Molecular architecture of the recombinant human MCM2-7 helicase in complex with nucleotides and DNA

DNA replication is a key biological process that involves different protein complexes whose assembly is rigorously regulated in a successive order. One of these complexes is a replicative hexameric helicase, the MCM complex, which is essential for the initiation and elongation phases of replication. After the assembly of a double heterohexameric MCM2-7 complex at replication origins in G1, the 2 heterohexamers separate from each other and associate with Cdc45 and GINS proteins in a CMG complex that is capable of unwinding dsDNA during S phase. Here, we have reconstituted and characterized the purified human MCM2-7 (hMCM2-7) hexameric complex by co-expression of its 6 different subunits in insect cells. The conformational variability of the complex has been analyzed by single particle electron microscopy in the presence of different nucleotide analogs and DNA. The interaction with nucleotide stabilizes the complex while DNA introduces conformational changes in the hexamer inducing a cylindrical shape. Our studies suggest that the assembly of GINS and Cdc45 to the hMCM2-7 hexamer would favor conformational changes on the hexamer bound to ssDNA shifting the cylindrical shape of the complex into a right-handed spiral conformation as observed in the CMG complex bound to DNA.     

Cache Domains That are Homologous to,but Different from PAS Domains Comprise the Largest Superfamily of Extracellular Sensors in Prokaryotes

Cellular receptors usually contain a designated sensory domain that recognizes the signal. Per/Arnt/Sim (PAS) domains are ubiquitous sensors in thousands of species ranging from bacteria to humans. Although PAS domains were described as intracellular sensors, recent structural studies revealed PAS-like domains in extracytoplasmic regions in several transmembrane receptors. However, these structurally defined extracellular PAS-like domains do not match sequence-derived PAS domain models, and thus their distribution across the genomic landscape remains largely unknown. Here we show that structurally defined extracellular PAS-like domains belong to the Cache superfamily, which is homologous to, but distinct from the PAS superfamily. Our newly built computational models enabled identification of Cache domains in tens of thousands of signal transduction proteins including those from important pathogens and model organisms. Furthermore, we show that Cache domains comprise the dominant mode of extracellular sensing in prokaryotes.     

Mechanisms governing radial heat fluxes in tokamak plasma

A method for analyzing the characteristics of turbulence responsible for radial heat transport is proposed. The method is based on the previously proposed hypotheses (to a great extent, confirmed experimentally) concerning the consistency of normalized pressure profiles in tokamak plasmas and the mechanism of internal transport barrier formation. Using the proposed approach, it is shown that, under an external action on the plasma, when the plasma heat flux onto the wall grows, the spectrum of turbulent modes broadens due to the excitation of modes with lower poloidal numbers m. Thus, in contrast to the conventional diffusion approach, the transport coefficient depends on the flux intensity. A mechanism of formation of internal transport barriers is proposed.     

Soil and entomopathogenic fungi with potential for biodegradation of insecticides: degradation of flubendiamide in vivo by fungi and in vitro by laccase

Purpose

Flubendiamide is a highly toxic and persistent insecticide that causes loss of insect muscle functions leading to paralysis and death. The objective was to screen for filamentous fungi in soils where insecticides had been applied, to isolate entomopathogenic fungi from insect larva (Anticarsia gemmatalis) that infest soybean crops, and to use these in biodegradation of insecticides.

Method

Filamentous fungi were isolated from soils, and growth inhibition was evaluated on solid medium containing commercial insecticides, Belt® (flubendiamide) and Actara® (thiamethoxam). A total of 133 fungi were isolated from soil and 80 entomopathogenic fungi from insect larva. Based on growth inhibition tests, ten soil fungi, 2 entomopathogenic fungi, and Botryosphaeria rhodina MAMB-05 (reference standard) were selected for growth on commercial insecticides in solid media. Fungi were grown in submerged fermentation on media containing commercial insecticides and assayed for laccase activity.

Result

Isolates JUSOLCL039 (soil), JUANT070 (insect), and MAMB-05 performed best, and were respectively inhibited by 48.41%, 75.97%, and 79.23% when cultivated on 35 g/L Actara®, and 0.0, 5.42%, and 43.39% on 39.04 g/L Belt®. JUSOLCL039 and JUANT070 were molecularly identified as Trichoderma koningiopsis and Neurospora sp., respectively. The three fungal isolates produced laccase constitutively, albeit at low activities. Fungal growth on pure flubendiamide and thiamethoxam resulted in only thiamethoxam inducing high laccase titers (10.16 U/mL) by JUANT070. Neurospora sp. and B. rhodina degraded flubendiamide by 27.4% and 9.5% in vivo, while a crude laccase from B. rhodina degraded flubendiamide by 20.2% in vitro.

Conclusion

This is the first report of fungi capable of degrading flubendiamide, which have applications in bioremediation.

     

Combination of sepsis biomarkers may indicate an invasive fungal infection in haematological patients

Background: Invasive fungal infections are a major threat to a large cohort of immunocompromised patients, including patients with chemotherapy-associated neutropenia. Early differential diagnosis with bacterial infections is often complicated, which leads to a delay in empirical antifungal therapy and increases risk for adverse outcome. Accessibility and performance of specific fungal antigen and PCR-tests are still limited, while sepsis biomarkers are more broadly used in most settings currently.

Methods: Haematological patients hospitalized to receive chemotherapy with proven or probable invasive fungal infection or microbiologically proven bacterial bloodstream infection were included in the study. C-reactive protein was assessed daily during the profound neutropenia period, while procalcitonin or presepsin were measured during the first 48?hours after the onset of febrile episode.

Results: There were totally 64 patients included in the study, 53 with bacterial bloodstream infections and 11 with invasive fungal infections. Combination of CRP >120 with PCT <1.25 or presepsin <170 was shown to be a possible combined biomarker for invasive fungal infections in immunocompromised patients, with areas under the ROC-curves: 0.962 (95% CI 0.868 to 0.995) for PCT-based combination and 0.907 (95% CI 0.692 to 0.990) for presepsin-based combination.     

Shining light on the alphaproteobacterial general stress response

The general stress response (GSR) allows many bacterial species to react to myriad different stressors. In Alphaproteobacteria, this signaling pathway proceeds through the partner‐switching PhyR‐EcfG sigma‐factor mechanism and is involved in multiple life processes, including virulence in Brucella abortus. To date, details of the alphaproteobacterial GSR signaling pathway have been determined using genetic and biochemical work on a diverse set of species distributed throughout the clade. Fiebig and co‐workers establish Erythrobacter litoralis DSM 8509 as a genetically tractable lab strain and use it to both directly and indirectly delineate photoresponsive GSR pathways mediated by multiple HWE/HisKA_2 histidine kinases. The existence of a new phototrophic lab strain allows researchers to compare the GSR across different Alphaproteobacteria, as well as study the interplay between the GSR and phototrophy. Additionally, the discovery of new HWE/HisKA_2 kinases regulating the GSR poses new questions about how different stimuli feed into this widespread stress pathway.     

Morphology and Phylogeny of Calyptospora paranaidji n. sp. (Eimeriorina: Calyptosporidae), an Apicomplexan Parasite of the Hepatic Tissue of Cichla piquiti Kullander & Ferreira, 2006, From a Reservoir in the Brazilian Amazon Region

The coccidians of the family Calyptosporidae are parasites of the tissue and organs of fish and aquatic invertebrates, in particular in the tropical region. In contrast with other apicomplexans of the suborder Eimeriorina, the diversity and ecology of the species of the genus Calyptospora have been poorly investigated, resulting in a lacuna that restricts the understanding of the distribution and prevalence of this group of eukaryote microparasites in the Amazon region. In the present study, the integrated comparative analysis of morphological characteristics, histological and structural traits, and the sequences of a fragment of the 18S rRNA gene, provides support for the identification of a new species of Calyptospora, found parasitizing the hepatic tissue of the piscivorous blue peacock bass, Cichla piquiti, captured in the reservoir of the Estreito hydroelectric dam on the middle Tocantins River in northern Brazil. This new species was named Calyptospora paranaidji n. sp.     

Salicylic acid influences the protease activity and posttranslation modifications of the secreted peptides in the moss Physcomitrella patens

Plant secretome comprises dozens of secreted proteins. However, little is known about the composition of the whole secreted peptide pools and the proteases responsible for the generation of the peptide pools. The majority of studies focus on target detection and characterization of specific plant peptide hormones. In this study, we performed a comprehensive analysis of the whole extracellular peptidome, using moss Physcomitrella patens as a model. Hundreds of modified and unmodified endogenous peptides that originated from functional and nonfunctional protein precursors were identified. The plant proteases responsible for shaping the pool of endogenous peptides were predicted. Salicylic acid (SA) influenced peptide production in the secretome. The proteasome activity was altered upon SA treatment, thereby influencing the composition of the peptide pools. These results shed more light on the role of proteases and posttranslational modification in the “active management” of the extracellular peptide pool in response to stress conditions. It also identifies a list of potential peptide hormones in the moss secretome for further analysis.     

Riparian zone as a main determinant of the structure of lizard assemblages in upland Amazonian forests

The use of lizards as model organisms in ecological studies is based on their success in occupying a great diversity of habitats, and some species are closely tied to the environment, which is disadvantaged by the legislation of several countries concerning land use. Our aim was to relate lizard species distribution patterns in rainforest environments to variation in environmental gradients, and provide ecologically based metrics for establishing buffer zones around streams. Lizards were sampled three times in 41 standardised transects near Manaus, Brazil, only in dry season, with Time Limited Visual Search associated with raking through leaf litter. We recorded 20 species from 10 families and used non‐metric multidimensional scaling to reduce the dimensionality of quantitative and qualitative compositions of species. Multiple linear regression models indicated that the environmental gradients distance to nearest stream, extent of canopy openness, vegetation density and slope did not significantly influence assemblage species distribution, with an indication of effect of litter depth. By means of piecewise linear regression, the use of riparian zone was estimated at ~190 m from quantitative species composition and ~211 m from qualitative species composition. Five species occurred only in the riparian zone. Our results suggest that conservation of the entire riparian lizard assemblage in Amazonian rainforest is likely to require protection of at least a 211 m buffer on either side of streams.