent neurons during development and in response to tissue‐damaging stimuli.
RAPID COMMUNICATION: Blockade of nociceptin/orphanin FQ transmission in rat substantia nigra reverses haloperidol‐induced akinesia and normalizes nigral glutamate release
Authors: Marti Matteo , Mela Flora , Guerrini Remo , Calò Girolamo , Bianchi Clementina , Morari Michele
Published: December 2004

We recently showed that pharmacological blockade of nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptors located in the substantia nigra stimulates the nigrostriatal dopaminergic pathway and motor behavior (Marti et al. J. Neurosci. 2004, 24, 6659–6666). To investigate whether such motor‐stimulating action was dependent on functional dopaminergic transmission, the selective NOP receptor peptide antagonist [Nphe1,Arg14,Lys15]N/OFQ‐NH2 (UFP‐101) was microinjected into the substantia nigra reticulata of rats made cataleptic by systemic haloperidol administration. UFP‐101 reduced haloperidol‐induced akinesia as measured by immobility time in the bar test. UFP‐101 also induced contralateral turning in cataleptic rats. To investigate the mechanisms involved in the anti‐akinetic action of UFP‐101, nigral glutamate release was monitored by microdialysis technique. The anti‐akinetic action of UFP‐101 correlated with normalization of nigral glutamate release, previously elevated by haloperidol injection. We conclude that endogenous N/OFQ in the substantia nigra sustains akinesia generated by impaired DA transmission and subthalamic nucleus overactivation. NOP receptor antagonists may be beneficial in the symptomatic therapy of parkinsonism, via normalization of subthalamonigral glutamatergic transmission.
Botulinum neurotoxin type D enables cytosolic delivery of enzymatically active cargo proteins to neurones via unfolded translocation intermediates
Authors: Bade Steffen , Rummel Andreas , Reisinger Clemens , Karnath Tino , Ahnert‐Hilger Gudrun , Bigalke Hans , Binz Thomas
Published: December 2004

Multi‐domain bacterial protein toxins are being explored as potential carriers for targeted delivery of biomolecules. Previous approaches employing isolated receptor binding subunits disallow entry into the cytosol. Strategies in which catalytic domains are replaced with cargo molecules are presumably inefficient due to co‐operation of domains during the endosomal translocation step. Here, we characterize a novel transport vehicle in which cargo proteins are attached to the amino terminus of the full‐length botulinum neurotoxin type D (BoNT/D). The intrinsic enzymatic activity of the neurotoxin allowed quantification of the efficacy of cargo delivery to the cytosol. Dihydrofolate reductase and BoNT type A (BoNT/A) light chain (LC) were efficiently conveyed into the cytosol, whereas attachment of firefly luciferase or green fluorescent protein drastically reduced the toxicity. Luciferase and BoNT/A LC retained their catalytic activity as evidenced by luciferin conversion or SNAP‐25 hydrolysis in the cytosol of synaptosomes, respectively. Conformationally stabilized dihydrofolate reductase as cargo considerably decreased the toxicity indicative for the requirement of partial unfolding of cargo protein and catalytic domain as prerequisite for efficient translocation across the endosomal membrane. Thus, enzymatically inactive clostridial neurotoxins may serve as effective, safe carriers for delivering proteins in functionally active form to the cytosol of neurones.
STAT6 transcription factor binding sites with mismatches within the canonical 5′‐TTC…GAA‐3′ motif involved in regulation of δ‐ and µ‐opioid receptors
Authors: Börner Christine , Wöltje Michael , Höllt Volker , Kraus Jürgen
Published: December 2004

Opioid receptors are expressed in neuronal and immune cells and regulated in response to immunological processes. Herein, we demonstrate up‐regulation of the δ‐opioid receptor gene by interleukin‐4 in immune cells (primary T and polymorphonuclear leukocytes, Jurkat E6 T cells), and in NG 108–15 neuronal cells. We identified an interleukin‐4‐responsive element at nt −671 on the murine gene promoter, to which the transcription factor STAT6 binds, as shown by reporter gene analysis and STAT6/DNA interaction studies in living cells with transcription factor decoy oligonucleotides. STAT6 normally binds to palindromic DNA motifs with a 5′‐TTC…GAA‐3′ core. Notably, the δ‐opioid receptor STAT6 site (5′‐TTC…GGA‐3′) is an imperfect palindrome with a mismatch within this core sequence. A systematic analysis of possible mismatch 5′‐TTC…GAA‐3′ motifs revealed that STAT6 also binds to the sequence 5′‐TTA…GAA‐3′. This motif occurs as a polymorphism in the human µ‐opioid receptor gene (Kraus et al. 2001 J. Biol. Chem 276, 43901–43908). We show that this mutated element has a significantly reduced STAT6 binding activity which correlates to its reduced interleukin (IL)‐4 inducibility. In contrast, the non‐canonical STAT6 site of the δ‐opioid receptor binds STAT6 with similar high activity as a perfectly palindromic STAT6 site and is strongly inducible by IL‐4.
Expression and regulation of cyclooxygenase‐2 in normal and neoplastic canine keratinocytes
Authors: Pronovost N. , Suter M. M. , Mueller E. , Sirois J. , Doré M.
Published: December 2004

Squamous cell carcinoma (SCC) is one of the most common cancers in dogs, yet relatively little is known about the molecular events involved in its development. Increasing evidence implicates cyclooxygenase‐2 (COX‐2) in the pathogenesis of various cancers in humans and animals. COX‐2 overexpression has recently been demonstrated in canine SCCs. The objective of our study was to characterize the expression and regulation of COX‐2 in normal and neoplastic canine keratinocytes (CKs) to provide anin vitrosystem to investigate the implication of COX‐2 in SCC oncogenesis in dogs. Cell lines derived from normal CKs and neoplastic CKs (SCCs) were cultured in the absence or presence of agonists, and immunoblots, immunocytochemistry, radioimmunoassays and a cell proliferation assay were used to characterize COX‐2 expression and action. Results showed that neoplastic keratinocytes had a higher basal COX‐2 expression than normal keratinocytes. In both cell lines, stimulation with the tumour promoter phorbol 12‐myristate 13‐acetate induced a time‐dependent increase in COX‐2 protein, with COX‐2 induction being stronger in cancerous SCC than in normal CK cells. Moreover, SCC cells produced significantly more PGE2than CK cells, under both baseline and stimulated conditions (P < 0.05). NS‐398, a selective COX‐2 inhibitor, inhibited prostaglandin (PG)E2synthesis and decreased proliferation of CK and SCC cells (P < 0.05). Collectively, our results indicate that the canine neoplastic keratinocyte SCC cell line expresses more COX‐2 and produces more PGE2than the normal keratinocyte CK cell line, thus providing anin vitrosystem to study the molecular basis of elevated COX‐2 expression in SCCs in dogs.
Short‐Term Indwelling Catheters (A Systematic Review): Evidence for a Primarily Nursing Decision
Authors: Brosnahan Jane Elizabeth , Kent Bridie
Published: December 2004

Program Overview: Urinary tract infection is the most common infection acquired by patients in hospital. The major associated cause is the presence of indwelling urinary catheters. A variety of specialised urethral catheters have been specifically designed to reduce the risk of infection. These include antiseptic impregnated catheters and antibiotic impregnated catheters. Catheter choice or selection is primarily a nursing decision, and nurses also take a leading role in decision‐making regarding catheters purchased for hospitals.

Methods: A systematic review (including metanalyses) was completed with the Cochrane Incontinence Review Group. Eighteen studies met the inclusion criteria: antiseptic impregnated catheters versus standard catheters, which included silver oxide (n = 3 trials) and silver alloy (n = 8 trials), antibiotic impregnated catheters versus standard catheters (n = 1 trial), and comparisons of different types of standard catheters (n = 6 trials).

Results: Silver oxide catheters were not found to prevent bacteriuria in short‐term catheterised hospitalised adults (RR 0.87, 95% CI 0.68 to 1.13). Silver alloy catheters were found to significantly reduce the incidence of asymptomatic bacteriuria (RR 0.36, 95% CI 0.25 to 0.52.) in hospitalised adults catheterised for less than 1 week. At greater than 1‐week catheterization, the risk of asymptomatic bacteriuria was still reduced with the use of silver alloy catheters (RR 0.67, 95% CI 0.50 to 0.90). The risk of symptomatic urinary tract infection was also found to be reduced with the use of silver alloy catheters (RR 0.60, 95% CI 0.50 to 0.73). Silver alloy indwelling catheters appear to reduce the risk of catheter‐acquired urinary tract infection in hospitalised adults catheterised short term. Cost‐benefit analyses indicate that compensation for increased cost of silver alloy catheters would come from the reduction in the cost of catheter‐acquired urinary tract infection.

Conclusions: This review provides not only nurses and nurse managers, but also other health professionals with evidence of effectiveness to influence catheter choice.
Identification of Barriers and Facilitators to Utilizing Research in Nursing Practice
Authors: Micevski Vaska , Sarkissian Sonia , Byrne Julie , Smirnis Jill
Published: December 2004

Background: Despite a growing body of literature, utilization of nursing research remains relatively low even within academic health care institutions.

Objective: The purpose of this study was to (1) investigate barriers for utilization of research in nursing practice in multiple nursing roles (staff nurses and nursing leadership) within an academic health care setting, and (2) identify strategies that may influence nurses' perceptions to utilizing research in nursing practice.

Design: This study was a prospective, descriptive, cross‐sectional study.

Population, Sample, Setting: The subjects were 181 nurses employed at a tertiary care hospital in Southern Ontario, Canada.

Methods: A self‐administered questionnaire identifying demographics, and perceptions of barriers and facilitators to utilizing nursing research in practice was sent to 640 nurses. Results were entered into an SPSS database, and descriptive statistics were used for data analysis. Ethical considerations included approval from the Institutional Ethics Review Board.

Findings: Twenty‐eight percentage of the invited participants responded to the questionnaire. The subjects identified four particular barriers that affected their utilization of research in practice to a great extent: awareness of the research (33%), lack of time to read the research (39%), feeling that the nurse does not have the authority to change practice procedures (41%), and insufficient time on the job to implement new ideas (49%).

Conclusions: Results from this study suggest that a strategic approach to enhancing utilization of nursing research in practice is essential. Organized activities enhance nurses' awareness of research in their field, and developing processes for nurses to participate in practice change is very important. In addition, providing time for nurses to read and implement research will have a positive impact on the utilization of nursing research in practice.
Optimizing parameters of supervised learning techniques (ANN) for precise mapping of the input–output relationship in TMCP steels
Authors: Datta Shubhabrata , Banerjee Malay K.
Published: December 2004

Optimization of artificial neural network architecture and training algorithm is undertaken to map the input–output relationship in thermomechanically processed high‐strength low‐alloy steels. Primarily, the model complexities are varied by varying the number of hidden layers and hidden units. A number of algorithms are tried for training the network. Also, different transfer functions are tried to find the best option. It is found that a four‐layer network with 48 hidden units can perform best in terms of attaining the lowest training error when the network uses hyperbolic tangent transfer function, and is trained with scaled conjugate gradient algorithm or Levenberg–Marquardt algorithm.
Kinetic modelling of chalcocite particle oxidation
Authors: Mihajlovic Ivan , Strbac Nada , Zivkovic Zivan
Published: December 2004

This paper presents the results of experimental investigations of chalcocite (Cu2S) oxidation process in an air atmosphere. X‐ray and chemical analysis were used for phase composition determination of the starting sample. The kinetics of the process was defined by the formula derived regarding simultaneous heat and mass transfer to a reacting front in a spherical particle, where oxidation takes place. The following kinetic model including the effect of the above chosen parameter was established:
image
The activation molar energy of the oxidation process was found to be 129 kJ/mol for the temperature interval 400–525°C.
Statistical analysis in Turkish chromite mining
Authors: Bayat Oktay , Arslan Volkan
Published: December 2004

Many of the techniques developed by mathematical statisticians for the analysis of data are used in the control of product quality. Statistical Process Control (SPC) may be used to cover all uses of statistical techniques for this purpose. However, it often relates particularly to four separate but related techniques that constitute the most common working statistical tools in quality control. One important tool in statistical quality control is the Shewhart control chart. Control schemes are the most widely known and used methods among SPC methods. They can be used to determine whether the differences in quality resulted from coincidences of operation conditions or from artificial reasons for which the source can be determined. The aim of this study is the determination of the lower, upper and central control limits of chromite concentrates obtained from Mikromaden Co., Akmetal Co. and Pinar Mining Co. in the Aladag mountain region of Adana, in Turkey.
Abrupt Temperature Changes in the Western Mediterranean over the Past 250,000 Years
Authors: Martrat Belen , Grimalt Joan O. , Lopez-Martinez Constancia , Cacho Isabel , Sierro Francisco J. , Flores Jose Abel , Zahn Rainer , Canals Miquel , Curtis Jason H. , Hodell David A.
Published: December 2004

A continuous high-resolution Western Mediterranean sea surface temperature (SST) alkenone record spanning the past 250,000 years shows that abrupt changes were more common at warming than at cooling. During marine isotope stage (MIS) 6, SST oscillated following a stadial-interstadial pattern but at lower intensities and rates of change than in the Dansgaard/Oeschger events of MIS 3. Some of the most prominent events occurred over MISs 5 and 7, after prolonged warm periods of high stability. Climate during the whole period was predominantly maintained in interglacial-interstadial conditions, whereas the duration of stadials was much shorter.
A Survey Method for Characterizing Daily Life Experience: The Day Reconstruction Method
Authors: Kahneman Daniel , Krueger Alan B. , Schkade David A. , Schwarz Norbert , Stone Arthur A.
Published: December 2004

The Day Reconstruction Method (DRM) assesses how people spend their time and how they experience the various activities and settings of their lives, combining features of time-budget measurement and experience sampling. Participants systematically reconstruct their activities and experiences of the preceding day with procedures designed to reduce recall biases. The DRM's utility is shown by documenting close correspondences between the DRM reports of 909 employed women and established results from experience sampling. An analysis of the hedonic treadmill shows the DRM's potential for well-being research.
First Atmospheric Science Results from the Mars Exploration Rovers Mini-TES
Authors: Smith Michael D. , Wolff Michael J. , Lemmon Mark T. , Spanovich Nicole , Banfield Don , Budney Charles J. , Clancy R. Todd , Ghosh Amitabha , Landis Geoffrey A. , Smith Peter , Whitney Barbara , Christensen Philip R. , Squyres Steven W.
Published: December 2004

Thermal infrared spectra of the martian atmosphere taken by the Miniature Thermal Emission Spectrometer (Mini-TES) were used to determine the atmospheric temperatures in the planetary boundary layer and the column-integrated optical depth of aerosols. Mini-TES observations show the diurnal variation of the martian boundary layer thermal structure, including a near-surface superadiabatic layer during the afternoon and an inversion layer at night. Upward-looking Mini-TES observations show warm and cool parcels of air moving through the Mini-TES field of view on a time scale of 30 seconds. The retrieved dust optical depth shows a downward trend at both sites.
The Cryptic Filtering House of an Invertebrate Larva
Authors: Pernet Bruno
Published: December 2004

Planktonic larvae of the annelid
Pectinaria californiensis
construct and inhabit cryptic houses through which they filter seawater to concentrate food. Although filtering houses may be unique to pectinariid larvae, other transparent and ephemeral structures are secreted by the larvae of many marine invertebrates and may be involved in the performance of a variety of critical larval functions.
Atmospheric Imaging Results from the Mars Exploration Rovers: Spirit and Opportunity
Authors: Lemmon M. T. , Wolff M. J. , Smith M. D. , Clancy R. T. , Banfield D. , Landis G. A. , Ghosh A. , Smith P. H. , Spanovich N. , Whitney B. , Whelley P. , Greeley R. , Thompson S. , Bell J. F. , Squyres S. W.
Published: December 2004

A visible atmospheric optical depth of 0.9 was measured by the Spirit rover at Gusev crater and by the Opportunity rover at Meridiani Planum. Optical depth decreased by about 0.6 to 0.7% per sol through both 90-sol primary missions. The vertical distribution of atmospheric dust at Gusev crater was consistent with uniform mixing, with a measured scale height of 11.56 ± 0.62 kilometers. The dust's cross section weighted mean radius was 1.47 ± 0.21 micrometers (μm) at Gusev and 1.52 ± 0.18 μ at Meridiani. Comparison of visible optical depths with 9-μ optical depths shows a visible-to-infrared optical depth ratio of 2.0 ± 0.2 for comparison with previous monitoring of infrared optical depths.
Getting Things Right
Authors: Adolphs Ralph , Woodward James
Published: December 2004

The Physiology of Truth
Neuroscience and Human Knowledge. by Jean-Pierre Changeux, translated by M. B. DeBevoise. Harvard University Press, Cambridge, MA, 2004. 324 pp. $45, £29.95. ISBN 0-674-01283-6.

Writing for both fellow scientists and a broad lay audience, the author argues that scientific progress in the neurosciences provides the grounds for concluding that we can know the world as it really is.
The Opportunity Rover's Athena Science Investigation at Meridiani Planum, Mars
Authors: Squyres S. W. , Arvidson R. E. , Bell J. F. , Brückner J. , Cabrol N. A. , Calvin W. , Carr M. H. , Christensen P. R. , Clark B. C. , Crumpler L. , Marais D. J. Des , d'Uston C. , Economou T. , Farmer J. , Farrand W. , Folkner W. , Golombek M. , Gorevan S. , Grant J. A. , Greeley R. , Grotzinger J. , Haskin L. , Herkenhoff K. E. , Hviid S. , Johnson J. , Klingelhöfer G. , Knoll A. H. , Landis G. , Lemmon M. , Li R. , Madsen M. B. , Malin M. C. , McLennan S. M. , McSween H. Y. , Ming D. W. , Moersch J. , Morris R. V. , Parker T. , Rice J. W. , Richter L. , Rieder R. , Sims M. , Smith M. , Smith P. , Soderblom L. A. , Sullivan R. , Wänke H. , Wdowiak T. , Wolff M. , Yen A.
Published: December 2004

The Mars Exploration Rover Opportunity has investigated the landing site in Eagle crater and the nearby plains within Meridiani Planum. The soils consist of fine-grained basaltic sand and a surface lag of hematite-rich spherules, spherule fragments, and other granules. Wind ripples are common. Underlying the thin soil layer, and exposed within small impact craters and troughs, are flat-lying sedimentary rocks. These rocks are finely laminated, are rich in sulfur, and contain abundant sulfate salts. Small-scale cross-lamination in some locations provides evidence for deposition in flowing liquid water. We interpret the rocks to be a mixture of chemical and siliciclastic sediments formed by episodic inundation by shallow surface water, followed by evaporation, exposure, and desiccation. Hematite-rich spherules are embedded in the rock and eroding from them. We interpret these spherules to be concretions formed by postdepositional diagenesis, again involving liquid water.
Stressed for Success
Authors: Garman Bryan
Published: December 2004

School of Dreams
Making the Grade at a Top American High School. by Edward Humes. Harcourt, New York, 2004. 400 pp. $25. ISBN 0-15-100703-9. Paper, $14. ISBN 0-15-603007-1.

The tensions between parental expectations and students' goals and between teaching test-taking and fostering learning provide focal points of an embedded journalist's chronicle of a year in an academically rigorous public high school.
Specialized Persister Cells and the Mechanism of Multidrug Tolerance in
Escherichia coli
Authors: Keren Iris , Shah Devang , Spoering Amy , Kaldalu Niilo , Lewis Kim
Published: December 2004

Bacterial populations produce persisters, cells that neither grow nor die in the presence of bactericidal agents, and thus exhibit multidrug tolerance (MDT). The mechanisms of MDT and the nature of persisters have remained elusive. Our previous research has shown that persisters are largely responsible for the recalcitrance of biofilm infections. A general method for isolating persisters was developed, based on lysis of regular cells by ampicillin. A gene expression profile of persisters contained toxin-antitoxin (TA) modules and other genes that can block important cellular functions such as translation. Bactericidal antibiotics kill cells by corrupting the target function (for example, aminoglycosides interrupt translation, producing toxic peptides). We reasoned that inhibition of translation will lead to a shutdown of cellular functions, preventing antibiotics from corrupting their targets, giving rise to MDT persister cells. Overproduction of the RelE toxin, an inhibitor of translation, caused a sharp increase in persisters. Functional expression of a putative HipA toxin also increased persisters, while deletion of the
hipBA
module caused a sharp decrease in persisters in both stationary and biofilm populations. HipA is thus the first validated persister-MDT gene. We suggest that random fluctuation in the levels of MDT proteins leads to the formation of rare persister cells. The function of these specialized dormant cells is to ensure the survival of the population in the presence of lethal factors.
Evolutionary Genetics of the Capsular Locus of Serogroup 6 Pneumococci
Authors: Mavroidi Angeliki , Godoy Daniel , Aanensen David M. , Robinson D. Ashley , Hollingshead Susan K. , Spratt Brian G.
Published: December 2004

The evolution of the capsular biosynthetic (
cps
) locus of serogroup 6
Streptococcus pneumoniae
was investigated by analyzing sequence variation within three serotype-specific
cps
genes from 102 serotype 6A and 6B isolates. Sequence variation within these
cps
genes was related to the genetic relatedness of the isolates, determined by multilocus sequence typing, and to the inferred patterns of recent evolutionary descent, explored using the eBURST algorithm. The serotype-specific
cps
genes had a low percent G+C, and there was a low level of sequence diversity in this region among serotype 6A and 6B isolates. There was also little sequence divergence between these serotypes, suggesting a single introduction of an ancestral
cps
sequence, followed by slight divergence to create serotypes 6A and 6B. A minority of serotype 6B isolates had
cps
sequences (class 2 sequences) that were ∼5% divergent from those of other serotype 6B isolates (class 1 sequences) and which may have arisen by a second, more recent introduction from a related but distinct source. Expression of a serotype 6A or 6B capsule correlated perfectly with a single nonsynonymous polymorphism within
wciP
, the rhamnosyl transferase gene. In addition to ample evidence of the horizontal transfer of the serotype 6A and 6B
cps
locus into unrelated lineages, there was evidence for relatively frequent changes from serotype 6A to 6B, and vice versa, among very closely related isolates and examples of recent recombinational events between class 1 and 2
cps
serogroup 6 sequences.
Characterization of LytA-LikeN-Acetylmuramoyl-l-Alanine Amidases from Two NewStreptococcus mitisBacteriophages Provides Insights into the Properties of the Major Pneumococcal Autolysin
Authors: Romero Patricia , López Rubens , García Ernesto
Published: December 2004

Two new temperate bacteriophages exhibiting aMyoviridae(φB6) and aSiphoviridae(φHER) morphology have been isolated fromStreptococcus mitisstrains B6 and HER 1055, respectively, and partially characterized. The lytic phage genes were overexpressed inEscherichia coli, and their encoded proteins were purified. ThelytAHERandlytAB6genes are very similar (87% identity) and appeared to belong to the group of the so-called typical LytA amidases (atypical LytA displays a characteristic two-amino-acid deletion signature). although they exhibited several differential biochemical properties with respect to the pneumococcal LytA, e.g., they were inhibited in vitro by sodium deoxycholate and showed a more acidic pH for optimal activity. However, and in sharp contrast with the pneumococcal LytA, a short dialysis of LytAHERor LytAB6resulted in reversible deconversion to the low-activity state (E-form) of the fully active phage amidases (C-form). Comparison of the amino acid sequences of LytAHERand LytAB6with that of the pneumococcal amidase suggested that Val317might be responsible for at least some of the peculiar properties ofS. mitisphage enzymes. Site-directed mutagenesis that changed Val317in the pneumococcal LytA amidase to a Thr residue (characteristic of LytAB6and LytAHER) produced a fully active pneumococcal enzyme that differs from the parental one only in that the mutant amidase can reversibly recover the low-activity E-form upon dialysis. This is the first report showing that a single amino acid residue is involved in the conversion process of the majorS. pneumoniaeautolysin. Our results also showed that some lysogenicS. mitisstrains possess alytA-like gene, something that was previously thought to be exclusive toStreptococcus pneumoniae. Moreover, the newly discovered phage lysins constitute a missing link between the typical and atypical pneumococcal amidases known previously.
Comparative Sequence Analysis of IS50/Tn5Transposase
Authors: Reznikoff William S. , Bordenstein Seth R. , Apodaca Jennifer
Published: December 2004

Comparative sequence analysis of IS50transposase-related protein sequences in conjunction with known structural, biochemical, and genetic data was used to determine domains and residues that play key roles in IS50transposase function. BLAST and ClustalW analyses have been used to find and analyze six complete protein sequences that are related to the IS50transposase. The protein sequence identity of these six homologs ranged from 25 to 55% in comparison to the IS50transposase. Homologous motifs were found associated with each of the three catalytic residues. Residues that play roles in transposase-DNA binding, protein autoregulation, and DNA hairpin formation were also found to be conserved in addition to other residues of unknown function. On the other hand, some homologous sequences did not appear to be competent to encode the inhibitor regulatory protein. The results were also used to compare the IS50transposase with the more distantly related transposase encoded by IS10.
Identification of the
Escherichia coli
K-12
ybhE
Gene as
pgl
, Encoding 6-Phosphogluconolactonase
Authors: Thomason Lynn C. , Court Donald L. , Datta Atin R. , Khanna Rita , Rosner Judah L.
Published: December 2004

We report identification of the
Escherichia coli ybhE
gene as the
pgl
gene that encodes 6-phosphogluconolactonase. A tentative identification was first made based on the known approximate location of the
pgl
gene and the similarity of the presumptive ybhE-encoded protein sequence to a known Pgl enzyme. To test this notion, the
ybhE
gene was deleted and replaced with a drug marker. Like previously characterized
pgl
mutants, the
ybhE
deletion mutant had a Blu
−
phenotype (dark-blue staining with iodine due to accumulation of starch after growth on minimal maltose) and demonstrated impaired growth on minimal glucose medium when combined with a
pgi
mutation. Biochemical assay of crude extracts for 6-phosphogluconolactonase enzymatic activity showed that
ybhE
encodes this activity. The
ybhE
gene was transferred from the
E. coli
chromosome to an expression vector. This
ybhE
clone complemented both the precise deletion of the
ybhE
gene and a larger deletion,
pgl
Δ
8
, for the Blu
−
phenotype and for phosphogluconolactonase activity, confirming that
ybhE
is the
pgl
gene. A newly observed phenotype of
pgl
strains is a lowered frequency of appearance of Bgl
+
mutants that can utilize the β-glucoside salicin. This is likely due to poor growth of Bgl
+
pgl
strains on salicin due to the accumulation of 6-phosphogluconolactone.
In Vivo Bypass of Chaperone by Extended Coiled-Coil Motif in T4 Tail Fiber
Authors: Qu Yun , Hyman Paul , Harrah Timothy , Goldberg Edward
Published: December 2004

The distal-half tail fiber of bacteriophage T4 is made of three gene products: trimeric gp36 and gp37 and monomeric gp35. Chaperone P38 is normally required for folding gp37 peptides into a P37 trimer; however, a temperature-sensitive mutation in T4 (ts3813) that suppresses this requirement at 30°C but not at 42°C was found in gene 37 (R. J. Bishop and W. B. Wood, Virology
72:
244-254, 1976). Sequencing of the temperature-sensitive mutant revealed a 21-bp duplication of wild-type gene 37 inserted into its C-terminal portion (S. Hashemolhosseini et al., J. Mol. Biol.
241:
524-533, 1994). We noticed that the 21-amino-acid segment encompassing this duplication in the ts3813 mutant has a sequence typical of a coiled coil and hypothesized that its extension would relieve the temperature sensitivity of the ts3813 mutation. To test our hypothesis, we crossed the T4 ts3813 mutant with a plasmid encoding an engineered pentaheptad coiled coil. Each of the six mutants that we examined retained two amber mutations in gene 38 and had a different coiled-coil sequence varying from three to five heptads. While the sequences varied, all maintained the heptad-repeating coiled-coil motif and produced plaques at up to 50°C. This finding strongly suggests that the coiled-coil motif is a critical factor in the folding of gp37. The presence of a terminal coiled-coil-like sequence in the tail fiber genes of 17 additional T-even phages implies the conservation of this mechanism. The increased melting temperature should be useful for “clamps” to initiate the folding of trimeric β-helices in vitro and as an in vivo screen to identify, sequence, and characterize trimeric coiled coils.
Phage φ29 Proteins p1 and p17 Are Required for Efficient Binding of Architectural Protein p6 to Viral DNA In Vivo
Authors: González-Huici Víctor , Alcorlo Martín , Salas Margarita , Hermoso José M.
Published: December 2004

Bacteriophage φ29 protein p6 is a viral architectural protein, which binds along the whole linear φ29 DNA in vivo and is involved in initiation of DNA replication and transcription control. Protein p1 is a membrane-associated viral protein, proposed to attach the viral genome to the cell membrane. Protein p17 is involved in pulling φ29 DNA into the cell during the injection process. We have used chromatin immunoprecipitation and real-time PCR to analyze in vivo p6 binding to DNA in cells infected with φ29
sus1
or
sus17
mutants; in both cases p6 binding is significantly decreased all along φ29 DNA. φ29 DNA is topologically constrained in vivo, and p6 binding is highly increased in the presence of novobiocin, a gyrase inhibitor that produces a loss of DNA negative superhelicity. Here we show that, in cells infected with φ29
sus1
or
sus17
mutants, the increase of p6 binding by novobiocin is even higher than in cells containing p1 and p17, alleviating the p6 binding deficiency. Therefore, proteins p1 and p17 could be required to restrain the proper topology of φ29 DNA, which would explain the impaired DNA replication observed in cells infected with
sus1
or
sus17
mutants.
The RpoH-Mediated Stress Response in
Neisseria gonorrhoeae
Is Regulated at the Level of Activity
Authors: Laskos Lina , Ryan Catherine S. , Fyfe Janet A. M. , Davies John K.
Published: December 2004

The general stress response in
Neisseria gonorrhoeae
was investigated. Transcriptional analyses of the genes encoding the molecular chaperones DnaK, DnaJ, and GrpE suggested that they are transcribed from σ
32
(RpoH)-dependent promoters upon exposure to stress. This was confirmed by mutational analysis of the σ
32
promoter of
dnaK
. The gene encoding the gonococcal RpoH sigma factor appears to be essential, as we could not isolate viable mutants. Deletion of an unusually long
rpoH
leader sequence resulted in elevated levels of transcription, suggesting that this region is involved in negative regulation of RpoH expression during normal growth. Transcriptional analyses and protein studies determined that regulation of the RpoH-mediated stress response is different from that observed in most other species, in which regulation occurs predominantly at the transcriptional and translational levels. We suggest that an increase in the activity of preformed RpoH is primarily responsible for induction of the stress response in
N. gonorrhoeae
.
Revising the Role of the Pneumococcalvex-vncRSLocus in Vancomycin Tolerance
Authors: Haas Wolfgang , Sublett Jack , Kaushal Deepak , Tuomanen Elaine I.
Published: December 2004

Vancomycin is used increasingly to treat invasive infections caused by multidrug-resistantStreptococcus pneumoniae. Although no vancomycin-resistant strains have been isolated to date, tolerant strains that fail to die rapidly and that cause relapsing disease have been described. Thevex123-pep27-vncRSlocus, consisting of an ABC transporter, a presumed signaling peptide, and a two-component system, respectively, has been implicated in vancomycin tolerance. Recent findings, however, challenged this model. The data presented here indicate that erythromycin in the growth medium induces a vancomycin-tolerant phenotype and that loss of function of Pep27or VncRS does not alter autolysis. However, a role for the ABC transporter encoded by thevex123genes in tolerance was confirmed. Avex3mutant was considerably more tolerant to vancomycin treatment than the wild-type strain T4, and the strength of the phenotype depended on the orientation of the resistance cassette used to construct the mutant. Microarray results suggested a number of genes that might be involved in tolerance in thevex3mutant. Although the exact function and regulation of thevex123-pep27-vncRSlocus remains to be determined, several factors influence the autolysis behavior ofS. pneumoniae, including the bacterial capsule, erythromycin, and thelytAandvex3gene products.
The Etiological Agent of Lyme Disease,Borrelia burgdorferi, Appears To Contain Only a Few Small RNA Molecules
Authors: Östberg Yngve , Bunikis Ignas , Bergström Sven , Johansson Jörgen
Published: December 2004

Small regulatory RNAs (sRNAs) have recently been shown to be the main controllers of several regulatory pathways. The function of sRNAs depends in many cases on the RNA-binding protein Hfq, especially for sRNAs with an antisense function. In this study, the genome ofBorrelia burgdorferiwas subjected to different searches for sRNAs, including direct homology and comparative genomics searches and ortholog- and annotation-based search strategies. Two new sRNAs were found, one of which showed complementarity to therpoSregion, which it possibly controls by an antisense mechanism. The role of the other sRNA is unknown, although observed complementarities against particular mRNA sequences suggest an antisense mechanism. We suggest that the low level of sRNAs observed inB. burgdorferiis at least partly due to the presumed lack of both functional Hfq protein and RNase E activity.
Regulation of σBby an Anti- and an Anti-Anti-Sigma Factor inStreptomyces coelicolorin Response to Osmotic Stress
Authors: Lee Eun-Jin , Cho You-Hee , Kim Hyo-Sub , Ahn Bo-Eun , Roe Jung-Hye
Published: December 2004

σB, a homolog of stress-responsive σBofBacillus subtilis, controls both osmoprotection and differentiation inStreptomyces coelicolorA3 (2). Its gene is preceded byrsbAandrsbBgenes encoding homologs of an anti-sigma factor, RsbW, and its antagonist, RsbV, ofB. subtilis, respectively. Purified RsbA bound to σBand prevented σB-directed transcription from thesigBp1 promoter in vitro. AnrsbA-null mutant exhibited contrasting behavior to thesigBmutant, with elevatedsigBp1 transcription, no actinorhodin production, and precocious aerial mycelial formation, reflecting enhanced activity of σBin vivo. Despite sequence similarity to RsbV, RsbB lacks the conserved phosphorylatable serine residue and its gene disruption produced no distinct phenotype. RsbV (SCO7325) from a putative six-gene operon (rsbV-rsbR-rsbS-rsbT-rsbU1-rsbU) was strongly induced by osmotic stress in a σB-dependent manner. It antagonized the inhibitory action of RsbA on σB-directed transcription and was phosphorylated by RsbA in vitro. These results support the hypothesis that the rapid induction of σBtarget genes by osmotic stress results from modulation of σBactivity by the kinase-anti-sigma factor RsbA and its phosphorylatable antagonist RsbV, which function by a partner-switching mechanism. Amplified induction could result from a rapid increase in the synthesis of both σBand its inhibitor antagonist.
Interaction between the TolC and AcrA Proteins of a Multidrug Efflux System of
Escherichia coli
Authors: Husain Fasahath , Humbard Matthew , Misra Rajeev
Published: December 2004

This paper provides the biochemical evidence for physical interactions between the outer membrane component, TolC, and the membrane fusion protein component, AcrA, of the major antibiotic efflux pump of
Escherichia coli
. Cross-linking between TolC and AcrA was independent of the presence of any externally added substrate of the efflux pump or of the pump protein, AcrB. The biochemical demonstration of a TolC-AcrA interaction is consistent with genetic studies in which extragenic suppressors of a mutant TolC strain were found in the
acrA
gene.