Structure of the proton-gated urea channel from the gastric pathogen Helicobacter pylori.

Half the world's population is chronically infected with Helicobacter pylori, causing gastritis, gastric ulcers and an increased incidence of gastric adenocarcinoma. Its proton-gated inner-membrane urea channel, HpUreI, is essential for survival in the acidic environment of the stomach. The channel is closed at neutral pH and opens at acidic pH to allow the rapid access of urea to cytoplasmic urease. Urease produces NH(3) and CO(2), neutralizing entering protons and thus buffering the periplasm to a pH of roughly 6.1 even in gastric juice at a pH below 2.0. Here we report the structure of HpUreI, revealing six protomers assembled in a hexameric ring surrounding a central bilayer plug of ordered lipids. Each protomer encloses a channel formed by a twisted bundle of six transmembrane helices. The bundle defines a previously unobserved fold comprising a two-helix hairpin motif repeated three times around the central axis of the channel, without the inverted repeat of mammalian-type urea transporters. Both the channel and the protomer interface contain residues conserved in the AmiS/UreI superfamily, suggesting the preservation of channel architecture and oligomeric state in this superfamily. Predominantly aromatic or aliphatic side chains line the entire channel and define two consecutive constriction sites in the middle of the channel. Mutation of Trp 153 in the cytoplasmic constriction site to Ala or Phe decreases the selectivity for urea in comparison with thiourea, suggesting that solute interaction with Trp 153 contributes specificity. The previously unobserved hexameric channel structure described here provides a new model for the permeation of urea and other small amide solutes in prokaryotes and archaea

Angiosperm phylogenetic diversity is lower in Africa than South America

Although originating from a common Gondwanan flora, the diversity and composition of the floras of Africa and South America have greatly diverged since continental breakup of Africa from South America now having much higher plant species richness. However, the phylogenetic diversity of the floras and what this tells us about their evolution remained unexplored. We show that for a given species richness and considering land surface area, topography, and present-day climate, angiosperm phylogenetic diversity in South America is higher than in Africa. This relationship holds regardless of whether all climatically matched areas or only matched areas in tropical climates are considered. Phylogenetic diversity is high relative to species richness in refugial areas in Africa and in northwestern South America, once the gateway for immigration from the north. While species richness is strongly influenced by massive plant radiations in South America, we detect a pervasive influence of historical processes on the phylogenetic diversity of both the South American and African floras

Successful Flash-Cooling of Xenon Derivatized Myoglobin Crystals

This paper demonstrates for the first time a method for preparing cryocooled xenon-derivatized protein crystals. The method is based upon the hypothesis and subsequent observation that the diffusion of a xenon atom from a tight binding site following depressurization occurs on a timescale of minutes. We have observed significant changes in diffraction intensities from myoglobin crystals for up to 5 min following depressurization from 1 MPa of xenon. In accordance with this observation, a xenon-derivatized myoglobin crystal was cryocooled at ~95 K within 20 s of complete depressurization. A crystallographic data set was then collected to 2.0 Å resolution and isomorphous and anomalous difference Patterson maps revealed the presence of a well ordered xenon site with an occupancy of approximately 0.5. Phasing statistics for this site were of good quality and demonstrate the practicality of this method. The ability to cryocool xenon-derivatized crystals will make this heavy-atom substitution method even more useful for single-isomorphous-replacement and multiple-isomorphous-replacement phasing of macromolecules

Review of the Application of Modern Cytogenetic Methods (FISH/GISH) to the Study of Reticulation (Polyploidy/Hybridisation).

The convergence of distinct lineages upon interspecific hybridisation, including when accompanied by increases in ploidy (allopolyploidy), is a driving force in the origin of many plant species. In plant breeding too, both interspecific hybridisation and allopolyploidy are important because they facilitate introgression of alien DNA into breeding lines enabling the introduction of novel characters. Here we review how fluorescence in situ hybridisation (FISH) and genomic in situ hybridisation (GISH) have been applied to: 1) studies of interspecific hybridisation and polyploidy in nature, 2) analyses of phylogenetic relationships between species, 3) genetic mapping and 4) analysis of plant breeding materials. We also review how FISH is poised to take advantage of nextgeneration sequencing (NGS) technologies, helping the rapid characterisation of the repetitive fractions of a genome in natural populations and agricultural plants.This work was supported by NSF grant DEB-0922003

Complete Plastid Genome Sequencing of Trochodendraceae Reveals a Significant Expansion of the Inverted Repeat and Suggests a Paleogene Divergence between the Two Extant Species

The early-diverging eudicot order Trochodendrales contains only two monospecific genera, Tetracentron and Trochodendron. Although an extensive fossil record indicates that the clade is perhaps 100 million years old and was widespread throughout the Northern Hemisphere during the Paleogene and Neogene, the two extant genera are both narrowly distributed in eastern Asia. Recent phylogenetic analyses strongly support a clade of Trochodendrales, Buxales, and Gunneridae (core eudicots), but complete plastome analyses do not resolve the relationships among these groups with strong support. However, plastid phylogenomic analyses have not included data for Tetracentron. To better resolve basal eudicot relationships and to clarify when the two extant genera of Trochodendrales diverged, we sequenced the complete plastid genome of Tetracentron sinense using Illumina technology. The Tetracentron and Trochodendron plastomes possess the typical gene content and arrangement that characterize most angiosperm plastid genomes, but both genomes have the same unusual ~4 kb expansion of the inverted repeat region to include five genes (rpl22, rps3, rpl16, rpl14, and rps8) that are normally found in the large single-copy region. Maximum likelihood analyses of an 83-gene, 88 taxon angiosperm data set yield an identical tree topology as previous plastid-based trees, and moderately support the sister relationship between Buxaceae and Gunneridae. Molecular dating analyses suggest that Tetracentron and Trochodendron diverged between 44-30 million years ago, which is congruent with the fossil record of Trochodendrales and with previous estimates of the divergence time of these two taxa. We also characterize 154 simple sequence repeat loci from the Tetracentron sinense and Trochodendron aralioides plastomes that will be useful in future studies of population genetic structure for these relict species, both of which are of conservation concern

Structural changes that occur upon photolysis of the Fe(II)a3–CO complex in the cytochrome ba3-oxidase of Thermus thermophilus: A combined X-ray crystallographic and infrared spectral study demonstrates CO binding to CuB

AbstractThe purpose of the work was to provide a crystallographic demonstration of the venerable idea that CO photolyzed from ferrous heme-a3 moves to the nearby cuprous ion in the cytochrome c oxidases. Crystal structures of CO-bound cytochrome ba3-oxidase from Thermus thermophilus, determined at ~2.8–3.2Å resolution, reveal a Fe–C distance of ~2.0Å, a Cu–O distance of 2.4Å and a Fe–C–O angle of ~126°. Upon photodissociation at 100K, X-ray structures indicate loss of Fea3–CO and appearance of CuB–CO having a Cu–C distance of ~1.9Å and an O–Fe distance of ~2.3Å. Absolute FTIR spectra recorded from single crystals of reduced ba3–CO that had not been exposed to X-ray radiation, showed several peaks around 1975cm−1; after photolysis at 100K, the absolute FTIR spectra also showed a significant peak at 2050cm−1. Analysis of the ‘light’ minus ‘dark’ difference spectra showed four very sharp CO stretching bands at 1970cm−1, 1977cm−1, 1981cm−1, and 1985cm−1, previously assigned to the Fea3–CO complex, and a significantly broader CO stretching band centered at ~2050cm−1, previously assigned to the CO stretching frequency of CuB bound CO. As expected for light propagating along the tetragonal axis of the P43212 space group, the single crystal spectra exhibit negligible dichroism. Absolute FTIR spectrometry of a CO-laden ba3 crystal, exposed to an amount of X-ray radiation required to obtain structural data sets before FTIR characterization, showed a significant signal due to photogenerated CO2 at 2337cm−1 and one from traces of CO at 2133cm−1; while bands associated with CO bound to either Fea3 or to CuB in “light” minus “dark” FTIR difference spectra shifted and broadened in response to X-ray exposure. In spite of considerable radiation damage to the crystals, both X-ray analysis at 2.8 and 3.2Å and FTIR spectra support the long-held position that photolysis of Fea3–CO in cytochrome c oxidases leads to significant trapping of the CO on the CuB atom; Fea3 and CuB ligation, at the resolutions reported here, are otherwise unaltered. This article is part of a Special Issue entitled: Respiratory Oxidases

STAT3 in the systemic inflammation of cancer cachexia

Weight loss is diagnostic of cachexia, a debilitating syndrome contributing mightily to morbidity and mortality in cancer. Most research has probed mechanisms leading to muscle atrophy and adipose wasting in cachexia; however cachexia is a truly systemic phenomenon. Presence of the tumor elicits an inflammatory response and profound metabolic derangements involving not only muscle and fat, but also the hypothalamus, liver, heart, blood, spleen and likely other organs. This global response is orchestrated in part through circulating cytokines that rise in conditions of cachexia. Exogenous Interleukin-6 (IL6) and related cytokines can induce most cachexia symptomatology, including muscle and fat wasting, the acute phase response and anemia, while IL-6 inhibition reduces muscle loss in cancer. Although mechanistic studies are ongoing, certain of these cachexia phenotypes have been causally linked to the cytokine-activated transcription factor, STAT3, including skeletal muscle wasting, cardiac dysfunction and hypothalamic inflammation. Correlative studies implicate STAT3 in fat wasting and the acute phase response in cancer cachexia. Parallel data in non-cancer models and disease states suggest both pathological and protective functions for STAT3 in other organs during cachexia. STAT3 also contributes to cancer cachexia through enhancing tumorigenesis, metastasis and immune suppression, particularly in tumors associated with high prevalence of cachexia. This review examines the evidence linking STAT3 to multi-organ manifestations of cachexia and the potential and perils for targeting STAT3 to reduce cachexia and prolong survival in cancer patients

Another look at the root of the angiosperms reveals a familiar tale

Since the advent of molecular phylogenetics more than 25 years ago, a major goal of plant systematists has been to discern the root of the angiosperms. While most studies indicate that Amborella trichopoda is sister to all remaining extant flowering plants, support for this position has varied with respect to both the sequence data sets and analyses employed. Recently, Goremykin et al. (2013) questioned the “Amborella-sister hypothesis” using a “noise-reduction” approach and reported a topology with Amborella + Nymphaeales (water lilies) sister to all remaining angiosperms. Through a series of analyses of both plastid genomes and mitochondrial genes, we continue to find mostly strong support for the Amborella-sister hypothesis and offer a rebuttal of Goremykin et al. (2013). The major tenet of Goremykin et al. is that the Amborella-sister position is determined by noisy data – i.e. characters with high rates of change and lacking true phylogenetic signal. To investigate the signal in these noisy data further, we analyzed the discarded characters from their noise-reduced alignments. We recovered a tree identical to that of the currently accepted angiosperm framework, including the position of Amborella as sister to all other angiosperms, as well as all other major clades. Thus, the signal in the “noisy” data is consistent with that of our complete data sets – arguing against the use of their noise-reduction approach. We also determined that one of the alignments presented by Goremykin et al. yields results at odds with their central claim – their data set actually supports Amborella as sister to all other angiosperms, as do larger plastid data sets we present here that possess more complete taxon sampling both within the monocots and for angiosperms in general. Previous unpartitioned, multi-locus analyses of mtDNA data have provided the strongest support for Amborella + Nymphaeales as sister to other angiosperms. However, our analysis of third codon positions from mtDNA sequence data also supports the Amborella-sister hypothesis. Finally, we challenge the conclusion of Goremykin et al. that the first flowering plants were aquatic and herbaceous, reasserting that even if Amborella + water lilies, or water lilies alone, are sister to the rest of the angiosperms, the earliest angiosperms were not necessarily aquatic and/or herbaceous

Angiosperm Phylogeny: 17 Genes, 640 Taxa

• Premise of the study : Recent analyses employing up to fi ve genes have provided numerous insights into angiosperm phylogeny, but many relationships have remained unresolved or poorly supported. In the hope of improving our understanding of angiosperm phylogeny, we expanded sampling of taxa and genes beyond previous analyses. • Methods : We conducted two primary analyses based on 640 species representing 330 families. The fi rst included 25 260 aligned base pairs (bp) from 17 genes (representing all three plant genomes, i.e., nucleus, plastid, and mitochondrion). The second included 19 846 aligned bp from 13 genes (representing only the nucleus and plastid). • Key results : Many important questions of deep-level relationships in the nonmonocot angiosperms have now been resolved with strong support. Amborellaceae, Nymphaeales, and Austrobaileyales are successive sisters to the remaining angiosperms ( Mesangiospermae ), which are resolved into Chloranthales + Magnoliidae as sister to Monocotyledoneae + [Ceratophyllaceae + Eudicotyledoneae ]. Eudicotyledoneae contains a basal grade subtending Gunneridae . Within Gunneridae , Gunnerales are sister to the remainder ( Pentapetalae ), which comprises (1) Superrosidae , consisting of Rosidae (including Vitaceae) and Saxifragales; and (2) Superasteridae , comprising Berberidopsidales, Santalales, Caryophyllales , Asteridae , and, based on this study, Dilleniaceae (although other recent analyses disagree with this placement). Within the major subclades of Pentapetalae , most deep-level relationships are resolved with strong support. • Conclusions : Our analyses confi rm that with large amounts of sequence data, most deep-level relationships within the angiosperms can be resolved. We anticipate that this well-resolved angiosperm tree will be of broad utility for many areas of biology, including physiology, ecology, paleobiology, and genomics