New systematic assignments in Gonyleptoidea (Arachnida, Opiliones, Laniatores)

As part of an ongoing revision of the family Gonyleptidac, we have identified many species that are synonyms of previously described species or misplaced in this family. This article summarizes these findings, adding previously unavailable information or correcting imprecise observations to justify the presented taxonomic changes. The following new familial or subfamilial assignments are proposed: Nemastygnus Roewer, 1929 and Taulisa Roewer, 1956 are transferred to Agoristenidae, Agoristeninae; Napostygnus Roewer, 1929 to Cranaidae; Ceropachylinus peruvianus Roewer, 1956 and Pirunipygus Roewer, 1936 are transferred to Gonyleptidae, Ampycinae; Gyndesops Roewer, 1943, Haversia Roewer, 1913 and Oxapampeus Roewer, 1963 are transferred to Gonyleptidae, Pachylinae. The following generic synonymies are proposed for the family Gonyleptidae: Acanthogonyleptes Mello-Leitao, 1922 = Centroleptes Roewer, 1943; Acrographinotus Roewer, 1929 = Unduavius Roewer, 1929; Gonyleptes Kirby, 1819 = Collonychium Bertkau, 1880; Mischonyx Bertkau, 1880 = Eugonyleptes Roewer, 1913 and Gonazula Roewer, 1930; Parampheres Roewer, 1913 = Metapachyloides Roewer, 1917; Pseudopucrolia Roewer, 19 12 = Meteusarcus Roewer, 1913; Haversia Roewer, 19 13 = Hoggellula Roewer, 1930. The following specific synonymies are proposed for the family Gonyleptidae: Acanthogonyleptes singularis (Mello-Leitao, 1935) = Centroleptes flavus Roewer, 1943, syn. n.; Geraeocormobius sylvarum Holmberg, 1887 = Discocyrtus serrifemur Roewer, 1943, syn. n.; Gonyleptellus bimaculatus (Sorensen, 1884) = Gonyleptes cancellatus Roewer, 1917, syn. n.; Gonyleptes atrus Mello-Leitao, 1923 = Weyhia brieni Giltay, 1928, syn. n.; Gonyleptes fragilis Mello-Leitao, 1923 = Gonyleptes banana Kury, 2003, syn. n.; Gonyleptes horridus Kirby, 1819 = Collonychium bicuspidatum Bertkau, 1880, syn. n., Gonyleptes borgmeyeri Mello-Leitao, 1932, syn. n., Gonyleptes curvicornis Mello-Leitao, 1932, syn. n., Metagonyleptes hamatus Roewer, 1913, syn. n. and Paragonyleptes simoni Roewer, 1930, syn. n.; Gonyleptes pustulatus Sorensen, 1884 = Gonyleptes guttatus Roewer, 1917, syn. n.; Haversia defensa (Butler, 1876) = Sadocus vallentini Hogg, 1913, syn. n.; Liogonyleptoides minensis (Piza, 1946) = Currala bahiensis Soares, 1972, syn. n.; Megapachylus grandis Roewer, 1913 = Metapachyloides almeidai Soares & Soares, 1946, syn. n.; Mischonyx cuspidatus (Roewer, 1913) = Gonazula gibbosa Roewer, 1930 syn. n.; Mischonyx scaber (Kirby, 1819) = Xundarava holacantha Mello-Leitao, 1927, syn. n.; Parampheres tibialis Roewer, 1917 = Metapachyloides rugosus Roewer, 1917, syn. n.; Parapachyloides uncinatus (Sorensen, 1879) = Goyazella armata Mello-Leitao, 1931, syn. n.; Pseudopucrolia mutica (Perry, 1833) = Meteusarcus armatus Roewer, 1913, syn. n. The following new combinations are proposed: Acrographinotus ornatus (Roewer, 1929), comb. n. (ex Unduavius); Gonyleptellus bimaculatus (Sorensen, 1884), comb. n. (ex Gonyleptes); Gonyleptes perlatus (Mello-Leitao, 1935), comb. n. (ex Moojenia); Mischonyx scaber (Kirby, 1819), comb. n. (ex Gonyleptes); and Neopachyloides peruvianus (Roewer, 1956), comb. n. (ex Ceropachylus). The following species of Gonyleptidae, Gonyleptinae are revalidated: Gonyleptes atrus Mello-Leitao, 1923 and Gonyleptes curvicornis (Roewer, 1913).

Evidence that a single bioluminescent system is shared by all known bioluminescent fungal lineages

Since the early 20th century, many researchers have attempted to determine how fungi are able to emit light. The first successful experiment was obtained using the classical luciferin-luciferase test that consists of mixing under controlled conditions hot (substrate/luciferin) and cold (enzyme/luciferase) water extracts prepared from bioluminescent fungi. Failures by other researchers to reproduce those experiments using different species of fungi lead to the hypothesis of a non-enzymatic luminescent pathway. Only recently, the involvement of a luciferase in this system was proven, thus confirming its enzymatic nature. Of the 100 000 described species in Kingdom Fungi, only 71 species are known to be luminescent and they are distributed unevenly amongst four distantly related lineages. The question we address is whether the mechanism of bioluminescence is the same in all four evolutionary lineages suggesting a single origin of luminescence in the Fungi, or whether each lineage has a unique mechanism for light emission implying independent origins. We prepared hot and cold extracts of numerous species representing the four bioluminescent fungal lineages and performed cross-reactions (luciferin x luciferase) in all possible combinations using closely related non-luminescent species as controls. All cross-reactions with extracts from luminescent species yielded positive results, independent of lineage, whereas no light was emitted in cross-reactions with extracts from non-luminescent species. These results support the hypothesis that all four lineages of luminescent fungi share the same type of luciferin and luciferase, that there is a single luminescent mechanism in the Fungi, and that fungal luciferin is not a ubiquitous molecule in fungal metabolism.

Analysis of three Xanthomonas axonopodis pv. citri effector proteins in pathogenicity and their interactions with host plant proteins

Xanthomonas axonopodis pv. citri, the bacterium responsible for citrus canker, uses effector proteins secreted by a type III protein secretion system to colonize its hosts. Among the putative effector proteins identified for this bacterium, we focused on the analysis of the roles of AvrXacE1, AvrXacE2 and Xac3090 in pathogenicity and their interactions with host plant proteins. Bacterial deletion mutants in avrXacE1, avrXacE2 and xac3090 were constructed and evaluated in pathogenicity assays. The avrXacE1 and avrXacE2 mutants presented lesions with larger necrotic areas relative to the wild-type strain when infiltrated in citrus leaves. Yeast two-hybrid studies were used to identify several plant proteins likely to interact with AvrXacE1, AvrXacE2 and Xac3090. We also assessed the localization of these effector proteins fused to green fluorescent protein in the plant cell, and observed that they co-localized to the subcellular spaces in which the plant proteins with which they interacted were predicted to be confined. Our results suggest that, although AvrXacE1 localizes to the plant cell nucleus, where it interacts with transcription factors and DNA-binding proteins, AvrXacE2 appears to be involved in lesion-stimulating disease 1-mediated cell death, and Xac3090 is directed to the chloroplast where its function remains to be clarified.

The 3D structure and function of digestive cathepsin L-like proteinases of Tenebrio molitor larval midgut

Cathepsin L-like proteinases (CAL) are major digestive proteinases in the beetle Tenebrio molitor. Procathepsin Ls 2 (pCAL2) and 3 (pCAL3) were expressed as recombinant proteins in Escherichia coil, purified and activated under acidic conditions. Immunoblot analyses of different T. molitor larval tissues demonstrated that a polyclonal antibody to pCAL3 recognized pCAL3 and cathepsin L 3 (CAD) only in the anterior two-thirds of midgut tissue and midgut luminal contents of T. molitor larvae. Furthermore, immunocytolocalization data indicated that pCAL3 occurs in secretory vesicles and microvilli in anterior midgut Therefore CAL3, like cathepsin L 2 (CAL2), is a digestive enzyme secreted by T. molitor anterior midgut CAD hydrolyses Z-FR-MCA and Z-RR-MCA (typical cathepsin substrates), whereas CAL2 hydrolyses only Z-FR-MCA. Active site mutants (pCAL2C25S and pCAL3C265) were constructed by replacing the catalytic cysteine with serine to prevent autocatalytic processing. Recombinant pCAL2 and pCAL3 mutants (pCAL2C25S and pCAL3C26S) were prepared, crystallized and their 3D structures determined at 1.85 and 2.1 angstrom, respectively. While the overall structure of these enzymes is similar to other members of the papain superfamily, structural differences in the S2 subsite explain their substrate specificities. The data also supported models for CAL trafficking to lysosomes and to secretory vesicles to be discharged into midgut contents. (C) 2012 Elsevier Ltd. All rights reserved.

Acute cocaine treatment increases thimet oligopeptidase in the striatum of rat brain

Many studies indicate that thimet oligopeptidase (EC3.4.24.15; TOP) can be implicated in the metabolism of bioactive peptides, including dynorphin 1-8, alpha-neoendorphin, beta-neoendorphin and GnRH. Furthermore, the higher levels of this peptidase are found in neuroendocrine tissue and testis. In the present study, we have evaluated the effect of acute cocaine administration in male rats on TOP specific activity and mRNA levels in prosencephalic brain areas related with the reward circuitry; ventral striatum, hippocampus, and frontal cortex. No significant differences on TOP specific activity were detected in the hippocampus and frontal cortex of cocaine treated animals compared to control vehicle group. However, a significant increase in activity was observed in the ventral striatum of cocaine treated-rats. The increase occurred in both, TOP specific activity and TOP relative mRNA amount determined by real time RT-PCR. As TOP can be implicated in the processing of many neuropeptides, and previous studies have shown that cocaine also alters the gene expression of proenkephalin and prodynorphin in the striatum, the present findings suggest that TOP changes in the brain could play important role in the balance of neuropeptide level correlated with cocaine effects. (C) 2012 Elsevier Inc. All rights reserved.

Dynamic complex formation between HD-GYP, GGDEF and PilZ domain proteins regulates motility in Xanthomonas campestris

RpfG is a member of a class of wide spread bacterial two-component regulators with an HD-GYP cyclic di-GMP phosphodiesterase domain. In the plant pathogen Xanthomonas campestris, RpfG together with the sensor kinase RpfC regulates multiple factors as a response to the cell-to-cell Diffusible Signalling Factor (DSF). A dynamic physical interaction of RpfG with two diguanylate cyclase (GGDEF) domain proteins controls motility. Here we show that, contrary to expectation, regulation of motility by the GGDEF domain proteins does not depend upon their cyclic di-GMP synthetic activity. Furthermore we show that the complex of RpfG and GGDEF domain proteins recruits a specific PilZ domain adaptor protein, and this complex then interacts with the pilus motor proteins PilU and PiIT. The results support a model in which DSF signalling influences motility through the highly regulated dynamic interaction of proteins that affect pilus action. A specific motif that we identify to be required for HD-GYP domain interaction is conserved in a number of GGDEF domain proteins, suggesting that regulation via interdomain interactions is of broad relevance.

Pimavanserin, a selective serotonin (5-HT)2A-inverse agonist, enhances the efficacy and safety of risperidone, 2 mg/day, but does not enhance efficacy of haloperidol, 2 mg/day: Comparison with reference dose risperidone, 6 mg/day

Most atypical antipsychotic drugs (APDs), e. g. risperidone (RIS), produce more extensive blockade of brain serotonin (5-HT)(2A) than dopamine (DA) D-2 receptors. This distinguishes them from typical APDs, e.g. haloperidol (HAL). Our objective was to test the hypothesis that augmentation of low doses of RIS or HAL (2 mg/day) with pimavanserin (PIM), a selective 5-HT2A inverse agonist, to enhance 5-HT2A receptor blockade, can achieve efficacy comparable to RIS, 6 mg/day, but with lesser side effects. In a multi-center, randomized, double-blind, 6 week trial, 423 patients with chronic schizophrenia experiencing a recent exacerbation of psychotic symptoms were randomized to RIS2mg + placebo (RIS2PBO), RIS2mg + PIM20mg (RIS2PIM), RIS6mg + PBO (RIS6PBO), HAL2mg + PBO (HAL2PBO), or HAL2mg + PIM20mg (HAL2PIM). Improvement in psychopathology was measured by the PANSS and CGI-S. The reduction in PANSS Total Score with RIS2PIM at endpoint was significantly greater than RIS2PBO: -23.0 vs. -16.3 (p = 0.007), and not significantly different from the RIS6PBO group: -23.2 points. The percentage of patients with >= 20% improvement at day 15 in the RIS2PIM group was 62.3%, significantly greater than the RIS6PBO (42.1%; p = 0.01) and the RIS2PBO groups (37.7%; p = 0.002). Weight gain and hyperprolactinemia were greater in the RIS6PBO group than the RIS2PIM group but there was no difference in extrapyramidal side effects (EPS). HAL2PBO and HAL2PIM were not significantly different from each other in efficacy but HAL2PIM had less EPS at end point. Both HAL groups and RIS6PBO showed equal improvement in psychopathology at endpoint, indicating HAL 2 mg/day is effective to treat an acute exacerbation in chronic schizophrenia patients. In conclusion, a sub-effective RIS dose combined with PIM to enhance 5-HT2A receptor blockade provided faster onset of action, and at endpoint, equal efficacy and better safety, compared to standard dose RIS. These results support the conclusion that 5-HT2A receptor blockade is a key component of the action of some atypical APDs and can reduce EPS due to a typical APD. (C) 2012 Elsevier B.V. All rights reserved.

Calcium Binding to Leptospira Outer Membrane Antigen LipL32 Is Not Necessary for Its Interaction with Plasma Fibronectin, Collagen Type IV, and Plasminogen

LipL32 is the most abundant outer membrane protein from pathogenic Leptospira and has been shown to bind extracellular matrix (ECM) proteins as well as Ca2+. Recent crystal structures have been obtained for the protein in the apo-and Ca2+-bound forms. In this work, we produced three LipL32 mutants (D163-168A, Q67A, and S247A) and evaluated their ability to interact with Ca2+ and with ECM glycoproteins and human plasminogen. The D163-168A mutant modifies aspartate residues involved in Ca2+ binding, whereas the other two modify residues in a cavity on the other side of the protein structure. Loss of calcium binding in the D163-D168A mutant was confirmed using intrinsic tryptophan fluorescence, circular dichroism, and thermal denaturation whereas the Q67A and S247A mutants presented the same Ca2+ affinity as the wild-type protein. We then evaluated if Ca2+ binding to LipL32 would be crucial for its interaction with collagen type IV and plasma proteins fibronectin and plasminogen. Surprisingly, the wild-type protein and all three mutants, including the D163-168A variant, bound to these ECM proteins with very similar affinities, both in the presence and absence of Ca2+ ions. In conclusion, calcium binding to LipL32 may be important to stabilize the protein, but is not necessary to mediate interaction with host extracellular matrix proteins.

First assessment of the endoparasitic nematode fauna of four psammophilous species of Tropiduridae (Squamata: Iguania) endemic to north-eastern Brazil

Tropiduridae (Squamata: Iguania) is a lizard taxon widely distributed in the neotropics. Among its representatives, some species are classified as generalists regarding habitat usage. Others exhibit a very restricted and probably relict distribution, and are strongly associated with predominantly sandy and dry habitats. Within this rather ecologically similar than phylogenetically closely related group we examined specimens of Eurolophosaurus amathites, E. divaricatus, Tropidurus hygomi, T. psammonastes for endoparasites. In all four species examined we recorded parasitic nematodes (Nemathelminthes: Nematoda). At least three nematode species were recovered: Parapharyngodon sp., Physaloptera lutzi and Strongyluris oscari, with Ph. lutzi being the most abundant parasite encountered in all lizard species examined. In spite of the hosts' habitat specialization, these parasites are also found frequently in non-psammophilous tropidurid species as well as in other squamates. Individual species richness per lizard was low, with usually just one species parasitizing at a time. These are the first parasites registered for these tropidurids and constitute a total of six new host records.

Extracytoplasmic function (ECF) sigma factor σF is involved in Caulobacter crescentus response to heavy metal stress

Background The α-proteobacterium Caulobacter crescentus inhabits low-nutrient environments and can tolerate certain levels of heavy metals in these sites. It has been reported that C. crescentus responds to exposure to various heavy metals by altering the expression of a large number of genes. Results In this work, we show that the ECF sigma factor σF is one of the regulatory proteins involved in the control of the transcriptional response to chromium and cadmium. Microarray experiments indicate that σF controls eight genes during chromium stress, most of which were previously described as induced by heavy metals. Surprisingly, σF itself is not strongly auto-regulated under metal stress conditions. Interestingly, σF-dependent genes are not induced in the presence of agents that generate reactive oxygen species. Promoter analyses revealed that a conserved σF-dependent sequence is located upstream of all genes of the σF regulon. In addition, we show that the second gene in the sigF operon acts as a negative regulator of σF function, and the encoded protein has been named NrsF (Negative regulator of sigma F). Substitution of two conserved cysteine residues (C131 and C181) in NrsF affects its ability to maintain the expression of σF-dependent genes at basal levels. Furthermore, we show that σF is released into the cytoplasm during chromium stress and in cells carrying point mutations in both conserved cysteines of the protein NrsF. Conclusion A possible mechanism for induction of the σF-dependent genes by chromium and cadmium is the inactivation of the putative anti-sigma factor NrsF, leading to the release of σF to bind RNA polymerase core and drive transcription of its regulon.

Structure of the PilZ-FimXEAL-c-di-GMP complex responsible for the regulation of bacterial type IV pilus biogenesis

Signal transduction pathways mediated by cyclic-bis(3'→5')-dimeric GMP (c-di-GMP) control many important and complex behaviors in bacteria. C-di-GMP is synthesized through the action of GGDEF domains that possess diguanylate cyclase activity and is degraded by EAL or HD-GYP domains with phosphodiesterase activity. There is mounting evidence that some important c-di-GMP-mediated pathways require protein-protein interactions between members of the GGDEF, EAL, HD-GYP and PilZ protein domain families. For example, interactions have been observed between PilZ and the EAL domain from FimX of Xanthomonas citri (Xac). FimX and PilZ are involved in the regulation of type IV pilus biogenesis via interactions of the latter with the hexameric PilB ATPase associated with the bacterial inner membrane. Here, we present the crystal structure of the ternary complex made up of PilZ, the FimX EAL domain (FimXEAL) and c-di-GMP. PilZ interacts principally with the lobe region and the N-terminal linker helix of the FimXEAL. These interactions involve a hydrophobic surface made up of amino acids conserved in a non-canonical family of PilZ domains that lack intrinsic c-di-GMP binding ability and strand complementation that joins β-sheets from both proteins. Interestingly, the c-di-GMP binds to isolated FimXEAL and to the PilZ-FimXEAL complex in a novel conformation encountered in c-di-GMP-protein complexes in which one of the two glycosidic bonds is in a rare syn conformation while the other adopts the more common anti conformation. The structure points to a means by which c-di-GMP and PilZ binding could be coupled to FimX and PilB conformational states