Species Identification of Tarantulas using Exuviae for International Wildlife Law Enforcement

This paper outlines a novel, non-invasive procedure to obtain DNA from Mexican tarantulas (Brachypelma spp.) using exuvia. These species are important in the pet trade and species identification is important for international wildlife law enforcement. Mitochondrial DNA sequence from the cytochrome c oxidase subunit I gene was used to investigate the relationship between various Brachypelma spp. This phylogeny was used as a framework to assign unknown specimens and spiderlings to species. The benefits to conservation, research, and international wildlife law enforcement that are gained by the ability to accurately identify species without the death of the specimen are explored. Our data also suggest that there is no support for the genus Brachypelmides as some authors have proposed and upholds the synonymy of Locht et al. (1999) J Arachnol 27:196–200.     

Mitochondria in stem cells

The current status of knowledge about mitochondrial properties in mouse, monkey and human embryonic, adult and precursor stem cells is discussed. Topics include mitochondrial localization patterns, oxygen consumption and ATP content in cells as they relate to the maintenance of stem cell properties and subsequent differentiation of stem cells into specific cell types. The significance of the perinuclear arrangement of mitochondria, which may be a characteristic feature of stem cells, as well as the expression of mitochondrial DNA regulatory proteins and mutations in the mitochondrial stem cell genome is also discussed.     

Shotgun glycopeptide capture approach coupled with mass spectrometry for comprehensive glycoproteomics

We present a robust and general shotgun glycoproteomics approach to comprehensively profile glycoproteins in complex biological mixtures. In this approach, glycopeptides derived from glycoproteins are enriched by selective capture onto a solid support using hydrazide chemistry followed by enzymatic release of the peptides and subsequent analysis by tandem mass spectrometry. The approach was validated using standard protein mixtures that resulted in a close to 100% capture efficiency. Our capture approach was then applied to microsomal fractions of the cisplatin-resistant ovarian cancer cell line IGROV-1/CP. With a Protein Prophet probability value greater than 0.9, we identified a total of 302 proteins with an average protein identification rate of 136 +/- 19 (n = 4) in a single linear quadrupole ion trap (LTQ) mass spectrometer nano-LC-MS experiment and a selectivity of 91 +/- 1.6% (n = 4) for the N-linked glycoconsensus sequence. Our method has several advantages. 1) Digestion of proteins initially into peptides improves the solubility of large membrane proteins and exposes all of the glycosylation sites to ensure equal accessibility to capture reagents. 2) Capturing glycosylated peptides can effectively reduce sample complexity and at the same time increase the confidence of MS-based protein identifications (more potential peptide identifications per protein). 3) The utility of sodium sulfite as a quencher in our capture approach to replace the solid phase extraction step in an earlier glycoprotein chemical capture approach for removing excess sodium periodate allows the overall capture procedure to be completed in a single vessel. This improvement minimizes sample loss, increases sensitivity, and makes our protocol amenable for high throughput implementation, a feature that is essential for biomarker identification and validation of a large number of clinical samples. 4) The approach is demonstrated here on the analysis of N-linked glycopeptides; however, it can be applied equally well to O-glycoprotein analysis.     

N-Glycan structure annotation of glycopeptides using a linearized glycan structure database (GlyDB)

While glycoproteins are abundant in nature, and changes in glycosylation occur in cancer and other diseases, glycoprotein characterization remains a challenge due to the structural complexity of the biopolymers. This paper presents a general strategy, termed GlyDB, for glycan structure annotation of N-linked glycopeptides from tandem mass spectra in the LC-MS analysis of proteolytic digests of glycoproteins. The GlyDB approach takes advantage of low-energy collision-induced dissociation of N-linked glycopeptides that preferentially cleaves the glycosidic bonds while the peptide backbone remains intact. A theoretical glycan structure database derived from biosynthetic rules for N-linked glycans was constructed employing a novel representation of branched glycan structures consisting of multiple linear sequences. The commonly used peptide identification program, Sequest, could then be utilized to assign experimental tandem mass spectra to individual glycoforms. Analysis of synthetic glycopeptides and well-characterized glycoproteins demonstrate that the GlyDB approach can be a useful tool for annotation of glycan structures and for selection of a limited number of potential glycan structure candidates for targeted validation.     

Putting beta-diversity on the map: broad-scale congruence and coincidence in the extremes

Beta-diversity, the change in species composition between places, is a critical but poorly understood component of biological diversity. Patterns of beta-diversity provide information central to many ecological and evolutionary questions, as well as to conservation planning. Yet beta-diversity is rarely studied across large extents, and the degree of similarity of patterns among taxa at such scales remains untested. To our knowledge, this is the first broad-scale analysis of cross-taxon congruence in beta-diversity, and introduces a new method to map beta-diversity continuously across regions. Congruence between amphibian, bird, and mammal beta-diversity in the Western Hemisphere varies with both geographic location and spatial extent. We demonstrate that areas of high beta-diversity for the three taxa largely coincide, but areas of low beta-diversity exhibit little overlap. These findings suggest that similar processes lead to high levels of differentiation in amphibian, bird, and mammal assemblages, while the ecological and biogeographic factors influencing homogeneity in vertebrate assemblages vary. Knowledge of beta-diversity congruence can help formulate hypotheses about the mechanisms governing regional diversity patterns and should inform conservation, especially as threat from global climate change increases.     

Protein oxidation implicated as the primary determinant of bacterial radioresistance

In the hierarchy of cellular targets damaged by ionizing radiation (IR), classical models of radiation toxicity place DNA at the top. Yet, many prokaryotes are killed by doses of IR that cause little DNA damage. Here we have probed the nature of Mn-facilitated IR resistance in Deinococcus radiodurans, which together with other extremely IR-resistant bacteria have high intracellular Mn/Fe concentration ratios compared to IR-sensitive bacteria. For in vitro and in vivo irradiation, we demonstrate a mechanistic link between Mn(II) ions and protection of proteins from oxidative modifications that introduce carbonyl groups. Conditions that inhibited Mn accumulation or Mn redox cycling rendered D. radiodurans radiation sensitive and highly susceptible to protein oxidation. X-ray fluorescence microprobe analysis showed that Mn is globally distributed in D. radiodurans, but Fe is sequestered in a region between dividing cells. For a group of phylogenetically diverse IR-resistant and IR-sensitive wild-type bacteria, our findings support the idea that the degree of resistance is determined by the level of oxidative protein damage caused during irradiation. We present the case that protein, rather than DNA, is the principal target of the biological action of IR in sensitive bacteria, and extreme resistance in Mn-accumulating bacteria is based on protein protection.     

Use of simulated data sets to evaluate the fidelity of metagenomic processing methods

Metagenomics is a rapidly emerging field of research for studying microbial communities. To evaluate methods presently used to process metagenomic sequences, we constructed three simulated data sets of varying complexity by combining sequencing reads randomly selected from 113 isolate genomes. These data sets were designed to model real metagenomes in terms of complexity and phylogenetic composition. We assembled sampled reads using three commonly used genome assemblers (Phrap, Arachne and JAZZ), and predicted genes using two popular gene-finding pipelines (fgenesb and CRITICA/GLIMMER). The phylogenetic origins of the assembled contigs were predicted using one sequence similarity-based (blast hit distribution) and two sequence composition-based (PhyloPythia, oligonucleotide frequencies) binning methods. We explored the effects of the simulated community structure and method combinations on the fidelity of each processing step by comparison to the corresponding isolate genomes. The simulated data sets are available online to facilitate standardized benchmarking of tools for metagenomic analysis.