Binding specificity of Bacillus thuringiensis Cry1Aa for purified, native Bombyx mori aminopeptidase N and cadherin-like receptors

BACKGROUND: To better understand the molecular interactions of Bt toxins with non-target insects, we have examined the real-time binding specificity and affinity of Cry1 toxins to native silkworm (Bombyx mori) midgut receptors. Previous studies on B. mori receptors utilized brush border membrane vesicles or purifed receptors in blot-type assays. RESULTS: The Bombyx mori (silkworm) aminopeptidase N (APN) and cadherin-like receptors for Bacillus thuringiensis insecticidal Cry1Aa toxin were purified and their real-time binding affinities for Cry toxins were examined by surface plasmon resonance. Cry1Ab and Cry1Ac toxins did not bind to the immobilized native receptors, correlating with their low toxicities. Cry1Aa displayed moderate affinity for B. mori APN (75 nM), and unusually tight binding to the cadherin-like receptor (2.6 nM), which results from slow dissociation rates. The binding of a hybrid toxin (Aa/Aa/Ac) was identical to Cry1Aa. CONCLUSIONS: These results indicate domain II of Cry1Aa is essential for binding to native B. mori receptors and for toxicity. Moreover, the high-affinity binding of Cry1Aa to native cadherin-like receptor emphasizes the importance of this receptor class for Bt toxin research

Characterization of Putative Iron Responsive Genes as Species-Specific Indicators of Iron Stress in Thalassiosiroid Diatoms

Iron (Fe) availability restricts diatom growth and primary production in large areas of the oceans. It is a challenge to assess the bulk Fe nutritional health of natural diatom populations, since species can differ in their physiological and molecular responses to Fe limitation. We assayed expression of selected genes in diatoms from the Thalassiosira genus to assess their potential utility as species-specific molecular markers to indicate Fe status in natural diatom assemblages. In this study, we compared the expression of the photosynthetic genes encoding ferredoxin (a Fe-requiring protein) and flavodoxin (a Fe-free protein) in culture experiments with Fe replete and Fe stressed Thalassiosira pseudonana (CCMP 1335) isolated from coastal waters and Thalassiosira weissflogii (CCMP 1010) isolated from the open ocean. In T. pseudonana, expression of flavodoxin and ferredoxin genes were not sensitive to Fe status but were found to display diel periodicities. In T. weissflogii, expression of flavodoxin was highly responsive to iron levels and was only detectable when cultures were Fe limited. Flavodoxin genes have been duplicated in most diatoms with available genome data and we show that T. pseudonana has lost its copy related to the Fe-responsive copy in T. weissflogii. We also examined the expression of genes for a putative high affinity, copper (Cu)-dependent Fe uptake system in T. pseudonana. Our results indicate that genes encoding putative Cu transporters, a multi-Cu oxidase, and a Fe reductase are not linked to Fe status. The expression of a second putative Fe reductase increased in Fe limited cultures, but this gene was also highly expressed in Fe replete cultures, indicating it may not be a useful marker in the field. Our findings highlight that Fe metabolism may differ among diatoms even within a genus and show a need to validate responses in different species as part of the development pipeline for genetic markers of Fe status in field populations

SNP Assay Development for Linkage Map Construction, Anchoring Whole-Genome Sequence, and Other Genetic and Genomic Applications in Common Bean.

A total of 992,682 single-nucleotide polymorphisms (SNPs) was identified as ideal for Illumina Infinium II BeadChip design after sequencing a diverse set of 17 common bean (Phaseolus vulgaris L) varieties with the aid of next-generation sequencing technology. From these, two BeadChips each with >5000 SNPs were designed. The BARCBean6K_1 BeadChip was selected for the purpose of optimizing polymorphism among market classes and, when possible, SNPs were targeted to sequence scaffolds in the Phaseolus vulgaris 14× genome assembly with sequence lengths >10 kb. The BARCBean6K_2 BeadChip was designed with the objective of anchoring additional scaffolds and to facilitate orientation of large scaffolds. Analysis of 267 F2 plants from a cross of varieties Stampede × Red Hawk with the two BeadChips resulted in linkage maps with a total of 7040 markers including 7015 SNPs. With the linkage map, a total of 432.3 Mb of sequence from 2766 scaffolds was anchored to create the Phaseolus vulgaris v1.0 assembly, which accounted for approximately 89% of the 487 Mb of available sequence scaffolds of the Phaseolus vulgaris v0.9 assembly. A core set of 6000 SNPs (BARCBean6K_3 BeadChip) with high genotyping quality and polymorphism was selected based on the genotyping of 365 dry bean and 134 snap bean accessions with the BARCBean6K_1 and BARCBean6K_2 BeadChips. The BARCBean6K_3 BeadChip is a useful tool for genetics and genomics research and it is widely used by breeders and geneticists in the United States and abroad

Blocking binding of Bacillus thuringiensis Cry1Aa to Bombyx mori cadherin receptor results in only a minor reduction of toxicity

<p>Abstract</p> <p>Background</p> <p><it>Bacillus thuringiensis </it>Cry1Aa insecticidal protein is the most active known <it>B. thuringiensis </it>toxin against the forest insect pest <it>Lymantria dispar </it>(gypsy moth), unfortunately it is also highly toxic against the non-target insect <it>Bombyx mori </it>(silk worm).</p> <p>Results</p> <p>Surface exposed hydrophobic residues over domains II and III were targeted for site-directed mutagenesis. Substitution of a phenylalanine residue (F328) by alanine reduced binding to the <it>Bombyx mori </it>cadherin by 23-fold, reduced biological activity against <it>B. mori </it>by 4-fold, while retaining activity against <it>Lymantria dispar</it>.</p> <p>Conclusion</p> <p>The results identify a novel receptor-binding epitope and demonstrate that virtual elimination of binding to cadherin BR-175 does not completely remove toxicity in the case of <it>B. mori</it>.</p

A New H I Survey of Active Galaxies

We have conducted a new Arecibo survey for H I emission for 113 galaxies with broad-line (type 1) active galactic nuclei (AGNs) out to recession velocities as high as 35,000 km/s. The primary aim of the study is to obtain sensitive H I spectra for a well-defined, uniformly selected sample of active galaxies that have estimates of their black hole masses in order to investigate correlations between H I properties and the characteristics of the AGNs. H I emission was detected in 66 out of the 101 (65%) objects with spectra uncorrupted by radio frequency interference, among which 45 (68%) have line profiles with adequate signal-to-noise ratio and sufficiently reliable inclination corrections to yield robust deprojected rotational velocities. This paper presents the basic survey products, including an atlas of H I spectra, measurements of H I flux, line width, profile asymmetry, optical images, optical spectroscopic parameters, as well as a summary of a number of derived properties pertaining to the host galaxies. To enlarge our primary sample, we also assemble all previously published H I measurements of type 1 AGNs for which can can estimate black hole masses, which total an additional 53 objects. The final comprehensive compilation of 154 broad-line active galaxies, by far the largest sample ever studied, forms the basis of our companion paper, which uses the H I database to explore a number of properties of the AGN host galaxies.Comment: To appear in ApJS; 31 pages. Preprint will full-resolution figures can be downloaded from http://www.ociw.edu/~lho/preprints/ms1.pd

Determination of minimal transcriptional signatures of compounds for target prediction

The identification of molecular target and mechanism of action of compounds is a key hurdle in drug discovery. Multiplexed techniques for bead-based expression profiling allow the measurement of transcriptional signatures of compound-treated cells in high-throughput mode. Such profiles can be used to gain insight into compounds' mode of action and the protein targets they are modulating. Through the proxy of target prediction from such gene signatures we explored important aspects of the use of transcriptional profiles to capture biological variability of perturbed cellular assays. We found that signatures derived from expression data and signatures derived from biological interaction networks performed equally well, and we showed that gene signatures can be optimised using a genetic algorithm. Gene signatures of approximately 128 genes seemed to be most generic, capturing a maximum of the perturbation inflicted on cells through compound treatment. Moreover, we found evidence for oxidative phosphorylation to be one of the most general ways to capture compound perturbation

Draft Nuclear Genome Sequence of the Liquid Hydrocarbon-Accumulating Green Microalga Botryococcus braunii Race B (Showa).

Botryococcus braunii has long been known as a prodigious producer of liquid hydrocarbon oils that can be converted into combustion engine fuels. This draft genome for the B race of B. braunii will allow researchers to unravel important hydrocarbon biosynthetic pathways and identify possible regulatory networks controlling this unusual metabolism

SNAVI: Desktop application for analysis and visualization of large-scale signaling networks

<p>Abstract</p> <p>Background</p> <p>Studies of cellular signaling indicate that signal transduction pathways combine to form large networks of interactions. Viewing protein-protein and ligand-protein interactions as graphs (networks), where biomolecules are represented as nodes and their interactions are represented as links, is a promising approach for integrating experimental results from different sources to achieve a systematic understanding of the molecular mechanisms driving cell phenotype. The emergence of large-scale signaling networks provides an opportunity for topological statistical analysis while visualization of such networks represents a challenge.</p> <p>Results</p> <p>SNAVI is Windows-based desktop application that implements standard network analysis methods to compute the clustering, connectivity distribution, and detection of network motifs, as well as provides means to visualize networks and network motifs. SNAVI is capable of generating linked web pages from network datasets loaded in text format. SNAVI can also create networks from lists of gene or protein names.</p> <p>Conclusion</p> <p>SNAVI is a useful tool for analyzing, visualizing and sharing cell signaling data. SNAVI is open source free software. The installation may be downloaded from: <url>http://snavi.googlecode.com</url>. The source code can be accessed from: <url>http://snavi.googlecode.com/svn/trunk</url></p

Properties of Active Galaxies Deduced from H I Observations

We completed a new survey for H I emission for a large, well-defined sample of 154 nearby (z < 0.1) galaxies with type 1 AGNs. We make use of the extensive database presented in a companion paper to perform a comprehensive appraisal of the cold gas content in active galaxies and to seek new strategies to investigate the global properties of the host galaxies and their relationship to their central black holes (BHs). We show that the BH mass obeys a strong, roughly linear relation with the host galaxy's dynamical mass. BH mass follows a looser, though still highly significant, correlation with the maximum rotation velocity of the galaxy, as expected from the known scaling between rotation velocity and central velocity dispersion. Neither of these H I-based correlations is as tight as the more familiar relations between BH mass and bulge luminosity or velocity dispersion, but they offer the advantage of being insensitive to the glare of the nucleus and therefore are promising new tools for probing the host galaxies of both nearby and distant AGNs. We present evidence for substantial ongoing BH growth in the most actively accreting AGNs. In these nearby systems, BH growth appears to be delayed with respect to the assembly of the host galaxy but otherwise has left no detectable perturbation to its mass-to-light ratio or its global gas content. The host galaxies of type 1 AGNs, including those luminous enough to qualify as quasars, are generally gas-rich systems, possessing a cold interstellar medium reservoir at least as abundant as that in inactive galaxies of the same morphological type. This calls into question current implementations of AGN feedback in models of galaxy formation that predict strong cold gas depletion in unobscured AGNs. (Abridged)Comment: To appear in ApJ; 14 page

Geographic Variation in Sexual Attraction of Spodoptera frugiperda Corn- and Rice-Strain Males to Pheromone Lures

The corn- and rice-strains of Spodoptera frugiperda exhibit several genetic and behavioral differences and appear to be undergoing ecological speciation in sympatry. Previous studies reported conflicting results when investigating male attraction to pheromone lures in different regions, but this could have been due to inter-strain and/or geographic differences. Therefore, we investigated whether corn- and rice-strain males differed in their response to different synthetic pheromone blends in different regions in North America, the Caribbean and South America. All trapped males were strain typed by two strain-specific mitochondrial DNA markers. In the first experiment, we found a nearly similar response of corn and rice-strain males to two different 4-component blends, resembling the corn- and rice-strain female blend we previously described from females in Florida. This response showed some geographic variation in fields in Canada, North Carolina, Florida, Puerto Rico, and South America (Peru, Argentina). In dose-response experiments with the critical secondary sex pheromone component (Z)-7-dodecenyl acetate (Z7-12:OAc), we found some strain-specific differences in male attraction. While the response to Z7-12:OAc varied geographically in the corn-strain, rice-strain males showed almost no variation. We also found that the minor compound (Z)-11-hexadecenyl acetate (Z11-16:OAc) did not increase attraction of both strains in Florida and of corn-strain males in Peru. In a fourth experiment, where we added the stereo-isomer of the critical sex pheromone component, (E)-7-dodecenyl acetate, to the major pheromone component (Z)-9-tetradecenyl acetate (Z9-14:OAc), we found that this compound was attractive to males in North Carolina, but not to males in Peru. Overall, our results suggest that both strains show rather geographic than strain-specific differences in their response to pheromone lures, and that regional sexual communication differences might cause geographic differentiation between populations.Fil: Unbehend, Melanie. Instituto Max Planck Institut Fur Chemische Okologie; AlemaniaFil: Hänniger, Sabine. Instituto Max Planck Institut Fur Chemische Okologie; AlemaniaFil: Vasquez, Gissella M.. University Of North Carolina; Estados UnidosFil: Juárez, María Laura. Gobierno de Tucumán. Ministerio de Desarrollo Productivo. Estación Experimental Agroindustrial Obispo Colombres; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán; ArgentinaFil: Reisig, Dominic. University Of North Carolina; Estados UnidosFil: Mcneil, Jeremy N.. University of Western Ontario. Department of Biology; CanadáFil: Meagher, Robert L.. United States Department Of Agriculture; Estados UnidosFil: Jenkins, David A.. United States Department of Agriculture; ArgentinaFil: Heckel, David G.. Instituto Max Planck Institut Fur Chemische Okologie; AlemaniaFil: Groot, Astrid T.. University Of Amsterdam; Países Bajos. Instituto Max Planck Institut Fur Chemische Okologie; Alemani