Molecular evolution of a multigene family in group A streptococci

The emm genes are members of a gene family in group A streptococci (GAS) that encode for antiphagocytic cell-surface proteins and/or immunoglobulin-binding proteins. Previously sequenced genes in this family have been named "emm," "fcrA," "enn," "arp," "protH," and "mrp"; herein they will be referred to as the "emm gene family." The genes in the emm family are located in a cluster occupying 3-6 kb between the genes mry and scpA on the chromosome of Streptococcus pyogenes. Most GAS strains contain one to three tandemly arranged copies of emm-family genes in the cluster, but the alleles within the cluster vary among different strains. Phylogenetic analysis of the conserved sequences at the 3' end of these genes differentiates all known members of this family into four evolutionarily distinct emm subfamilies. As a starting point to analyze how the different subfamilies are related evolutionarily, the structure of the emm chromosomal region was mapped in a number of diverse GAS strains by using subfamily-specific primers in the polymerase chain reaction. Nine distinct chromosomal patterns of the genes in the emm gene cluster were found. These nine chromosomal patterns support a model for the evolution of the emm gene family in which gene duplication followed by sequence divergence resulted in the generation of four major-gene subfamilies in this locus.      

Role of the reticulum in the stability and shape of the isolated human erythrocyte membrane

In order to examine the widely held hypothesis that the reticulum of proteins which covers the cytoplamsic surface of the human erythrocyte membrane controls cell stability and shape, we have assessed some of its properties. The reticulum, freed of the bilayer by extraction with Triton X-100, was found to be mechanically stable at physiological ionic strength but physically unstable at low ionic strength. The reticulum broke down after a characteristic lag period which decreased 500-fold between 0 degrees and 37 degrees C. The release of polypeptide band 4.1 from the reticulum preceded that of spectrin and actin, suggesting that band 4.1 might stabilize the ensemble but is not essential to its integrity. The time-course of breakdown was similar for ghosts, the reticulum inside of ghosts, and the isolated reticulum. However, at very low ionic strength, the reticulum was less stable within the ghost than when free; at higher ionic strength, the reverse was true. Over a wide range of conditions the membrane broke down to vesicles just as the reticulum disintegrated, presumably because the bilayer was mechanically stabilized by this network. The volume of both ghosts and naked reticula varied inversely and reversibly with ionic strength. The volume of the naked reticulum varied far more widely than the ghost, suggesting that its deformation was normally limited by the less extensible bilayer. The contour of the isolated reticulum was discoid and often dimpled or indented, as visualized in the fluorescence microscope after labeling of the ghosts with fluoroscein isothiocyanate. Reticula derived from ghosts which had lost the ability to crenate in isotonic saline were shriveled, even though the bilayer was smooth and expanded. Conversly, ghosts crenated by dinitrophenol yielded smooth, expanded reticula. We conclude that the reticulum is a durable, flexible, and elastic network which assumes and stabilizes the contour of the membrane but is not responsible for its crenation.     

Diversity of lactic acid bacteria of the bioethanol process

Background  

Bacteria may compete with yeast for nutrients during bioethanol production process, potentially causing economic losses. This is the first study aiming at the quantification and identification of Lactic Acid Bacteria (LAB) present in the bioethanol industrial processes in different distilleries of Brazil.     

The hydrophilic isoform of glutamate decarboxylase, GAD67, is targeted to membranes and nerve terminals independent of dimerization with the hydrophobic membrane-anchored isoform, GAD65

GAD67, the larger isoform of the gamma-aminobutyric acid-synthesizing enzyme glutamic acid decarboxylase, is a hydrophilic soluble molecule, postulated to localize at nerve terminals and membrane compartments by heterodimerization with the smaller membrane-anchored isoform GAD65. We here show that the dimerization region in GAD65 is distinct from the NH(2)-terminal membrane-anchoring region and that a membrane anchoring GAD65 subunit can indeed target a soluble subunit to membrane compartments by dimerization. However, only a fraction of membrane-bound GAD67 is engaged in a heterodimer with GAD65 in rat brain. Furthermore, in GAD65-/- mouse brain, GAD67, which no longer partitions into the Triton X-114 detergent phase, still anchors to membranes at similar levels as in wild-type mice. Similarly, in primary cultures of neurons derived from GAD65-/- mice, GAD67 is targeted to nerve terminals, where it co-localizes with the synaptic vesicle marker SV2. Thus, axonal targeting and membrane anchoring is an intrinsic property of GAD67 and does not require GAD65. The results suggest that three distinct moieties of glutamate decarboxylase localize to membrane compartments, an amphiphilic GAD65 homodimer, an amphiphilic GAD65/67 heterodimer, tethered to membranes via the GAD65 subunit, and a hydrophilic GAD67 homodimer, which associates with membranes by a distinct mechanism.     

Microbial Transport, Survival, and Succession in a Sequence of Buried Sediments

Abstract Two chronosequences of unsaturated, buried loess sediments, ranging in age from <10,000 years to >1 million years, were investigated to reconstruct patterns of microbial ecological succession that have occurred since sediment burial. The relative importance of microbial transport and survival to succession was inferred from sediment ages, porewater ages, patterns of abundance (measured by direct counts, counts of culturable cells, and total phospholipid fatty acids), activities (measured by radiotracer and enzyme assays), and community composition (measured by phospholipid fatty acid patterns and Biolog substrate usage). Core samples were collected at two sites 40 km apart in the Palouse region of eastern Washington State, near the towns of Washtucna and Winona. The Washtucna site was flooded multiple times during the Pleistocene by glacial outburst floods; the Winona site elevation is above flood stage. Sediments at the Washtucna site were collected from near surface to 14.9 m depth, where the sediment age was approximately 250 ka and the porewater age was 3700 years; sample intervals at the Winona site ranged from near surface to 38 m (sediment age: approximately 1 Ma; porewater age: 1200 years). Microbial abundance and activities declined with depth at both sites; however, even the deepest, oldest sediments showed evidence of viable microorganisms. Same-age sediments had equal quantities of microorganisms, but different community types. Differences in community makeup between the two sites can be attributed to differences in groundwater recharge and paleoflooding. Estimates of the microbial community age can be constrained by porewater and sediment ages. In the shallower sediments (<9 m at Washtucna, <12 m at Winona), the microbial communities are likely similar in age to the groundwater; thus, microbial succession has been influenced by recent transport of microorganisms from the surface. In the deeper sediments, the populations may be considerably older than the porewater ages, since microbial transport is severely restricted in unsaturated sediments. This is particularly true at the Winona site, which was never flooded.     

<Emphasis Type="Italic">Chlamydia pneumoniae</Emphasis> aggravates vein graft intimal hyperplasia in a rat model

Background  

Along with angioplasty, autologus vein grafts are commonly used for artery bypass grafting in patients with advanced arterial stenosis and drug-resistant angina pectoris. Although initially a successful procedure, long-term functionality is limited due to proliferation and migration of smooth muscle cells. Like in atherosclerosis, common chronic infections caused by viruses and bacteria may contribute to this process of vein graft failure. Here we investigated the possible role of Chlamydia pneumoniae (Cpn) in the pathogenesis of venous graft failure in an experimental animal model. In 2 groups (n = 10 rats/group), an epigastric vein-to-common femoral artery interposition graft was placed. Immediately thereafter, rats were infected with Cpn (5*10⁸ IFU) or injected with control solutions. Rats were sacrificed three weeks after surgery and the grafts were harvested for morphometrical and immunohistochemical analysis.     

Cyclic 3'', 5''-AMP relay in dictyostelium discoideum. IV. Recovery of the cAMP signaling response after adaptation to cAMP

In dictyoselium discoideum, an increase in extracellular cAMP activates adenylate cyclase, leading to an increase in intracellular cAMP and the rate of cAMP secretion. Cells adapt to any constant cAMP stimulus after several minutes, but still respond to an increase in the concentration of the stimulus. We have now characterized the decay of adaptation (deadaptation) after the removal of cAMP stimuli. Levels of adaptation were established by the perfusion of [(3)H]adenosine-labeled amoebae with a defined cAMP stimulus. After a variable recovery period, the magnitude of the signaling response to a second stimulus was measured; its attenuation was taken as a measure of residual adaption to the first stimulus. The level of adaptation established by the first stimulus depended on both its magnitude and duration. Deadaptation began as soon as the first stimulus was removed. The magnitude of the response to the second stimulus increased with the recovery time in a first-order fashion, with a t(1/2)=3-4 min for stimuli of 10(-8) M to 10(-5) M cAMP. Responses to test stimuli, although reduced in magnitude, had an accelerated time-course when they closely followed a prior response that had not completely subsided. This effect is called priming; we believe it reveals a reversible, rate-limiting step that modulates the onset and termination of the signaling responses of amoebae that have not recently responded to a cAMP stimulus. We have suggested that the cAMP signaling response is controlled by two antagonistic cellular processes, excitation and adaptation. The data reported here imply that both the rate of rise in the adaptation process and the final level reached depend on the occupancy of cAMP surface receptors and that the decay of adaptation when external cAMP is removed proceeds with first-order kinetics.     

Integrated circuits and molecular components for stress and feeding: implications for eating disorders

Eating disorders are complex brain disorders that afflict millions of individuals worldwide. The etiology of these diseases is not fully understood, but a growing body of literature suggests that stress and anxiety may play a critical role in their development. As our understanding of the genetic and environmental factors that contribute to disease in clinical populations like anorexia nervosa, bulimia nervosa and binge eating disorder continue to grow, neuroscientists are using animal models to understand the neurobiology of stress and feeding. We hypothesize that eating disorder clinical phenotypes may result from stress‐induced maladaptive alterations in neural circuits that regulate feeding, and that these circuits can be neurochemically isolated using animal model of eating disorders.     

The geographic apportionment of mitochondrial genetic diversity in east African chimpanzees, Pan troglodytes schweinfurthii

This study is a geographically systematic genetic survey of the easternmost subspecies of chimpanzee, Pan troglodytes schweinfurthii. DNA was noninvasively collected in the form of shed hair from chimpanzees of known origin in Uganda, Rwanda, Tanzania, and Zaire. Two hundred sixty-two DNA sequences from hypervariable region 1 of which of the mitochondrial control region were generated. Eastern chimpanzees display levels of mitochondrial genetic variation which are low and which are similar to levels observed in humans (Homo sapiens). Also like humans, between 80% and 90% of the genetic variability within the eastern chimpanzees is apportioned within populations. Spatial autocorrelation analysis shows that genetic similarity between eastern chimpanzees decreases clinically with distance, in a pattern remarkably similar to one seen for humans separated by equivalent geographic distances. Eastern chimpanzee mismatch distributions (frequency distributions of pairwise genetic differences between individuals) are similar in shape to those for humans, implying similar population histories of recent demographic expansion. The overall pattern of genetic variability in eastern chimpanzees is consistent with the hypothesis that the subject has responded demographically to paleoclimatically driven changes in the distribution of eastern African forests during the recent Pleistocene.