Enhancer trap expression patterns provide a novel teaching resource

A collection of Arabidopsis enhancer trap transposants has been identified for use as a teaching tool. This collection serves to assist students in understanding the patterning and organization of plant tissues and cells, and will be useful in plant anatomy, morphology, and developmental biology courses. Each transposant exhibits reporter gene expression in a specific tissue, cell type, or domain, and these lines collectively offer a glimpse of compartments of gene expression. Some compartments correspond to classical definitions of botanical anatomy and can assist in anatomical identification. Other patterns of reporter gene expression are more complex and do not necessarily correspond to known anatomical features. The sensitivity of the beta-glucuronidase histochemical stain provides the student with a colorful and direct way to visualize difficult aspects of plant development and anatomy, and provides the teacher with an invaluable tool for a practical laboratory session.     

New approaches for high-yield purification of Müllerian inhibiting substance improve its bioactivity

We have established a new method to purify Müllerian inhibiting substance (MIS) with higher purity and recovery over existing procedures. Recombinant human MIS was expressed in Chinese hamster ovary cells and secreted into chemically defined serum-free media. The secreted products were concentrated by either precipitation with ammonium sulfate or lectin-affinity chromatography, each of which was followed by anion-exchange chromatography. Further separation of the active carboxy-terminal domain of MIS was achieved after cleavage by plasmin followed by lectin-affinity chromatography. This method may be applicable to other members of the transforming growth factor beta family with which MIS shares sequence homology.     

Inferring protein interactions from phylogenetic distance matrices

Finding the interacting pairs of proteins between two different protein families whose members are known to interact is an important problem in molecular biology. We developed and tested an algorithm that finds optimal matches between two families of proteins by comparing their distance matrices. A distance matrix provides a measure of the sequence similarity of proteins within a family. Since the protein sets of interest may have dozens of proteins each, the use of an efficient approximate solution is necessary. Therefore the approach we have developed consists of a Metropolis Monte Carlo optimization algorithm which explores the search space of possible matches between two distance matrices. We demonstrate that by using this algorithm we are able to accurately match chemokines and chemokine-receptors as well as the tgfbeta family of ligands and their receptors.     

Stomatal neighbor cell polarity and division in <Emphasis Type="Italic">Arabidopsis</Emphasis>

Asymmetric divisions are key to regulating the number and patterning of stomata in Arabidopsis thaliana (L.) Heynh. Many formative asymmetric divisions take place in neighbor cells (NCs), cells adjacent to a stoma or stomatal precursor. TOO MANY MOUTHS is a receptor-like protein required for the correct plane of NC division, resulting in the placement of the new precursor distal to the pre-existing stoma. Because plant cells usually become polarized before asymmetric division, we studied whether NCs display a cytological asymmetry as a function of cell stage and of possible division behavior. Cells that divided in the developing leaf epidermis were smaller than 400 micro m(-2) in area and included NCs as well as isolated cells. All NCs in the youngest complexes divided with comparable frequencies, but divisions became restricted to the smaller and most recently produced NCs as the stomatal complex matured. The majority of developing NCs had distally located nuclei, suggesting that nuclear position is actively regulated in NCs. NC stages exhibiting distally located nuclei were the likeliest to divide asymmetrically. However, a distal nucleus did not necessarily predict an asymmetric division, because more NCs had distal nuclei than were likely to divide. No defect was detected in nuclear distribution in tmm NCs. These data suggest that TMM uses intercellular signals to control the plane of asymmetric division after or independently of nuclear positioning.     

Viewing and annotating sequence data with Artemis

Artemis is a widely used software tool for annotating and viewing sequence data. No database is required to use Artemis. Instead, individual sequence data files can be analysed with little or no formatting, making it particularly suited to the study of small genomes and chromosomes, and straightforward for a novice user to get started. Since its release in 1999, Artemis has been used to annotate a diverse collection of prokaryotic and eukaryotic genomes, ranging from Streptomyces coelicolor to, more recently, a large proportion of the Plasmodium falciparum genome. Artemis allows annotated genomes to be easily browsed and makes it simple to add useful biological information to raw sequence data. This paper gives an overview of some of the features of Artemis and includes how it facilitates manual gene prediction and can provide an overview of entire chromosomes or small compact genomes--useful for uncovering unusual features such as pathogenicity islands.     

Retinoic acid-induced developmental defects are mediated by RARbeta/RXR heterodimers in the pharyngeal endoderm

Fusion and hypoplasia of the first two branchial arches, a defect typically observed in retinoic acid (RA) embryopathy, is generated in cultured mouse embryos upon treatment with BMS453, a synthetic compound that exhibits retinoic acid receptor beta (RARbeta) agonistic properties in transfected cells. By contrast, no branchial arch defects are observed following treatment with synthetic retinoids that exhibit RARalpha or RARgamma agonistic properties. The BMS453-induced branchial arch defects are mediated through RAR activation, as they are similar to those generated by a selective pan-RAR agonist, are prevented by a selective pan-RAR antagonist and cannot be mimicked by exposure to a pan-RXR agonist alone. They are enhanced in the presence of a pan-RXR agonist, and cannot be generated in Rarb-null embryos. Furthermore, they are accompanied, in the morphologically altered region, by ectopic expression of Rarb and of several other direct RA target genes. Therefore, craniofacial abnormalities characteristic of the RA embryopathy are mediated through ectopic activation of RARbeta/RXR heterodimers, in which the ligand-dependent activity of RXR is subordinated to that of RARbeta. Endodermal cells lining the first two branchial arches respond to treatment with the RARbeta agonist, in contrast to neural crest cells and ectoderm, which suggests that a faulty endodermal regionalization is directly responsible for RA-induced branchial arch dysmorphologies. Additionally, we provide the first in vivo evidence that the synthetic RARbeta agonist BMS453 exhibits an antagonistic activity on the two other RAR isotypes.     

EpsR Modulates Production of Extracellular Polysaccharides in the Bacterial Wilt Pathogen Ralstonia (Pseudomonas) solanacearum

Ralstonia solanacearum is the causal agent of bacterial wilt of many agriculturally important crops. Exopolysaccharide synthesized by products of the epsI operon is the major virulence factor for R. solanacearum. Expression of epsI has been demonstrated to be under the control of several proteins, including several two-component regulators. Overexpression of EpsR was found previously to reduce the amount of synthesis specifically from the epsI promoter. Here we present data that a single chromosomal copy of epsR activates the epsI promoter, suggesting that EpsR is a concentration-dependent effector of epsI gene expression. Furthermore, the ability of EpsR to modulate epsI expression is dependent on the phosphorylation state of EpsR. Gel mobility shift assays suggest that EpsR can specifically bind the epsI promoter and that this binding requires a phosphorylated form of EpsR.     

The Quantity and Turnover of Dead Wood in Permanent Forest Plots in Six Life Zones of Venezuela1

Dead wood can be an important component of the carbon pool in many forests, but few measurements have been made of this pool in tropical forests, To fill this gap, we determined the quantity of dead wood (downed and standing dead) in 25 long-term (up to 30 yr) permanent forest plots located in six different life zones of Venezuela. Downed wood was separated into fine (< 10 cm in diameter) and coarse (≥ 10 cm in diameter) classes, and three decomposition states (sound, intermediate, or rotten). The total quantity of dead wood, averaged by life zone, was lowest in the dry (2.43 Mg/ha), reached a peak in the moist (42.33 Mg/ha) and decreased slightly in the wet (34.50 Mg/ha) life zone. Most of the dead wood was in the standing dead category (about 42–76% of the total). The decomposition state of dead wood in all plots was mostly rotten (45%) or intermediate (44%); there was little sound wood (11%). Turnover rates of dead wood generally ranged between 0.03/yr to 0.52/yr with no clear trend with life zone. The large amount of dead wood in some plots was equivalent to about 20 percent or less of aboveground biomass, indicating that dead wood can represent a significant amount of carbon in these forests.     

Graphical Aids to the Estimation and Discrimination of Uncertain Numerical Data

This research investigates the performance of graphical dot arrays designed to make discrimination of relative numerosity as effortless as possible at the same time as making absolute (quantitative) numerosity estimation as effortful as possible. Comparing regular, random, and hybrid (randomized regular) configurations of dots, the results indicate that both random and hybrid configurations reduce absolute numerosity estimation precision, when compared with regular dots arrays. However, discrimination of relative numerosity is significantly more accurate for hybrid dot arrays than for random dot arrays. Similarly, human subjects report significantly lower levels of subjective confidence in judgments when using hybrid dot configurations as compared with regular configurations; and significantly higher levels of subjective confidence as compared with random configurations. These results indicate that data graphics based on the hybrid, randomized-regular configurations of dots are well-suited to applications that require decisions to be based on numerical data in which the absolute quantities are less certain than the relative values. Examples of such applications include decision-making based on the outputs of empirically-based mathematical models, such as health-related policy decisions using data from predictive epidemiological models.