Proton Magnetic Resonance Spectroscopy of Klebsiella Capsular Polysaccharides

The presence of acetate and pyruvate groups in Klebsiella capsular polysaccharides may be demonstrated and estimated quantitatively by running the proton magnetic resonance spectrum of the polysaccharide (as sodium salt) in deuterium oxide at 95 C. Such spectra also permit an assessment to be made of the number of alpha- and beta-linkages in the repeat unit of the polysaccharide structure.     

Amino acid residues in the Ler protein critical for derepression of the <Emphasis Type="Italic">LEE5</Emphasis> promoter in enteropathogenic <Emphasis Type="Italic">E. coli</Emphasis>

Enteropathogenic E. coli causes attaching and effacing (A/E) intestinal lesions. The genes involved in the formation of A/E lesions are encoded within a chromosomal island comprising of five major operons, LEE1-5. The global regulator H-NS represses the expression of these operons. Ler, a H-NS homologue, counteracts the H-NS–mediated repression. Using a novel genetic approach, we identified the amino acid residues in Ler that are involved in the interaction with H-NS: I20 and L23 in the C-terminal portion of α-helix 3, and I42 in the following unstructured linker region.     

Alpha 5/6 helix domains together with N-terminus determine the apoptotic potency of the Bcl-2 family proteins

A critical process in apoptosis is the permeabilization of the mitochondrial outer membrane (MOM). This process is known to be regulated by the multi-domain Bcl-2 family proteins. For example, the pro-apoptotic proteins Bax and Bak are responsible for forming pores at MOM. The anti-apoptotic proteins (including Bcl-2, Mcl-1 and Bcl-xL), on the other hand, can inhibit this pore-forming process. Interestingly, although these two subgroups of proteins perform opposite apoptotic functions, their structures are very similar. This raises two highly interesting questions: (1) Why do these structurally similar proteins play opposite roles in apoptosis? (2) What are the roles of different functional domains of a Bcl-2 family protein in determining its apoptotic property? In this study, we generated a series of deletion mutants and substitution chimera, and used a combination of molecular biology, bio-informatics and living cell imaging techniques to answer these questions. Our major findings are: (1) All of the Bcl-2 family proteins appear to possess an intrinsic pro-apoptotic property. (2) The N-termini of these proteins play an active role in suppressing their pro-apoptotic function. (3) The apoptotic potency is positively correlated with membrane affinity of the alpha 5/6 helix domains. (4) Charge distribution flanking the alpha 5/6 helices is also important for the apoptotic potency. These findings explain why different members of Bcl-2 family proteins with similar domain composition can function oppositely in the apoptotic process.     

Mouse models of rhinovirus-induced disease and exacerbation of allergic airway inflammation

Rhinoviruses cause serious morbidity and mortality as the major etiological agents of asthma exacerbations and the common cold. A major obstacle to understanding disease pathogenesis and to the development of effective therapies has been the lack of a small-animal model for rhinovirus infection. Of the 100 known rhinovirus serotypes, 90% (the major group) use human intercellular adhesion molecule-1 (ICAM-1) as their cellular receptor and do not bind mouse ICAM-1; the remaining 10% (the minor group) use a member of the low-density lipoprotein receptor family and can bind the mouse counterpart. Here we describe three novel mouse models of rhinovirus infection: minor-group rhinovirus infection of BALB/c mice, major-group rhinovirus infection of transgenic BALB/c mice expressing a mouse-human ICAM-1 chimera and rhinovirus-induced exacerbation of allergic airway inflammation. These models have features similar to those observed in rhinovirus infection in humans, including augmentation of allergic airway inflammation, and will be useful in the development of future therapies for colds and asthma exacerbations.     

Proteasome inhibition: an early or late event in nitric oxide-induced neuronal death?

Nitric oxide (NO), ubiquitously expressed in the central nervous system, has been perceived to be a potential neuromodulator. Employing cultured murine primary cortical neurons, NO resulted in an inhibition of the ubiquitin-proteasome system (UPS) with a dose- and time-dependent decrease in cell viability. This is consistent with a previous study that reported a dysfunction of UPS with consequential apoptotic death in macrophage cell with NO treatment. However, it cannot be unclear if the drop in UPS efficiency is directly imposed on by NO. Therefore by using microarray analysis, our study revealed an early down-regulation or non-significant differential expression of genes encoding UPS proteins in NOC-18 (NO donor)-treated neurons as compared to an observed elevation of corresponding gene expression genes in lactacystin (classical proteasome inhibitor)-treated neurons (conducted earlier). Furthermore, time-course analysis of proteasome activity in NOC-18-treated neurons demonstrated a late onset of reduction. This is intriguing as it is well established that in an exclusive proteasome dysfunction-induced cell death, a compensatory feedback mechanism will be activated with an initial and concerted up-regulation of genes encoding proteins involved in UPS as seen when neurons were treated with lactacystin. Thus, it is highly suggestive that NO-triggered neuronal death takes on a different signaling cascade from that of a classical proteasome inhibitor, and that the late reduction of proteasome activity is a downstream event following the activation of apoptotic cellular signaling cascade. In intracellular condition, the proteasome is not NO preferred primary target responsible for the trigger of the cell death machinery. In conclusion, we presented novel findings that shed light into NO-induced cell death signaling cascade, which would be important in understanding the pathogenesis of neurodegenerative disorders such as Parkinson's disease.     

The coup de grâce for the nested clade phylogeographic analysis?

Nested clade phylogeographic analysis (NCPA) has become a popular method for reconstructing the history of populations across species ranges. Ever since its invention in 1995, criticisms have been formulated, but the method, which has been regularly updated, continues to attract investigators. Molecular Ecology has published a large fraction of the literature on the topic — both pro and con. A recent study by Panchal and Beaumont (2007) finally allows a precise evaluation of the method by developing software that automates the somewhat complicated NCPA procedure. Using simulations of random-mating populations, Panchal and Beaumont find a high frequency of false-positives with their automated NCPA procedure (over 75%). These findings, which echo and amplify earlier warnings, appear serious enough to suggest to researchers to await further evaluation of the method. Although no other all-encompassing method such as the NCAP currently exists to evaluate phylogeographic data sets, researchers have many alternative methods to test ever more refined hypotheses.     

Weekly pattern of stroke onset in an Asian country: a nationwide population-based study

This study used a nationwide population-based dataset to explore the variation among the days of week of stroke onset within population subgroups defined by age, sex, and stroke type. We used ambulatory care data from the 2002 Taiwan National Health Insurance Research Database, focusing on 42,779 emergency room (ER) visits for stroke that year. All analyses were stratified by sex, age (<60 and > or =60 yrs), and type of stroke. Auto-Regressive Integrated Moving Average (ARIMA) was performed to investigate the relationship between daily number of stroke events and holidays and days of the week after adjusting for the effects of seasonality and trends. One-way ANOVA revealed significant differences in stroke ER admissions according to day of week according to age <60 (p<0.01), age > or =60 (p<0.001), male (p<0.001), female (p<0.001), ischemic stroke (IS) (p<0.001), and unspecified stroke (UNSP) (p<0.001). However, the analysis by type-subarachnoid hemorrhage and intracerebral hemorrhage-did not show significant relationships between daily emergency room stroke admissions, holidays, or day of the week. The ARIMA regression analyses also showed that Mondays had the highest rate of emergency room admissions for stroke regardless of sex, age, or IS and UNSP types of stroke, after adjusting for seasonality and trends. We conclude that stroke occurs more frequently on Mondays than on the other days of the week, which might be associated with short-term changes in lifestyle or due to the sudden return of stress on the first working day of the week, and on holidays.     

Cyclooxygenase expression is elevated in retinoic acid-differentiated U937 cells

Cyclooxygenase (COX) is the rate-limiting enzyme for the biosynthesis of prostaglandins in monocytes/macrophages. The COX-1 is constitutively expressed in most tissues and may be involved in cellular homeostasis, whereas the COX-2 is an inducible enzyme that may play an important role in inflammation and mitogenesis. When U937 monocytic cells were incubated with retinoic acid (RA) for 48 h, cell differentiation took place with concomitant increases in prostaglandin E₂ (PGE₂) production and COX activity. In this study, the mechanism of RA (all-trans- or 9-cis-RA)-induced enhancement of PGE₂ biosynthesis in U937 cells was examined. Treatment of cells with all-trans- or 9-cis-RA up to 48 h caused an increase in PGE₂ production in a time- and dose-dependent manner. Both RA isomers caused the enhancement of PGE₂ production and the up-regulation of COX-1 expression at the protein and mRNA levels. The increase in COX-1 mRNA was found to precede the increase in COX-1 protein expression. Interestingly, the COX-2 protein and COX-2 mRNA were not detected in U937 cells, and their levels remained undetectable during the entire course of RA treatment. We conclude that treatment of U937 cells by RA for 48 h caused the initiation of cell differentiation, which was found to be concomitant with a significant increase in PGE₂ production mediated via the up-regulation of COX-1 mRNA and protein expression.     

The Drosophila homologue of Arf-GAP GIT1, dGIT, is required for proper muscle morphogenesis and guidance during embryogenesis

GIT1-like proteins are GTPase-activating proteins (GAPs) for Arfs and interact with a variety of signaling molecules to function as integrators of pathways controlling cytoskeletal organization and cell motility. In this report, we describe the characterization of a Drosophila homologue of GIT1, dGIT, and show that it is required for proper muscle morphogenesis and myotube guidance in the fly embryo. The dGIT protein is concentrated at the termini of growing myotubes and localizes to muscle attachment sites in late stage embryos. dgit mutant embryos show muscle patterning defects and aberrant targeting in subsets of their muscles. dgit mutant muscles fail to localize the p21-activated kinase, dPak, to their termini. dPak and dGIT form a complex in the presence of dPIX and dpak mutant embryos show similar muscle morphogenesis and targeting phenotypes to that of dgit. We propose that dGIT and dPak are part of a complex that promotes proper muscle morphogenesis and myotube targeting during embryogenesis.