Novel 2-(2-(benzylthio)-1H-benzo[d]imidazol-1-yl)acetic acids: discovery and hit-to-lead evolution of a selective CRTh2 receptor antagonist chemotype

Hit-to-lead evolution of 2-(2-((2-(4-chlorophenoxy)ethyl)thio)-1H-benzo[d]imidazol-1-yl)acetic acid (1), discovered in a high-throughput screening campaign as a novel chemotype of CRTh2 receptor antagonist, is presented. SAR development as well as in vitro and in vivo DMPK properties of selected representatives of substituted 2-(2-(benzylthio)-1H-benzo[d]imidazol-1-yl)acetic acids are discussed.     

Construction and functionalization of fused pyridine ring leading to novel compounds as potential antitubercular agents

A series of fused and functionalized pyridine derivatives were designed, synthesized and tested for their potential antitubercular properties. All these novel compounds were prepared by using multistep methods involving the construction of pyridine ring as a key synthetic step. Some of these compounds were found to be interesting when tested for their antitubercular properties in vitro and one of them appeared as an attractive and potential antitubercular agent.     

Linking landscape history and dispersal traits in grassland plant communities

Dispersal limitation and long-term persistence are known to delay plant species’ responses to habitat fragmentation, but it is still unclear to what extent landscape history may explain the distribution of dispersal traits in present-day plant communities. We used quantitative data on long-distance seed dispersal potential by wind and grazing cattle (epi- and endozoochory), and on persistence (adult plant longevity and seed bank persistence) to quantify the linkages between dispersal and persistence traits in grassland plant communities and current and past landscape configurations. The long-distance dispersal potential of present-day communities was positively associated with the amounts of grassland in the historical (1835, 1938) landscape, and with a long continuity of grazing management—but was not associated with the properties of the current landscape. The study emphasises the role of history as a determinant of the dispersal potential of present-day grassland plant communities. The importance of long-distance dispersal processes has declined in the increasingly fragmented modern landscape, and long-term persistent species are expected to play a more dominant role in grassland communities in the future. However, even within highly fragmented landscapes, long-distance dispersed species may persist locally—delaying the repayment of the extinction debt.     

Genomics and Localization of the Arabidopsis DHHC-Cysteine-Rich Domain S-Acyltransferase Protein Family

Protein lipid modification of cysteine residues, referred to as S-palmitoylation or S-acylation, is an important secondary and reversible modification that regulates membrane association, trafficking, and function of target proteins. This enzymatic reaction is mediated by protein S-acyl transferases (PATs). Here, the phylogeny, genomic organization, protein topology, expression, and localization pattern of the 24 PAT family members from Arabidopsis (Arabidopsis thaliana) is described. Most PATs are expressed at ubiquitous levels and tissues throughout the development, while few genes are expressed especially during flower development preferentially in pollen and stamen. The proteins display large sequence and structural variations but exhibit a common protein topology that is preserved in PATs from various organisms. Arabidopsis PAT proteins display a complex targeting pattern and were detected at the endoplasmic reticulum, Golgi, endosomal compartments, and the vacuolar membrane. However, most proteins were targeted to the plasma membrane. This large concentration of plant PAT activity to the plasma membrane suggests that the plant cellular S-acylation machinery is functionally different compared with that of yeast (Saccharomyces cerevisiae) and mammalians.Cellular functions are regulated by various protein modifications. These modifications can occur in a controlled, reversible, and transient manner in response to different signals (Huber and Hardin, 2004; Stulemeijer and Joosten, 2008). Protein phosphorylation is the most prominent modification and is regulated by the balanced activities of protein kinases and protein phosphatases (Hardie, 1999; Luan, 2003). However, other modifications, like ubiquitination and sumoylation (Miura and Hasegawa, 2010) or nitrosylation (Lamattina et al., 2003), also regulate protein functions.A less well characterized reversible lipid modification is the thioesterification of Cys residues, which is known as S-palmitoylation or more generally referred to as S-acylation (Hemsley and Grierson, 2008; Sorek et al., 2009). S-acyl modifications mainly affect membrane attachment and trafficking of proteins. They are required for the dynamic association of proteins with membrane subdomains (Sorek et al., 2007, 2010) and for the cycling between different cellular membranes (Rocks et al., 2005). In addition, S-acylation can influence the stability of proteins (Valdez-Taubas and Pelham, 2005; Abrami et al., 2006), modulates the functions of proteins (Gubitosi-Klug et al., 2005), or mediates the interaction between different proteins (Charrin et al., 2002; Flannery et al., 2010).S-acyl modification of proteins is brought about by palmitoyltransferases, correctly referred to as protein S-acyltransferases (PATs; Mitchell et al., 2006). Originally, these enzymes were identified in yeast (Saccharomyces cerevisiae; Lobo et al., 2002; Roth et al., 2002). Sequence analyses of the yeast PAT enzymes revealed that they share a common structure mainly composed of four predicted transmembrane domains (TMDs) and a stretch of Asp-His-His-Cys (DHHC) within a Cys-rich domain (CRD; Pfam accession, PF01529; InterPro entry, IPR001594). Mutational analyses revealed that the Cys residue of the DHHC stretch is necessary for autoacylation and for the modification of target proteins. Therefore, this domain appears to represent the active site of these enzymes (Lobo et al., 2002; Roth et al., 2002). Detailed analyses of the lipid modifications of Rho-of-plants6 (ROP6; Sorek et al., 2007) and of the Calcineurin-B like (CBL) proteins 1 and 2 (Batistič et al., 2008, 2012) revealed that plant PATs either transfer palmitate or stearate to these substrates.In plants, so far only one PAT was characterized in detail. A screen of Arabidopsis (Arabidopsis thaliana) mutants resulted in the identification of TIP GROWTH DEFECTIVE1 (TIP1), which is affected in root hair formation and growth (Schiefelbein et al., 1993; Ryan et al., 1998). Mapping of the mutant allele revealed that TIP1 encodes a DHHC-CRD-containing protein, and it was shown that this protein is indeed S-acylated (Hemsley et al., 2005, 2008). Moreover, TIP1 contains N-terminal ankyrin repeats, related to the yeast PAT Akr1, and is able to complement the PAT function in the akr1Δ yeast strain, corroborating that TIP1 has a PAT enzyme function (Hemsley et al., 2005). Although our knowledge about the plant S-acylome and S-acylation machinery is still limited (Hemsley, 2009), previous studies on the S-acylation of plant proteins revealed that these modifications could occur at different cellular membranes like the endoplasmic reticulum (ER; Adjobo-Hermans et al., 2006; Batistič et al., 2008), the Golgi (Zeng et al., 2007), or directly at the plasma membrane (PM; Sorek et al., 2007). Furthermore, recent studies on the lipid modification of the tonoplast-associated CBL2 protein from Arabidopsis implicated that S-acylation can also occur at the vacuolar membrane (Batistič et al., 2012). These findings suggested that the PAT enzymes that modify these targets reside at different membranes within the plant cell.In this report, the complement of the Arabidopsis PAT gene loci was examined to perform a comprehensive analysis of this gene and protein family. These analyses revealed that at least 24 PAT gene loci are encoded in the Arabidopsis genome. All Arabidopsis PATs are expressed. Expression occurs mainly at steady levels throughout development and in most tissues examined. However, some PATs are preferentially expressed at certain developmental stages or cell types especially during flowering and in pollen, respectively. Phylogenetic analysis revealed that the proteins are highly divergent. Nevertheless, most PAT proteins harbor a similar structural topology and preserved sequence motifs that are representative of this class of enzymes. Functional analyses on several proteins by yeast complementation analyses indicate that Arabidopsis PATs represent active enzymes, even if their sequence deviates from the regular standard sequence composition previously reported for functional PATs. Moreover, the protein localizations of all 24 Arabidopsis PATs were examined. This revealed a complex targeting pattern of PATs showing that S-acylation can occur at different cellular membranes within the cell. Remarkably, the plant PATs analyzed are mainly targeted to the PM, indicating that the plant S-acylation machinery has a different organization and function compared with the mammalian and yeast PAT machinery.     

Genomic and morphological evidence converge to resolve the enigma of Strepsiptera

The phylogeny of insects, one of the most spectacular radiations of life on earth, has received considerable attention. However, the evolutionary roots of one intriguing group of insects, the twisted-wing parasites (Strepsiptera), remain unclear despite centuries of study and debate. Strepsiptera exhibit exceptional larval developmental features, consistent with a predicted step from direct (hemimetabolous) larval development to complete metamorphosis that could have set the stage for the spectacular radiation of metamorphic (holometabolous) insects. Here we report the sequencing of a Strepsiptera genome and show that the analysis of sequence-based genomic data (comprising more than 18 million nucleotides from nearly 4,500 genes obtained from a total of 13 insect genomes), along with genomic metacharacters, clarifies the phylogenetic origin of Strepsiptera and sheds light on the evolution of holometabolous insect development. Our results provide overwhelming support for Strepsiptera as the closest living relatives of beetles (Coleoptera). They demonstrate that the larval developmental features of Strepsiptera, reminiscent of those of hemimetabolous insects, are the result of convergence. Our analyses solve the long-standing enigma of the evolutionary roots of Strepsiptera and reveal that the holometabolous mode of insect development is more malleable than previously thought.     

Heterocyclic methylsulfone hydroxamic acid LpxC inhibitors as Gram-negative antibacterial agents

The synthesis and antibacterial activity of heterocyclic methylsulfone hydroxamates is presented. Compounds in this series are potent inhibitors of the LpxC enzyme, a key enzyme involved in the production of lipopolysaccharide (LPS) found in the outer membrane of Gram-negative bacteria. SAR evaluation of compounds in this series revealed analogs with potent antibacterial activity against challenging Gram-negative species such as Pseudomonas aeruginosa and Klebsiella pneumoniae.     

Multiple nuclear genes and retroposons support vicariance and dispersal of the palaeognaths,and an Early Cretaceous origin of modern birds

The origin and timing of the diversification of modern birds remains controversial, primarily because phylogenetic relationships are incompletely resolved and uncertainty persists in molecular estimates of lineage ages. Here, we present a species tree for the major palaeognath lineages using 27 nuclear genes and 27 archaic retroposon insertions. We show that rheas are sister to the kiwis, emu and cassowaries, and confirm ratite paraphyly because tinamous are sister to moas. Divergence dating using 10 genes with broader taxon sampling, including emu, cassowary, ostrich, five kiwis, two rheas, three tinamous, three extinct moas and 15 neognath lineages, suggests that three vicariant events and possibly two dispersals are required to explain their historical biogeography. The age of crown group birds was estimated at 131 Ma (95% highest posterior density 122–138 Ma), similar to previous molecular estimates. Problems associated with gene tree discordance and incomplete lineage sorting in birds will require much larger gene sets to increase species tree accuracy and improve error in divergence times. The relatively rapid branching within neoaves pre-dates the extinction of dinosaurs, suggesting that the genesis of the radiation within this diverse clade of birds was not in response to the Cretaceous–Paleogene extinction event.     

Robust and fast whole brain mapping of the RF transmit field B1 at 7T

In-vivo whole brain mapping of the radio frequency transmit field B(1) (+) is a key aspect of recent method developments in ultra high field MRI. We present an optimized method for fast and robust in-vivo whole-brain B(1) (+) mapping at 7T. The method is based on the acquisition of stimulated and spin echo 3D EPI images and was originally developed at 3T. We further optimized the method for use at 7T. Our optimization significantly improved the robustness of the method against large B(1) (+) deviations and off-resonance effects present at 7T. The mean accuracy and precision of the optimized method across the brain was high with a bias less than 2.6 percent unit (p.u.) and random error less than 0.7 p.u. respectively.     

Confocal laser scanning microscopy, a new in vivo diagnostic tool for schistosomiasis

Background

The gold standard for the diagnosis of schistosomiasis is the detection of the parasite''s characteristic eggs in urine, stool, or rectal and bladder biopsy specimens. Direct detection of eggs is difficult and not always possible in patients with low egg-shedding rates. Confocal laser scanning microscopy (CLSM) permits non-invasive cell imaging in vivo and is an established way of obtaining high-resolution images and 3-dimensional reconstructions. Recently, CLSM was shown to be a suitable method to visualize Schistosoma mansoni eggs within the mucosa of dissected mouse gut. In this case, we evaluated the suitability of CLSM to detect eggs of Schistosoma haematobium in a patient with urinary schistosomiasis and low egg-shedding rates.

Methodology/Principal Findings

The confocal laser scanning microscope used in this study was based on a scanning laser system for imaging the retina of a living eye, the Heidelberg Retina Tomograph II, in combination with a lens system (image modality). Standard light cystoscopy was performed using a rigid cystoscope under general anaesthesia. The CLSM endoscope was then passed through the working channel of the rigid cystoscope. The mucosal tissue of the bladder was scanned using CLSM. Schistoma haematobium eggs appeared as bright structures, with the characteristic egg shape and typical terminal spine.

Conclusion/Significance

We were able to detect schistosomal eggs in the urothelium of a patient with urinary schistosomiasis. Thus, CLSM may be a suitable tool for the diagnosis of schistosomiasis in humans, especially in cases where standard diagnostic tools are not suitable.