bluuebunny/crossref_metadata_2025 huggingface dataset:
Cutting the cost: Laparoscopic inguinal hernia repair by totally extraperitoneal approach without disposable instruments
Authors: Chiang David Ting‐Wei , Bohmer Robert
Published: September 2006

Background:  One of the disadvantages of laparoscopic inguinal hernioplasty by the totally extraperitoneal approach is the cost associated with disposable instrumentation that is used to establish and maintain the preperitoneal space. A prospective audit and cost‐analysis of laparoscopic inguinal hernia repair with and without disposable instruments was conducted at the Royal Hobart Hospital, Australia.

Methods:  The data from 94 patients undergoing laparoscopic inguinal hernioplasty (47 with and 47 without disposable instruments) were reviewed. In the group without disposable instruments, digital dissection supplemented by movements of a 30° laparoscope, and followed by dissection with laparoscopic forceps, was used instead of disposable balloon expanders to establish and maintain the preperitoneal space during hernioplasty. Detailed pre‐, intra‐, and postoperative findings were audited for each laparoscopic inguinal hernioplasty technique. In‐hospital cost analysis was conducted for theatre time, and costs of laparoscopic and disposable instruments were compared.

Result:  The overall cost saving of not using disposable instrumentation at the initial stage was almost $A20 000 for 47 patients (approximately $A425 per case). The difference in theatre time used was not statistically significant. In the group of hernioplasty without disposable instruments, disposable balloon expanders were still required in three patients (8.2%) due to gas leakage, and one (2.6%) required conversion to open procedure due to profuse bleeding. There were two patients in this group with postoperative complications: one with seroma and one with urinary retention.

Conclusion:  The preperitoneal space was established and maintained in most patients without using the specially designed balloon expanders. Therefore, this modification of the totally extraperitoneal approach could be considered safe and effective, and should be attempted initially. However, in difficult cases, the disposable balloon expanders may still be required.
Medicare Part D: Practical practice information for healthcare providers
Authors: Moczygemba Leticia R
Published: September 2006

Purpose: To provide an overview of Medicare Part D and useful practice information for healthcare providers to help guide Medicare beneficiaries when choosing a Part D plan.

Data sources: Documents written by the Centers for Medicare & Medicaid Services, Kaiser Family Foundation, and United States Department of Health & Human Services.

Conclusions: Although Medicare Part D can provide assistance with prescription drug costs to Medicare beneficiaries, the transition to Medicare Part D in 2006 will be challenging for both beneficiaries and healthcare providers. It is important that both Medicare beneficiaries and healthcare providers understand the complex drug benefit to facilitate the transition to Medicare Part D.

Implications for practice: It is likely that Medicare Part D formularies will require adjustments to many Medicare beneficiaries’ medication regimens. Additionally, it is possible that there will be an increase in the number of prior authorizations and other requirements such as step therapy requests.
Synthesis and Reactivity of 6,7‐dihydrogeranylazides: Reagents for Primary Azide Incorporation into Peptides and Subsequent Staudinger Ligation
Authors: Xu Juhua , DeGraw Amanda J , Duckworth Benjamin P , Lenevich Stepan , Tann Cheng‐Min , Jenson Emily C , Gruber Simon J , Barany George , Distefano Mark D
Published: September 2006

Protein farnesyltransferase (PFTase) catalyzes the attachment of a geranylazide moiety to a peptide substrate, N‐dansyl‐GCVIA. Because geranylazide is actually a mixture of isomeric, interconverting primary and secondary azides, incorporation of this isoprenoid into peptides can potentially result in a corresponding mixture of prenylated peptides. Here, we first examined the reactivity of geranyl azide in a model Staudinger reaction and determined that a mixture of products is formed. We then describe the synthesis of 6,7‐dihydrogeranylazide diphosphate and demonstrate that this compound allows exclusive incorporation of a primary azide into a peptide. The resulting azide‐containing peptide was derivatized with a triphenylphosphine‐based reagent to generate an O‐alkyl imidate‐linked product. Finally, we show, using a series of model reactions, that the Staudinger ligation frequently produces small amounts of O‐alkyl imidate products in addition to the major amide‐linked products. Thus, the alkoxyimidates we have observed as the exclusive products in the reactions of peptides containing prenylated azides also appear to be a common type of product formed using other azide‐containing reactants, although at greatly reduced levels. This method for chemical modification of the C‐terminus of a protein should be useful for a variety of applications in protein chemistry.
Personal Protective Equipment Use and Safety Behaviors Among Farm Adolescents: Gender Differences and Predictors of Work Practices
Authors: Reed Deborah B. , Browning Steven R. , Westneat Susan C. , Kidd Pamela S.
Published: September 2006

ABSTRACT: Context:Children on farms perform work that places them at risk for acute and chronic negative health outcomes. Despite strategies for preventing and reducing the risk of disease and injury, children’s use of personal protective equipment and safely equipped farm machinery has generally remained unreported.Purpose:This paper reports the use of personal protective equipment, self‐protective work behaviors, and selected risk exposures of children aged 14‐19, who perform farm work.Methods:Survey results of adolescent high school students (n = 593) enrolled in agriculture class in Kentucky, Iowa, and Mississippi. Students were part of the sample that participated in the Agricultural Disability Awareness and Risk Education Project.Findings:Boys were at a significantly higher risk of exposure compared to girls, and boys engaged more frequently in risky behavior. Hearing and respiratory protection was used minimally and sporadically. Physical symptoms influenced use of hearing and respirator use, as did physician recommendation to use such protection. Of students who operated farm tractors, only half most frequently operated tractors with safety bars and seat belts. Sixty percent of the students reported using equipment with damaged or missing safety shields.Conclusions:In addition to the usual risks of farm work, adolescents may be at even greater risk by not using personal protective equipment or not having access to machinery that is properly equipped for maximum protection. Health care providers should incorporate advice to adolescents and their parents on risk reduction, particularly on the use of personal protective equipment.
Swimmer syndrome in a Devon rex kitten and an English bulldog puppy
Authors: Verhoeven G. , De Rooster H. , Risselada M. , Wiemer P. , Scheire L. , Van Bree H.
Published: September 2006

A three‐week‐old Devon rex kitten and a four‐week‐old English bulldog puppy were presented with “swimmer syndrome”. The owners consulted several veterinarians who suggested euthanasia as the only possible solution for this condition. Physiotherapy in the puppy, and physiotherapy and bandaging in the kitten led to the resolution of the clinical signs and resulted in normal ambulation after several weeks. The authors concluded that intensive physiotherapy and dedication of the owner can lead to success.
Meningoencephalitis associated with Staphylococcus warneri in a dog
Authors: Espino L. , Bérmudez R. , Fidalgo L. E. , González A. , Miño N. , Quiroga M. I.
Published: September 2006

A case of meningoencephalitis in a dog caused by Staphylococcus warneri is reported here. The history and clinical signs were suggestive of possible central nervous system infection. Analysis of cerebrospinal fluid documented a neutrophilic pleocytosis (890 cells/μl) and the presence of occasional intracellular cocci. Staphylococcus warneri was isolated from the microbiological culture of the cerebrospinal fluid. Treatment consisted of intravenous antibiotics, supportive care and anticonvulsants for the generalised seizures that developed after admission. Histological assessment confirmed the location and extension of bacterial meningoencephalitis. Thrombotic meningoencephalitis associated with Staphylococcus warneri infection has not, to the authors’ knowledge, been previously reported in dogs.
Dementia with Cerebrovascular Disease
Authors: Wright Clinton B. , Vonsattel Jean Paul G. , Bell Karen , Honig Lawrence S.
Published: September 2006

In this case study, we review the symptoms, cognitive testing, brain imaging, and brain pathology of a woman with dementia, for whom the neuropathological findings suggest a prominent contribution of cerebrovascular disease. Vascular dementia is the term commonly used for persons with dementia resulting from strokes, either clinically evident or subclinical "silent" events. "Mixed dementia" is the term used when there is an admixture of pathological findings related to Alzheimer's disease (AD) and cerebrovascular disease, as in this situation. In some cases of mixed dementia, the pathological involvement of AD may be the principal contributory cause of the cognitive symptoms, and in others, the vascular changes may give the greater contribution.
The Age of Cancer
Authors: Leslie Mitch
Published: September 2006

The biggest risk factor for cancer isn't smoking or bad diet or anything a person can avoid. It's growing old. New findings might help explain the connection between aging and cancer. According to the study, a gene that's faulty in a disorder that resembles accelerated aging shuts down in many cancers.
Death-Bed Prophecy
Authors: Leslie Mitch
Published: September 2006

What's left to learn about aging? The burning question for many researchers is whether life-stretching pathways and genes from model organisms boost human life span. Researchers might be able to track down additional genes and pathways that adjust longevity by studying a broader range of organisms or by tracking the evolution of genes that promote aging. An alternative way to extend our lives might be to identify the genes behind late-life killers such as heart disease and diabetes. Lab animals last longer on a very low-cal diet, and scientists are probing whether humans can benefit from this austerity. Or better yet, perhaps researchers can design molecules that deliver the gain of calorie reduction without the pain. Scientists are also focusing on which parts of the cell incur damage as we age and how growth and reproduction tie in to longevity. The speed of the next round of advances will depend on whether movers and shakers in funding organizations recognize the importance of the research and are willing to pay for it.
The Role of Mitochondria in Conserved Mechanisms of Aging
Authors: Scheckhuber Christian , Osiewacz Heinz D.
Published: September 2006

The European research project MiMage, supported by the European Community's Sixth Framework for Research and Technological Development, focuses on elucidating the role of mitochondria in conserved mechanisms of aging. This Perspective summarizes a selection of talks presented in April 2006 at the second MiMage symposium by members from participating laboratories and invited speakers.
Aging On the Job
Authors: Levine James , Heet Justin , Burlingame Barbara
Published: September 2006

The older population is growing faster than the population as a whole. Paid work is the main mechanism for physical activity in humans. We, therefore, wanted to ascertain whether work practices (such as employment status and the types of tasks workers perform) follow the biological decline in physical activity that occurs with aging in humans and many other species. We recorded work practices in 3352 adults in the Ivory Coast to establish how work burdens are distributed across different age groups in a community that is minimally fettered by legal constraints. We found a decrease in the amount of work performed with increasing age, and we found that elderly persons performed more skilled, less exothermic tasks than younger workers. These data mirror global workforce distribution trends expressed by age. If the trend continues, a major portion of the population will be unlikely to contribute to the labor force in the near future, producing potentially adverse economic consequences in some populations. Although the problem might be averted or at least minimized by implementing different employment policies, biological factors could overlay and supervene any economic planning.
Developing a Research Agenda in Biogerontology: Physiological Systems
Authors: Carrington Jill L. , Bellino Francis L.
Published: September 2006

The Biology of Aging Program (BAP) at the National Institute on Aging supports research in many areas, including processes of cell senescence and apoptosis, genetic influences on aging, and how aging leads to tissue dysfunction. Several approaches to research on aging physiological systems are described, along with BAP programmatic efforts to enhance and support that research. Understanding the relation between aging and tissue dysfunction has led to new insights into how health can be improved for aged individuals.
WRN's Tenth Anniversary
Authors: Hisama Fuki M. , Bohr Vilhelm A. , Oshima Junko
Published: September 2006

Werner syndrome (WS) is a segmental progeroid syndrome in which patients display pleiotropic features of aging seen in the normal population. The advent of positional cloning in the 1990s markedly accelerated the identification of human disease-causing genes. In 1996, mutations in
WRN
, which was shown to encode a new, putative member of the family of RecQ DNA helicases, were identified in four patients as the cause of WS. Ten years after the identification of WRN, what have we learned about its role in WS, and its contribution to normal aging?
A Work in Progress: The Metabolic Syndrome
Authors: Luchsinger Jose A.
Published: September 2006

Metabolic syndrome refers to a constellation of risk factors for cardiovascular disease. They include elevated plasma glucose concentrations, dyslipidemia, hypertension, and abdominal obesity. These conditions typically occur during middle age or later in life. Although there is no clear consensus on the diagnosis of metabolic syndrome, it is a potentially important entity to recognize and manage once traditional cardiovascular risk factors, such as smoking, hypertension, dyslipidemia, and diabetes, have been treated individually. This Perspective summarizes our current knowledge of the metabolic syndrome. Lifestyle change, including diet and exercise, is probably the best available option for treating the metabolic syndrome. However, rigorous lifestyle interventions are difficult to implement outside of a clinical trial setting, especially among elderly patients.
The Unusual Genetics of Human Longevity
Authors: De Benedictis Giovanna , Franceschi Claudio
Published: September 2006

In no species other than humans do cultural, social, and biological factors interact with each other in modulating complex phenotypes. Thus, the identification of genetic factors that affect human longevity is a true challenge. The model of centenarians provides us a unique opportunity to tackle this challenge. In this Perspective, we discuss some recent findings (the impact of geography and demography on the longevity phenotype, the relationship between longevity and homozygosity, the role of the nuclear-mitochondrial genome cross-talk) by which new ideas are suggested, such as the concept of a complex allele timing as a pivotal process in modulating the probability of achieving longevity.
From Bedside to Bench: Does Mental and Physical Activity Promote Cognitive Vitality in Late Life?
Authors: Studenski Stephanie , Carlson Michelle C. , Fillit Howard , Greenough William T. , Kramer Arthur , Rebok George W.
Published: September 2006

A wide range of animal and human studies provide evidence for the potential of physical and cognitive exercise in promoting cognitive health later in life. The effects of such activities on intermediate outcomes, such as cognitive performance, are becoming clearer, as are the molecular mechanisms involved. Physical and cognitive exercise might increase "cognitive reserve" and increase the overall health of the brain, thereby reducing or delaying cognitive impairment and dementia. However, conclusive evidence for such benefits is not yet established. The third annual
Bedside to Bench conference
, cosponsored by
The American Geriatrics Society
and the National Institutes of Health's
National Institute on Aging
, reviewed current knowledge regarding the role of physical and cognitive exercise in promoting cognitive vitality. Conference attendees identified gaps in our current understanding of these processes and recommended next steps for research. In particular, researchers will need to explore clinical issues related to the timing, intensity, and duration of various types and combinations of physical and cognitive activities in animal models to elucidate the mechanisms involved and inform the design of future human studies. The concept of the enriched environment currently employed in animal studies to promote physical activity, socialization, and problem solving should be explored in human studies.
Targeting the Role of the Endosome in the Pathophysiology of Alzheimer's Disease: A Strategy for Treatment
Authors: Tate Barbara A. , Mathews Paul M.
Published: September 2006

Membrane-bound endosomal vesicles play an integral role in multiple cellular events, including protein processing and turnover, and often critically regulate the cell-surface availability of receptors and other plasma membrane proteins in many different cell types. Neurons are no exception, being dependent on endosomal function for housekeeping and synaptic events. Growing evidence suggests a link between neuronal endosomal function and Alzheimer's disease (AD) pathophysiology. Endosomal abnormalities invariably occur within neurons in AD brains, and endocytic compartments are one likely site for the production of the pathogenic β-amyloid peptide (Aβ), which accumulates within the brain during the disease and is generated by proteolytic processing of the amyloid precursor protein (APP). The enzymes and events involved in APP processing are appealing targets for therapeutic agents aimed at slowing or reversing the pathogenesis of AD. The neuronal endosome may well prove to be the intracellular site of action for inhibitors of β-amyloidogenic APP processing. We present here the view that knowledge of the endosomal system in the disease can guide drug discovery of AD therapeutic agents.
I Come Not to Bury SAGE KE But to Appraise It
Authors: Strauss Evelyn
Published: September 2006

This article serves as a eulogy for the Science of Aging Knowledge Environment (SAGE KE). This online resource--
Science
's Web site on aging--is publishing its last issue today. The piece is a personal recollection of co-creating the site--and includes some thoughts on how the field of aging has changed over the last six years.
Volatile Chemical Cues Guide Host Location and Host Selection by Parasitic Plants
Authors: Runyon Justin B. , Mescher Mark C. , De Moraes Consuelo M.
Published: September 2006

The importance of plant volatiles in mediating interactions between plant species is much debated. Here, we demonstrate that the parasitic plant
Cuscuta pentagona
(dodder) uses volatile cues for host location.
Cuscuta pentagona
seedlings exhibit directed growth toward nearby tomato plants (
Lycopersicon esculentum
) and toward extracted tomato-plant volatiles presented in the absence of other cues. Impatiens (
Impatiens wallerana
) and wheat plants (
Triticum aestivum
) also elicit directed growth. Moreover, seedlings can distinguish tomato and wheat volatiles and preferentially grow toward the former. Several individual compounds from tomato and wheat elicit directed growth by
C. pentagona
, whereas one compound from wheat is repellent. These findings provide compelling evidence that volatiles mediate important ecological interactions among plant species.
DNA, Evolution, and the Moral Law
Authors: Pollack Robert
Published: September 2006

The Language of God
. A Scientist Presents Evidence for Belief.
By Francis S. Collins
. Free Press (Simon and Schuster), New York, 2006. 303 pp. $26, C$32.95. ISBN 0-7432-8639-1.

Holding that the principles of faith are complementary with those of science, the author presents his case for the existence of God and of miracles.
A High-Throughput Screening Method to Identify Small Molecule Inhibitors of Thyroid Hormone Receptor Coactivator Binding
Authors: Arnold Leggy A. , Estébanez-Perpiñá Eva , Togashi Marie , Shelat Anang , Ocasio Cory A. , McReynolds Andrea C. , Nguyen Phuong , Baxter John D. , Fletterick Robert J. , Webb Paul , Guy R. Kiplin
Published: September 2006

To provide alternative methods for regulation of gene transcription initiated by the binding of thyroid hormone (T3) to the thyroid receptor (TR), we have developed a high-throughput method for discovering inhibitors of the interaction of TR with its transcriptional coactivators. The screening method is based on fluorescence polarization (FP), one of the most sensitive and robust high-throughput methods for the study of protein-protein interactions. A fluorescently labeled coactivator is excited by polarized light. The emitted polarized light is a function of the molecular properties of the labeled coactivator, especially Brownian molecular rotation, which is very sensitive to changes in the molecular mass of the labeled complex. Dissociation of hormone receptor from fluorescently labeled coactivator peptide in the presence of small molecules can be detected by this competition method, and the assay can be performed in a high-throughput screening format. Hit compounds identified by this method are evaluated by several secondary assay methods, including a dose-response analysis, a semiquantitative glutathione-
S
-transferase assay, and a hormone displacement assay. Subsequent in vitro transcription assays can detect inhibition of thyroid signaling at low micromolar concentrations of small molecules in the presence of T3.
Receptors as Microprocessors: Pharmacological Nuance on Metabotropic Glutamate Receptors 1α
Authors: Kenakin Terry
Published: September 2006

G protein–coupled receptors have revealed themselves to be complex information-processing units that may be exploited for subtle therapeutic signaling effects. Thus, ligands may not only turn receptors on and off, but may also select from their repertoire of signaling effects to further refine drug response.
Touched and Moved by STAT3
Authors: Gao S. Paul , Bromberg Jacqueline F.
Published: September 2006

STAT3, a member of the signal transducer and activator of transcription (STAT) family of transcription factors, is a known regulator of cell motility through its transcriptional activating functions. However, new evidence suggests a novel role for non–tyrosine-phosphorylated and cytoplasmically localized STAT3 in mediating cell migration by disrupting an interaction between microtubules and one of its partners, stathmin. The association of STAT3 with stathmin potentiates microtubule polymerization and cell movement.
Rules for Modeling Signal-Transduction Systems
Authors: Hlavacek William S. , Faeder James R. , Blinov Michael L. , Posner Richard G. , Hucka Michael , Fontana Walter
Published: September 2006

Formalized rules for protein-protein interactions have recently been introduced to represent the binding and enzymatic activities of proteins in cellular signaling. Rules encode an understanding of how a system works in terms of the biomolecules in the system and their possible states and interactions. A set of rules can be as easy to read as a diagrammatic interaction map, but unlike most such maps, rules have precise interpretations. Rules can be processed to automatically generate a mathematical or computational model for a system, which enables explanatory and predictive insights into the system’s behavior. Rules are independent units of a model specification that facilitate model revision. Instead of changing a large number of equations or lines of code, as may be required in the case of a conventional mathematical model, a protein interaction can be introduced or modified simply by adding or changing a single rule that represents the interaction of interest. Rules can be defined and visualized by using graphs, so no specialized training in mathematics or computer science is necessary to create models or to take advantage of the representational precision of rules. Rules can be encoded in a machine-readable format to enable electronic storage and exchange of models, as well as basic knowledge about protein-protein interactions. Here, we review the motivation for rule-based modeling; applications of the approach; and issues that arise in model specification, simulation, and testing. We also discuss rule visualization and exchange and the software available for rule-based modeling.
Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) for the Identification and Analysis of Multiprotein Complexes
Authors: Swamy Mahima , Siegers Gabrielle M. , Minguet Susana , Wollscheid Bernd , Schamel Wolfgang W. A.
Published: September 2006

Multiprotein complexes (MPCs) play crucial roles in cell signaling. Two kinds of MPCs can be distinguished: (i) Constitutive, abundant MPCs--for example, multisubunit receptors or transcription factors; and (ii) signal-induced, transient, low copy number MPCs--for example, complexes that form upon binding of Src-homology 2 (SH2) domain-containing proteins to tyrosine-phosphorylated proteins. Blue native polyacrylamide gel electrophoresis (BN-PAGE) is a separation method with a higher resolution than gel filtration or sucrose density ultracentrifugation that can be used to analyze abundant, stable MPCs from 10 kD to 10 MD. In contrast to immunoprecipitation and two-hybrid approaches, it allows the determination of the size, the relative abundance, and the subunit composition of an MPC. In addition, it shows how many different complexes exist that share a common subunit, whether free monomeric forms of individual subunits exist, and whether these parameters change upon cell stimulation. Here, we give a detailed protocol for the separation of MPCs from total cellular lysates or of prepurified MPCs by one-dimensional BN-PAGE or by two-dimensional BN-PAGE and SDS-PAGE.
Role of Insulin, Adipocyte Hormones, and Nutrient-Sensing Pathways in Regulating Fuel Metabolism and Energy Homeostasis: A Nutritional Perspective of Diabetes, Obesity, and Cancer
Authors: Marshall Stephen
Published: September 2006

Traditionally, nutrients such as glucose and amino acids have been viewed as substrates for the generation of high-energy molecules and as precursors for the biosynthesis of macromolecules. However, it is now apparent that nutrients also function as signaling molecules in functionally diverse signal transduction pathways. Glucose and amino acids trigger signaling cascades that regulate various aspects of fuel and energy metabolism and control the growth, proliferation, and survival of cells. Here, we provide a functional and regulatory overview of three well-established nutrient signaling pathways—the hexosamine signaling pathway, the mTOR (mammalian target of rapamycin) signaling pathway, and the adenosine monophosphate–activated protein kinase (AMPK) signaling pathway. Nutrient signaling pathways are interconnected, coupled to insulin signaling, and linked to the release of metabolic hormones from adipose tissue. Thus, nutrient signaling pathways do not function in isolation. Rather, they appear to serve as components of a larger "metabolic regulatory network" that controls fuel and energy metabolism (at the cell, tissue, and whole-body levels) and links nutrient availability with cell growth and proliferation. Understanding the diverse roles of nutrients and delineating nutrient signaling pathways should facilitate drug discovery research and the search for novel therapeutic compounds to prevent and treat various human diseases such as diabetes, obesity, and cancer.
Transducing the Signals: A G Protein Takes a New Identity
Authors: Dhanasekaran Danny N.
Published: September 2006

The prevailing dogma is that heterotrimeric G proteins exclusively transduce signals from the seven-transmembrane motif–containing cell surface receptors, also known as G protein–coupled receptors (GPCRs). New evidence indicates that Gα
13
, the α subunit of the G protein G
13
, breaks away from this traditional exclusive signaling alliance with GPCRs to transmit signals from receptor tyrosine kinases (RTKs), such as platelet-derived growth factor receptor (PDGFR), epidermal growth factor receptor (EGFR), and vascular endothelial growth factor receptor (VEGFR). Gα
13
is involved in cell migration in response to GPCRs activated by lysophosphatidic acid (LPA) or thrombin. A new report indicates that Gα
13
is also required for cell migration induced by the growth factors, such as PDGF, EGF, or VEGF. GPCR coupling is not required for such RTK-to-Gα
13
signaling. This new identity for Gα
13
as a signal transducer for both GPCRs and RTKs may be a forerunner for similar findings involving other Gα subunits. This expanding role of G proteins in both GPCR signaling and RTK signaling is likely to have a great impact not only on our understanding of cell signaling in general, but also more specifically where the dysregulation of signaling by GPCRs, RTKs, and G proteins cause pathophysiological changes such as in the case of tumorigenesis, tumor progression and/or metastasis.
A RSK(y) Relationship with Promiscuous PKA
Authors: Houslay Miles D.
Published: September 2006

Where, when, and with "whom" do molecules interact? Such relations in space and time are key concepts that currently engage investigators of cellular signaling processes. The notion of compartmentalized signaling grew out of studies of adenosine 3′,5′-monophosphate (cAMP) signaling processes, and this area continues to generate exciting new paradigms. Distinct clouds of cAMP are formed and shaped within cells by tethered cAMP phosphodiesterases (PDEs). AKAPosomes, formed from distinct subpopulations of cAMP-dependent protein kinase (PKA) tethered to anchoring proteins (AKAPs) together with specific substrate molecules, interpret these gradients to generate individualized responses. PKA activity is also regulated by the interaction of other proteins with the regulatory (R) or catalytic (C) subunits of PKA, and a mechanism has been uncovered in which ribosomal S6 kinase (RSK1) interacts with either PKA subunit, depending on whether RSK1 has been phosphorylated and activated by extracellular signal–regulated kinase (ERK). Thus, inactive RSK1 binds the RI subunit of PKA to sensitize it to activation, whereas activated RSK1 binds the C subunit to desensitize PKA to cAMP activation. Cross-talk between the key cAMP and ERK signaling pathways provides a mechanism that, along with distinct mechanisms of both positive and negative attenuation provided by Raf and PDE4 isoforms, can be tailored on a cell type–specific basis.
Localizing NADPH Oxidase–Derived ROS
Authors: Ushio-Fukai Masuko
Published: September 2006

Reactive oxygen species (ROS) function as signaling molecules to mediate various biological responses, including cell migration, growth, and gene expression. ROS are diffusible and short-lived molecules. Thus, localizing the ROS signal at the specific subcellular compartment is essential for activating redox signaling events after receptor activation. NADPH (nicotinamide adenine dinucleotide phosphate) oxidase is one of the major sources of ROS in vasculature; it consists of a catalytic subunit (Nox1, Nox2, Nox3, Nox4, or Nox5), p22phox, p47phox, p67phox, and the small guanosine triphosphatase Rac1. Targeting of NADPH oxidase to focal complexes in lamellipodia and membrane ruffles through the interaction of p47phox with the scaffold proteins TRAF4 and WAVE1 provides a mechanism for achieving localized ROS production, which is required for directed cell migration. ROS are believed to inactivate protein tyrosine phosphatases, which concentrate in specific subcellular compartments, thereby establishing a positive feedback system that activates redox signaling pathways to promote cell movement. Additionally, ROS production may be localized through interactions of NADPH oxidase with signaling platforms associated with lipid rafts and caveolae, as well as with endosomes. There is also evidence that NADPH oxidase is found in the nucleus, indicating its involvement in redox-responsive gene expression. This review focuses on targeting of NADPH oxidase to discrete subcellular compartments as a mechanism of localizing ROS and activation of downstream redox signaling events that mediate various cell functions.
Multifunctional Potassium Channels: Electrical Switches and Redox Enzymes, All in One
Authors: Heinemann Stefan H. , Hoshi Toshinori
Published: September 2006

Kv1-type K
+
channels are protein complexes containing both voltage-sensing, pore-forming α subunits and modulatory Kvβ subunits. Although some Kvβ subunits include an amino-terminal region that allows them to transform noninactivating Kv1 channels into rapidly inactivating channels, the function of Kvβ subunits that do not possess these inactivating amino-terminal regions has been less clear. Recent research demonstrates that Kvβ2 acts as an NADPH (the reduced form of nicotinamide adenine dinucleotide phosphate)–dependent redox enzyme and that its catalytic activity can regulate the speed with which the Kv1.4-Kvβ2 complex undergoes inactivation, suggesting that Kvβ2 may link cellular metabolic activity and redox state with electrical signaling.
Real-Time Measurements of Protein Affinities on Membrane Surfaces by Fluorescence Spectroscopy
Authors: Philip Finly , Scarlata Suzanne
Published: September 2006

Signal transduction in cells involves transitory interactions between proteins and membranes and between different proteins of the interacting species. These associations depend on the strength of the interactions and on the local concentration. Because the energy and intensity of the fluorescence of many probes are very sensitive to the local environment, fluorescence measurements can report on events, such as membrane binding and protein association, in real time. We describe methods to monitor associations both in vitro and in vivo by fluorescence.
Assessing Undergraduate Laboratory Performance
Authors: Adler Elizabeth M. , Gough Nancy R.
Published: September 2006

The lab notebook is one element for assessing student laboratory performance. However, it is also important to be able to communicate research results in a journal article format and a visual poster format. Another key aspect to scientific research is the ability to present a research plan. This article describes four assessment tools that can be used in conjunction with undergraduate lab courses that provide the opportunity for the students to practice each of these essential communication skills.
Differentiation of PC12 Cells
Authors: Adler Elizabeth M. , Gough Nancy R. , Blundon Jay A.
Published: September 2006

This 3-week-long series of collaborative laboratory exercises explores how to use a cultured cell system (PC12 cells) to study signaling pathways involved in cellular differentiation. The laboratory would be useful in a neurobiology or cell biology course for advanced undergraduate students. The background and details for performing the lab are provided along with suggestions for assessing student performance and understanding.
Systems Biology of AGC Kinases in Fungi
Authors: Sobko Alex
Published: September 2006

Sch9 appears to be the
Saccharomyces cerevisiae
homolog of protein kinase B and is involved in the control of numerous nutrient-sensitive processes, including regulation of cell size, cell cycle progression, and stress resistance. Sch9 has also been implicated in the regulation of replicative and chronological life span. Systematic comparison of the phenotypes of
sch9
and other AGC kinase mutants in fungal species with their counterparts in model eukaryotic organisms provides insight into the functions of AGC kinases. The availability of data from global studies of protein-protein interactions now makes it possible to predict and validate functional connections between Sch9, its putative substrates, and other proteins. This review highlights several emerging paradigms of AGC kinase signaling that are relevant for growth, development, and aging.
Cross-Linking Transglutaminases with G Protein–Coupled Receptor Signaling
Authors: Iismaa Siiri E. , Begg Gillian E. , Graham Robert M.
Published: September 2006

Transglutaminases are a family of calcium- and thiol-dependent acyl transferases that catalyze the formation of an amide bond between the γ-carboxamide groups of peptide-bound glutamine residues and the primary amino groups in various compounds, including the ε-amino group of lysines in certain proteins. As a result, these enzymes effect posttranslational modification of proteins by amine incorporation, or stabilization of protein assemblies by their cross-linking; such actions profoundly influence critical biological processes such as blood clotting and protection from infection and dehydration by establishing the barrier function of skin. In addition, transglutaminases have other more diverse actions, including involvement in signaling by the superfamily of heterotrimeric guanine nucleotide–binding protein (G protein)–coupled receptors (GPCRs) in one of three ways: (i) through actions as guanosine triphosphate–binding proteins that activate intracellular effectors, such as phospholipase C; (ii) by cross-linking GPCR monomers to enhance signaling as a result of covalent dimer formation; or (iii) by interacting with an apparent growth inhibitory orphan GPCR, GPR56, to limit metastatic spread of melanoma cells. The implications of these receptor-coupled actions of transglutaminases are discussed.
Simultaneous Optical Measurements of Cytosolic Ca
2+
and cAMP in Single Cells
Authors: Harbeck Mark C. , Chepurny Oleg , Nikolaev Viacheslav O. , Lohse Martin J. , Holz George G. , Roe Michael W.
Published: September 2006

Understanding the temporal and spatial integration of the Ca
2+
and adenosine 3′,5′-monophosphate (cAMP) signaling pathways requires concurrent measurements of both second messengers. Here, we describe an optical technique to simultaneously image cAMP and Ca
2+
concentration gradients in MIN6 mouse insulinoma cells using Epac1-camps, a Förster (or fluorescence) resonance energy transfer (FRET)-based cAMP biosensor, and Fura-2, a fluorescent indicator of Ca
2+
. This real-time imaging method allows investigation of the dynamic organization and integration of multiple levels of signal processing in single living cells.
Irrelevant Apoptosis?
Authors: Adler Elizabeth M.
Published: September 2006

Research from two groups suggests that the ability of p53 to stimulate apoptosis in response to DNA damage is independent of its role as a tumor suppressor. p53, which is frequently mutated in human cancers, mediates cellular responses to DNA damage and to oncogenic stress. In wild-type mice, radiation elicits DNA damage-dependent cell death, whereas strains of mice that lack functional p53 are resistant to radiation-induced cell death but rapidly develop radiation-induced lymphomas. Christophorou
et al
. used transgenic mice that express a 4-hydroxytamoxifen (4-OHT)-sensitive fusion protein that allows p53 to be reversibly switched between functional (in the presence of 4-OHT) and nonfunctional states [homozygous

p53ER
TAM

knockin (

p53
KI/KI

) mice] to investigate the relation between p53’s roles in radiation-dependent apoptosis and as a tumor suppressor.

p53
KI/KI

mice treated with 4-OHT during the 6 days before irradiation, so that functional p53 was present during irradiation, showed widespread apoptosis of primary lymphoid organs and intestinal epithelium and sustained leukocytopenia (low white blood cell count) but no protection from lymphoma, compared with

p53
KI/KI

mice that were not treated with 4-OHT. In contrast,

p53
KI/KI

mice in which p53 function was restored for 6 days starting 8 days after irradiation failed to experience radiation-dependent apoptosis but were protected from radiation-induced lymphomagenesis, with tumor onset delayed about 100 days. The ability of delayed p53 restoration to suppress lymphomagenesis depended on p19
ARF
, which is involved in oncogenic signaling to p53, rather than in the DNA damage response.

In the second study, Efeyan
et al
. showed that the protective effect of an extra copy of the p53 gene against spontaneous tumors and tumors following exposure to 3-methyl cholanthrene critically depended on p19
ARF
. Thus, both groups conclude that oncogenic signaling to p53, rather than DNA damage-dependent apoptosis, is critical to p53’s role as a tumor suppressor. Berns discusses implications of the research with regard to controlling toxic side effects of cancer therapy.

M. A. Christophorou, I. Ringshausen, A. J. Finch, L. Brown Swigart, G. I. Evan, The pathological response to DNA damage does not contribute to p53-mediated tumour suppression.
Nature
443
, 214-217 (2006).
[PubMed]

A. Efeyan, I. Garcia-Cao, D. Herranz, S. Velasco-Miguel, M. Serrano, Policing of oncogene activity by p53.
Nature
443
, 159 (2006).
[PubMed]

A. Berns, Can less be more for p53?
Nature
443
, 153-154 (2006).
[PubMed]
Promoting Regeneration Rather Than Death?
Authors: Adler Elizabeth M.
Published: September 2006

Di Giovanni
et al
., who previously identified a group of temporally correlated genes that appear after spinal cord injury, including some known to be involved in neurite outgrowth and axon regeneration, undertook a search for a transcription factor that might coordinately regulate their expression. In silico promoter analysis revealed that two of these genes, the gene encoding the actin-binding Coronin 1b and the small guanosine triphosphatase Rab13 (both implicated in neurite outgrowth), had multiple binding sites for the transcription factor p53. Chromatin immunoprecipitation (ChIP) analysis confirmed that p53 bound to the Coronin 1b and Rab13 promoters, and p53 expression enhanced expression of their mRNA and protein in various cell lines. In cultured rat cortical neurons exposed to neuronal medium or PC12 cells exposed to nerve growth factor, nuclear p53 abundance increased together with expression of Coronin 1b and Rab13 during neurite outgrowth, whereas a p53 dominant negative inhibited Coronin 1b and Rab13 expression and neurite outgrowth. In NT2/D1 human teratocarcinoma cells, which develop neurites and synapses in response to treatment with retinoic acid (RA), RA stimulated acetylation of p53 at lysine 320. ChIP analysis and experiments involving p53 acetylation mutants indicated that p53 acetylated at lysine 320 binds to the Coronin 1b and Rab13 promoters to stimulate Coronin 1b and Rab13 expression and neurite outgrowth. Mice lacking p53 showed decreased nerve regeneration after axotomy compared with wild-type mice and, unlike regenerating neurons in wild-type mice, those in p53-null mice did not show enhanced expression of Coronin 1b and Rab13. Thus, the authors conclude that the well-known tumor suppressor p53, already implicated in neuronal apoptosis, likely plays a role in stimulating neurite outgrowth and neuronal regeneration.

S. Di Giovanni, C. D. Knights, M. Rao, A. Yakovlev, J. Beers, J. Catania, M. L. Avantaggiati, A. I. Faden, The tumor suppressor protein p53 is required for neurite outgrowth and axon regeneration.
EMBO J.
25
, 4084-4096 (2006).
[PubMed]
Challenging the JNK2 Knockout Data
Authors: Gough Nancy R.
Published: September 2006

Data from mouse knockouts suggested that c-Jun N-terminal kinases 1 and 2 (JNK1 and JNK2) exhibited opposing effects, with JNK1 stimulating the JNK pathway and JNK2 inhibiting the pathway. Jaeschke
et al
. challenge this result using a chemical genetics approach in which JNK2 activity is acutely inhibited in cells expressing a mutant form of JNK2 (JNK2
MG
) that is sensitive to derivatives of the protein kinase inhibitor PP1. The authors genetically engineered mice to be either
JNK1
+/+
,
JNK2
MG/MG
, or
JNK1
−/−
,
JNK2
MG/MG
. In multiple assays, in the presence of the inhibitor, the response of the
JNK1
+/+
,
JNK
MG/MG
mouse embryo fibroblasts (MEFs) was the same as that of wild-type MEFs, suggesting that JNK1 and JNK2 are redundant activators of the pathway. Furthermore, the responses of the
JNK1
−/−
,
JNK2
MG/MG
MEFs to activation of the JNK pathway in the presence of the inhibitor were decreased. This was unexpected, given that
JNK1
+/+
,
JNK2
−/−
MEFs showed enhanced responses and
JNK1
−/−
,
JNK2
+/+
MEFs showed decreased responses to activators of the JNK pathway, such as ultraviolet radiation, which suggested a negative role for JNK2. The authors showed that
JNK2
−/−
MEFs exhibited enhanced JNK1 activity, suggesting that under the chronic absence of JNK2, there is a compensatory increase in JNK1, which explains why JNK signaling appears to be enhanced in the absence of JNK2 activity. This compensatory increase in JNK1 does not occur when the chemical genetics approach is used to modulate the activity of JNK2, because both enzymes are present at wild-type abundance and only the activity of JNK2 is acutely inhibited, allowing the contribution of this protein to be assessed in a more normal environment.

A. Jaeschke, M. Karasarides, J.-J. Ventura, A. Ehrhardt, C. Zhang, R. A. Flavell, K. M. Shokat, R. J. Davis, JNK2 is a positive regulator of the cJun transcription factor.
Mol. Cell
23
, 899-911 (2006).
[Online Journal]
Caveolin and Liver Regeneration
Authors: Hurtley Stella M.
Published: September 2006

Caveolin is a key component of caveolae, cell surface invaginations involved in the internalization of a variety of signaling molecules and the uptake of certain viruses. Surprisingly enough, when caveolin knockout mice were generated a few years ago, they appeared to be healthy. Fernández
et al
. have now examined these mice in more detail and discovered a phenotype in these animals—a profound defect in liver regeneration leading to reduced survival after partial hepatectomy. Problems uncovered included changes in lipid metabolism and cell cycle progression. Treating mutant mice with glucose could circumvent the defect and improve survival after liver damage

M. A. Fernández, C. Albor, M. Ingelmo-Torres, S. J. Nixon, C. Ferguson, T. Kurzchalia, F. Tebar, C. Enrich, R. G. Parton, A. Pol, Caveolin-1 is essential for liver regeneration.
Science
313
, 1628-1632 (2006).
[Abstract]
[Full Text]

D. L. Brasaemle, A metabolic push to proliferate.
Science
313
, 1581-1582 (2006).
[Summary]
[Full Text]