Metabolic interdependence of obligate intracellular bacteria and their insect hosts.

Mutualistic associations of obligate intracellular bacteria and insects have attracted much interest in the past few years due to the evolutionary consequences for their genome structure. However, much less attention has been paid to the metabolic ramifications for these endosymbiotic microorganisms, which have to compete with but also to adapt to another metabolism--that of the host cell. This review attempts to provide insights into the complex physiological interactions and the evolution of metabolic pathways of several mutualistic bacteria of aphids, ants, and tsetse flies and their insect hosts.     

Functional fluorescent Ca2+ indicator proteins in transgenic mice under TET control

Genetically encoded fluorescent calcium indicator proteins (FCIPs) are promising tools to study calcium dynamics in many activity-dependent molecular and cellular processes. Great hopes—for the measurement of population activity, in particular—have therefore been placed on calcium indicators derived from the green fluorescent protein and their expression in (selected) neuronal populations. Calcium transients can rise within milliseconds, making them suitable as reporters of fast neuronal activity. We here report the production of stable transgenic mouse lines with two different functional calcium indicators, inverse pericam and camgaroo-2, under the control of the tetracycline-inducible promoter. Using a variety of in vitro and in vivo assays, we find that stimuli known to increase intracellular calcium concentration (somatically triggered action potentials (APs) and synaptic and sensory stimulation) can cause substantial and rapid changes in FCIP fluorescence of inverse pericam and camgaroo-2.     

Epiphytic Refugium: Are Two Species of Invading Freshwater Bivalves Partitioning Spatial Resources?

Enumeration of benthic (bottom dwelling) and epiphytic (attached to plants) zebra and quagga mussels (Dreissena polymorpha and D. bugensis, respectively) at Lake Erie near-shore sites in fall of 2000 revealed an unexpected prevalence of the zebra mussel on submerged plants. Even at Buffalo, New York, USA, where benthic dreissenids have been 92–100% quagga mussel since 1996, zebra mussels constituted 30–61% of epiphytes numerically. This may reflect a partitioning of settling space consistent with interspecific competition. A seasonal epiphytic refugium might allow the zebra mussel to persist even where the benthos is almost exclusively quagga mussel. This revised version was published online in July 2006 with corrections to the Cover Date.     

What can we learn from noncoding regions of similarity between genomes?

Background  

In addition to known protein-coding genes, large amounts of apparently non-coding sequence are conserved between the human and mouse genomes. It seems reasonable to assume that these conserved regions are more likely to contain functional elements than less-conserved portions of the genome.     

Troponin T isoforms alter the tolerance of transgenic mouse cardiac muscle to acidosis

Troponin T (TnT) is an essential protein in the Ca²⁺ regulatory system of striated of muscle. Three fiber type-specific TnT genes have evolved in higher vertebrates to encode cardiac, slow and fast skeletal muscle TnT isoforms. To understand the functional significance of TnT isoforms, we studied the effects of acidosis on the contractility of transgenic mouse cardiac muscle that expresses fast skeletal muscle TnT. Contractility analysis of intact cardiac muscle strips showed that while no differences were detected at physiological pH, the transgenic cardiac muscle had significantly greater decreases in +dF/dt_max at acidic pH than that of the wild-type control. Contractility of skinned cardiac muscles demonstrated that the presence of fast TnT resulted in significantly larger decreases in force and Ca²⁺ sensitivity at acidic pH than that of the wild-type control. The effect of TnT isoforms on the tolerance of muscle to acidosis may explain the higher tolerance of embryonic versus adult cardiac muscles. The results are consistent with the hypothesis that charge differences in TnT isoforms contribute to the contractility of muscle. The data further support a hypothesis that slow TnT is similar to the cardiac, but not fast, and TnT may contribute to the higher tolerance of slow muscles to stress conditions. Therefore, TnT isoform diversity may contribute to the compatibility of muscle thin filaments to cellular environments in different fiber types, during development and functional adaptation.     

Photooxidation of pterin in aqueous solutions: biological and biomedical implications

Studies of the photochemical reactivity of pterin (= 2-aminopteridin-4(3H)-one; PT) in acidic (pH 5.0-6.0) and alkaline (pH 10.2-10.8) aqueous solutions have been performed. The photochemical reactions were followed by UV/VIS spectrophotometry, thin layer chromatography (TLC), high-performance liquid chromatography (HPLC), and an enzymatic method for H2O2 determination. PT is not light-sensitive in the absence of molecular oxygen, but it undergoes photooxidation in the presence of O2, yielding several nonpteridinic products. The quantum yields for PT disappearance were found to be 8.2 (+/-0.6) x 10(-4) and 1.2 (+/-0.2) x 10(-3) in acidic and alkaline media, respectively. H2O2 was detected and quantified in irradiated solutions of PT; and its importance from a biomedical point of view is discussed. The rate constant of the chemical reaction between singlet oxygen ((1)O2) and PT was determined to be 2.5 (+/-0.2) x 10(5) l mol(-1) s(-1) in alkaline medium, and the role of (1)O2 in the photooxidation of pterin was evaluated.     

Characterization of ligand binding to the bifunctional key enzyme in the sialic acid biosynthesis by NMR: II. Investigation of the ManNAc kinase functionality

N-Acetylmannosamine (ManNAc) is the physiological precursors to all sialic acids that occur in nature. As variations in the sialic acid decoration of cell surfaces can profoundly affect cell-cell, pathogen-cell, or drug-cell interactions, the enzymes that convert ManNAc into sialic acid are attractive targets for the development of drugs that specifically interrupt sialic acid biosynthesis or lead to modified sialic acids on the surface of cells. The first step in the enzymatic conversion of ManNAc into sialic acid is phosphorylation, yielding N-acetylmannosamine-6-phosphate. The enzyme that catalyzes this conversion is the N-acetylmannosamine kinase (ManNAc kinase) as part of the bifunctional enzyme UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase. Here, we employed saturation transfer difference (STD) NMR experiments to study the binding of ManNAc and related ligands to the ManNAc kinase. It is shown that the configuration of C1 and C4 of ManNAc is crucial for binding to the enzyme, whereas the C2 position not only accepts variations in the attached N-acyl side chain but also tolerates inversion of configuration. Our experiments also show that ManNAc kinase maintains its functionality, even in the absence of Mg(2+). From the analysis of the STD NMR-derived binding epitopes, it is concluded that the binding mode of the N-acylmannosamines critically depends on the N-acyl side chain. In conjunction with the relative binding affinities of the ligands obtained from STD NMR titrations, it is possible to derive a structure-binding affinity relationship. This provides a cornerstone for the rational design of drugs for novel therapeutic applications by altering the sialic acid decorations of cell walls.     

Evaluation of a modified selective differential medium for the isolation of Stenotrophomonas maltophilia

VIA medium for Stenotrophomonas maltophilia was modified by substituting meropenem (16 mg/L) for imipenem. S. maltophilia grew from 40% of drains sampled within a hospital and surrounding locations in Perth, Western Australia. The specificity of the new medium for S. maltophilia was 62%, and all contaminating bacteria were easily distinguishable by phenotypic tests and PCR.