Potassium channel blocker crafted by α-hairpinin scaffold engineering

The α-Hairpinins are a family of plant defense peptides with a common fold presenting two short α-helices stabilized by two invariant S–S-bridges. We have shown previously that substitution of just two amino acid residues in a wheat α-hairpinin Tk-AMP-X2 leads to Tk-hefu-2 that features specific affinity to voltage-gated potassium channels K_V1.3. Here, we utilize a combined molecular modeling approach based on molecular dynamics simulations and protein surface topography technique to improve the affinity of Tk-hefu-2 to K_V1.3 while preserving its specificity. An important advance of this work compared with our previous studies is transition from the analysis of various physicochemical properties of an isolated toxin molecule to its consideration in complex with its target, a membrane-bound ion channel. As a result, a panel of computationally designed Tk-hefu-2 derivatives was synthesized and tested against K_V1.3. The most active mutant Tk-hefu-10 showed a half-maximal inhibitory concentration of ～150 nM being >10 times more active than Tk-hefu-2 and >200 times more active than the original Tk-hefu. We conclude that α-hairpinins provide an attractive disulfide-stabilized scaffold for the rational design of ion channel inhibitors. Furthermore, the success rate can be considerably increased by the proposed “target-based” iterative strategy of molecular design.     

Pathological and immunological evaluation of different regimens of praziquantel treatment in a mouse model of Schistosoma mansoni infection

BackgroundOne of the considerable challenges of schistosomiasis chemotherapy is the inefficacy of praziquantel (PZQ) at the initial phase of the infection. Immature schistosomes are not susceptible to PZQ at the curative dose. Here, we investigated the efficacy of different PZQ regimens administered during the initial stage of Schistosoma mansoni infection in mice.Methodology/Principal findingsTwo months-old mice were individually infected with 80 S. mansoni cercariae and divided into one infected-untreated control group (IC) and four PZQ-treated groups: PZQ at 100 mg/kg/day for five consecutive days (group PZQ1), PZQ at 100 mg/kg/day for 28 days (group PZQ2), PZQ at 18 mg/kg/day for 28 days (group PZQ3) and a single dose of PZQ at 500 mg/kg (group PZQ4). The treatment started on day one post-infection (p.i), and each group of mice was divided into two subgroups euthanized on day 36 or 56 p.i, respectively. We determined the mortality rate, the parasitological burden, the hepatic and intestinal granulomas, the serum levels of Th-1, Th-2, and Th-17 cytokines, and gene expression. The treatment led to a significant (p < 0.001) reduction of worm burden and egg counts in the intestine and liver in groups PZQ2 and PZQ3. On 56^th day p.i, there was a significant reduction (p < 0.001) of the number and volume of the hepatic granulomas in groups PZQ2 and PZQ3 compared to group PZQ1 or PZQ4. Moreover, in group PZQ3, the serum levels of IFN-γ, TNF-α, IL-13, and IL-17 and their liver mRNA expressions were significantly reduced while IL-10 and TGF-β gene expression significantly increased. The highest mortality rate (81.25%) was recorded in group PZQ2.Conclusion/SignificanceThis study revealed that the administration of PZQ at 18 mg/kg/day for 28 consecutive days was the optimal effective posology for treating S. mansoni infection at the initial stage in a murine model.     

Heat stress responses and population genetics of the kelp Laminaria digitata (Phaeophyceae) across latitudes reveal differentiation among North Atlantic populations

To understand the thermal plasticity of a coastal foundation species across its latitudinal distribution, we assess physiological responses to high temperature stress in the kelp Laminaria digitata in combination with population genetic characteristics and relate heat resilience to genetic features and phylogeography. We hypothesize that populations from Arctic and cold‐temperate locations are less heat resilient than populations from warm distributional edges. Using meristems of natural L. digitata populations from six locations ranging between Kongsfjorden, Spitsbergen (79°N), and Quiberon, France (47°N), we performed a common‐garden heat stress experiment applying 15°C to 23°C over eight days. We assessed growth, photosynthetic quantum yield, carbon and nitrogen storage, and xanthophyll pigment contents as response traits. Population connectivity and genetic diversity were analyzed with microsatellite markers. Results from the heat stress experiment suggest that the upper temperature limit of L. digitata is nearly identical across its distribution range, but subtle differences in growth and stress responses were revealed for three populations from the species’ ecological range margins. Two populations at the species’ warm distribution limit showed higher temperature tolerance compared to other populations in growth at 19°C and recovery from 21°C (Quiberon, France), and photosynthetic quantum yield and xanthophyll pigment responses at 23°C (Helgoland, Germany). In L. digitata from the northernmost population (Spitsbergen, Norway), quantum yield indicated the highest heat sensitivity. Microsatellite genotyping revealed all sampled populations to be genetically distinct, with a strong hierarchical structure between southern and northern clades. Genetic diversity was lowest in the isolated population of the North Sea island of Helgoland and highest in Roscoff in the English Channel. All together, these results support the hypothesis of moderate local differentiation across L. digitata's European distribution, whereas effects are likely too weak to ameliorate the species’ capacity to withstand ocean warming and marine heatwaves at the southern range edge.     

Cosolute modulation of protein oligomerization reactions in the homeostatic timescale

Protein oligomerization processes are widespread and of crucial importance to understand degenerative diseases and healthy regulatory pathways. One particular case is the homo-oligomerization of folded domains involving domain swapping, often found as a part of the protein homeostasis in the crowded cytosol, composed of a complex mixture of cosolutes. Here, we have investigated the effect of a plethora of cosolutes of very diverse nature on the kinetics of a protein dimerization by domain swapping. In the absence of cosolutes, our system exhibits slow interconversion rates, with the reaction reaching the equilibrium within the average protein homeostasis timescale (24–48 h). In the presence of crowders, though, the oligomerization reaction in the same time frame will, depending on the protein's initial oligomeric state, either reach a pure equilibrium state or get kinetically trapped into an apparent equilibrium. Specifically, when the reaction is initiated from a large excess of dimer, it becomes unsensitive to the effect of cosolutes and reaches the same equilibrium populations as in the absence of cosolute. Conversely, when the reaction starts from a large excess of monomer, the reaction during the homeostatic timescale occurs under kinetic control, and it is exquisitely sensitive to the presence and nature of the cosolute. In this scenario (the most habitual case in intracellular oligomerization processes), the effect of cosolutes on the intermediate conformation and diffusion-mediated encounters will dictate how the cellular milieu affects the domain-swapping reaction.     

Occurrence of species of the genus Pityophthorus Eichhoff (Coleoptera,Curculionidae, Scolytinae) in?the?province of Quebec,Canada

Twig beetles in the genus Pityophthorus Eichhoff, 1864 include more than 300 species worldwide, with maximum diversity in tropical and subtropical regions. To date, approximately 50 species of Pityophthorus have been recorded in Canada, and these species are associated mainly with coniferous trees. Since 1981, no comprehensive study on this difficult taxonomic group has been conducted in Quebec, Canada, most likely due to their limited significance as forest pests. Based on data gathered from five years of field sampling in conifer seed orchards and compiled from various entomological collections, the distribution of Pityophthorus species in Quebec is presented. Approximately 291 new localities were recorded for the Pityophthorus species. Five species-group taxa, namely Pityophthorus puberulus (LeConte, 1868), Pityophthorus pulchellus pulchellus Eichhoff, 1869, Pityophthorus pulicarius (Zimmermann, 1868), Pityophthorus nitidus Swaine, 1917,and Pityophthorus cariniceps LeConte&Horn, 1876 were the most widespread. In contrast, Pityophthorus consimilis LeConte, 1878, Pityophthorus intextus Swaine, 1917, Pityophthorus dentifrons Blackman, 1922, Pityophthorus ramiperda Swaine, 1917, and Pityophthorus concavus Blackman, 1928 display a notably limited distribution. In addition, the first distribution records of Pityophthorus intextus and Pityophthorus biovalis Blackman, 1922 are furnished, and the subspecies Pityophthorus murrayanae murrayanae Blackman, 1922is reported from Quebec for the second time. Moreover, distribution maps are provided for all Pityophthorus species recorded in the province of Quebec.     

In vitro brown and “brite”/“beige” adipogenesis: Human cellular models and molecular aspects

Brown adipose tissue (BAT) has long been thought to be absent or very scarce in human adults so that its contribution to energy expenditure was not considered as relevant. The recent discovery of thermogenic BAT in human adults opened the field for innovative strategies to combat overweight/obesity and associated diseases. This energy-dissipating function of BAT is responsible for adaptive thermogenesis in response to cold stimulation. In this context, adipocytes can be converted, within white adipose tissue (WAT), into multilocular adipocytes expressing UCP1, a mitochondrial protein that plays a key role in heat production by uncoupling the activity of the respiratory chain from ATP synthesis. These adipocytes have been named “brite” or “beige” adipocytes. Whereas BAT has been studied for a long time in murine models both in vivo and in vitro, there is now a strong demand for human cellular models to validate and/or identify critical factors involved in the induction of a thermogenic program within adipocytes. In this review we will discuss the different human cellular models described in the literature and what is known regarding the regulation of their differentiation and/or activation process. In addition, the role of microRNAs as novel regulators of brown/“brite” adipocyte differentiation and conversion will be depicted. Finally, investigation of both the conversion and the metabolism of white-to-brown converted adipocytes is required for the development of therapeutic strategies targeting overweight/obesity and associated diseases. This article is part of a Special Issue entitled Brown and White Fat: From Signaling to Disease.     

Prediction of Severe Disease in Children with Diarrhea in a Resource-Limited Setting

Objective

To investigate the accuracy of three clinical scales for predicting severe disease (severe dehydration or death) in children with diarrhea in a resource-limited setting.

Methods

Participants included 178 children admitted to three Rwandan hospitals with diarrhea. A local physician or nurse assessed each child on arrival using the World Health Organization (WHO) severe dehydration scale and the Centers for Disease Control (CDC) scale. Children were weighed on arrival and daily until they achieved a stable weight, with a 10% increase between admission weight and stable weight considered severe dehydration. The Clinical Dehydration Scale was then constructed post-hoc using the data collected for the other two scales. Receiver Operator Characteristic (ROC) curves were constructed for each scale compared to the composite outcome of severe dehydration or death.

Results

The WHO severe dehydration scale, CDC scale, and Clinical Dehydration Scale had areas under the ROC curves (AUCs) of 0.72 (95% CI 0.60, 0.85), 0.73 (95% CI 0.62, 0.84), and 0.80 (95% CI 0.71, 0.89), respectively, in the full cohort. Only the Clinical Dehydration Scale was a significant predictor of severe disease when used in infants, with an AUC of 0.77 (95% CI 0.61, 0.93), and when used by nurses, with an AUC of 0.78 (95% CI 0.63, 0.93).

Conclusions

While all three scales were moderate predictors of severe disease in children with diarrhea, scale accuracy varied based on provider training and age of the child. Future research should focus on developing or validating clinical tools that can be used accurately by nurses and other less-skilled providers to assess all children with diarrhea in resource-limited settings.     

Two New Kremastochrysopsis species,K. austriaca sp. nov. and K. americana sp. nov. (Chrysophyceae)1

Melting summer snow in the Austrian Alps exhibited a yellowish bloom that was mainly comprised of an unidentified unicellular chrysophyte. Molecular data (18S rRNA and rbcL genes) showed a close relationship to published sequences from an American pond alga formerly identified as Kremastochrysis sp. The genera Kremastochrysis and Kremastochrysopsis are morphologically distinguished by the number of flagella observed with the light microscope, and therefore we assigned the Austrian snow alga and an American pond alga to the genus Kremastochrysopsis. Transmission and scanning electron microscopy revealed that swimming cells had two flagella oriented in opposite directions, typical for the Hibberdiales. Molecular phylogenetic analyses showed that both new species were closely related to Hibberdia. Kremastochrysopsis ocellata, the type species and only known species, has two chloroplasts per cell and the zoospores have red eyespots. Our two organisms had only a single chloroplast and no zoospore eyespot, but their gene sequences differed substantially. Therefore, we described two new species, Kremastochrysopsis austriaca sp. nov and Kremstochrysopsis americana sp. nov. When grown in culture, both taxa showed a characteristic hyponeustonic growth (hanging below the water surface), whereas older immotile cells grew at the bottom of the culture vessel. Ecologically, Kremastochrysopsis austriaca sp. nov., which caused snow discolorations, had no close phylogenetic relationships to other psychrophilic chrysophytes, for example, Chromulina chionophilia, Hydrurus sp., and Ochromonas-like flagellates.