Gap junction structures: Analysis of the x-ray diffraction data

Models for the spatial distribution of protein, lipid and water in gap junction structures have been constructed from the results of the analysis of X-ray diffraction data described here and the electron microscope and chemical data presented in the preceding paper (Caspar, D. L. D., D. A. Goodenough, L. Makowski, and W.C. Phillips. 1977. 74:605-628). The continuous intensity distribution on the meridian of the X-ray diffraction pattern was measured, and corrected for the effects of the partially ordered stacking and partial orientation of the junctions in the X-ray specimens. The electron density distribution in the direction perpendicular to the plane of the junction was calculated from the meridional intensity data. Determination of the interference function for the stacking of the junctions improved the accuracy of the electron density profile. The pair-correlation function, which provides information about the packing of junctions in the specimen, was calculated from the interference function. The intensities of the hexagonal lattice reflections on the equator of the X-ray pattern were used in coordination with the electron microscope data to calculate to the two-dimensional electron density projection onto the plane of the membrane. Differences in the structure of the connexons as seen in the meridional profile and equatorial projections were shown to be correlated to changes in lattice constant. The parts of the junction structure which are variable have been distinguished from the invariant parts by comparison of the X-ray data from different specimens. The combination of these results with electron microscope and chemical data provides low resolution three- dimensional representations of the structures of gap junctions.     

Phylogenomics of the reproductive parasite Wolbachia pipientis wMel: a streamlined genome overrun by mobile genetic elements

The complete sequence of the 1,267,782 bp genome of Wolbachia pipientis wMel, an obligate intracellular bacteria of Drosophila melanogaster, has been determined. Wolbachia, which are found in a variety of invertebrate species, are of great interest due to their diverse interactions with different hosts, which range from many forms of reproductive parasitism to mutualistic symbioses. Analysis of the wMel genome, in particular phylogenomic comparisons with other intracellular bacteria, has revealed many insights into the biology and evolution of wMel and Wolbachia in general. For example, the wMel genome is unique among sequenced obligate intracellular species in both being highly streamlined and containing very high levels of repetitive DNA and mobile DNA elements. This observation, coupled with multiple evolutionary reconstructions, suggests that natural selection is somewhat inefficient in wMel, most likely owing to the occurrence of repeated population bottlenecks. Genome analysis predicts many metabolic differences with the closely related Rickettsia species, including the presence of intact glycolysis and purine synthesis, which may compensate for an inability to obtain ATP directly from its host, as Rickettsia can. Other discoveries include the apparent inability of wMel to synthesize lipopolysaccharide and the presence of the most genes encoding proteins with ankyrin repeat domains of any prokaryotic genome yet sequenced. Despite the ability of wMel to infect the germline of its host, we find no evidence for either recent lateral gene transfer between wMel and D. melanogaster or older transfers between Wolbachia and any host. Evolutionary analysis further supports the hypothesis that mitochondria share a common ancestor with the α-Proteobacteria, but shows little support for the grouping of mitochondria with species in the order Rickettsiales. With the availability of the complete genomes of both species and excellent genetic tools for the host, the wMel–D. melanogaster symbiosis is now an ideal system for studying the biology and evolution of Wolbachia infections.     

Hydrolysis of filter-paper cellulose to glucose by two recombinant endogenous glycosyl hydrolases of Coptotermes formosanus

Abstract Genes encoding for glycosyl hydrolases (GH) in multiple families were recovered from an expression sequence tag library of Coptotermes formosanus, a xylophagous lower termite species. Functional analyses of these genes not only shed light on the mechanisms the insect employs to successfully use cellulosic materials as energy sources, which may serve as strategic targets for designing molecular-based bio-pesticides, but also enrich discoveries of new cellulolytic enzymes for conversion of biomass into biofuel. Our study demonstrated that cellulose could be converted to glucose by two recombinant endogenous glycosyl hydrolases (endo-β-1,4 glucanase in GH9 and β-glucosidase in GH1). While the former cleaved cellulose to cellobiose and cellotriose, the resulting simple cellodextrins were digested to glucose. Both of the Escherichia coli-expressed recombinant proteins showed properties that could be incorporated in a glucose-based ethanol production program.     

Elucidation of <Emphasis Type="Italic">veA</Emphasis>-dependent genes associated with aflatoxin and sclerotial production in <Emphasis Type="Italic">Aspergillus flavus</Emphasis> by functional genomics

The aflatoxin-producing fungi, Aspergillus flavus and A. parasiticus, form structures called sclerotia that allow for survival under adverse conditions. Deletion of the veA gene in A. flavus and A. parasiticus blocks production of aflatoxin as well as sclerotial formation. We used microarray technology to identify genes differentially expressed in wild-type veA and veA mutant strains that could be involved in aflatoxin production and sclerotial development in A. flavus. The DNA microarray analysis revealed 684 genes whose expression changed significantly over time; 136 of these were differentially expressed between the two strains including 27 genes that demonstrated a significant difference in expression both between strains and over time. A group of 115 genes showed greater expression in the wild-type than in the veA mutant strain. We identified a subgroup of veA-dependent genes that exhibited time-dependent expression profiles similar to those of known aflatoxin biosynthetic genes or that were candidates for involvement in sclerotial production in the wild type.     

<Emphasis Type="Italic">Burkholderia</Emphasis> Hep_Hag autotransporter (BuHA) proteins elicit a strong antibody response during experimental glanders but not human melioidosis

Background  

The bacterial biothreat agents Burkholderia mallei and Burkholderia pseudomallei are the cause of glanders and melioidosis, respectively. Genomic and epidemiological studies have shown that B. mallei is a recently emerged, host restricted clone of B. pseudomallei.     

Genome-wide expression analysis of iron regulation in Burkholderia pseudomallei and Burkholderia mallei using DNA microarrays

Burkholderia pseudomallei and B. mallei are the causative agents of melioidosis and glanders, respectively. As iron regulation of gene expression is common in bacteria, in the present studies, we have used microarray analysis to examine the effects of growth in different iron concentrations on the regulation of gene expression in B. pseudomallei and B. mallei. Gene expression profiles for these two bacterial species were similar under high and low iron growth conditions irrespective of growth phase. Growth in low iron led to reduced expression of genes encoding most respiratory metabolic systems and proteins of putative function, such as NADH-dehydrogenases, cytochrome oxidases, and ATP-synthases. In contrast, genes encoding siderophore-mediated iron transport, heme-hemin receptors, and a variety of metabolic enzymes for alternative metabolism were induced under low iron conditions. The overall gene expression profiles suggest that B. pseudomallei and B. mallei are able to adapt to the iron-restricted conditions in the host environment by up-regulating an iron-acquisition system and by using alternative metabolic pathways for energy production. The observations relative to the induction of specific metabolic enzymes during bacterial growth under low iron conditions warrants further experimentation.