Critical Role of the Secondary Binding Pocket in Modulating the Enzymatic Activity of DUSP5 toward Phosphorylated ERKs

DUSP5 is an inducible nuclear dual-specificity phosphatase that specifically interacts with and deactivates extracellular signal-regulated kinases ERK1 and ERK2, which are responsible for cell proliferation, differentiation, and survival. The phosphatase domain (PD) of DUSP5 has unique structural features absent from other nuclear DUSPs, such as the presence of a secondary anion-binding site in the proximity of the reaction center and a glutamic acid E264 positioned next to the catalytic cysteine C263, as well as a remote intramolecular disulfide linkage. The overall 400 ns molecular dynamics simulations indicate that the secondary binding site of DUSP5 PD acts as an allosteric regulator of the phosphatase activity of DUSP5. Our studies have identified E264 as a critical constituent of the dual binding pocket, which regulates the catalytic activity of DUSP5 by forming a salt bridge with arginine R269. Molecular dynamics studies showed that initial occupation of the secondary binding pocket leads to the breakage of the salt bridge, which then allows the occupation of the active site. Indeed, biochemical analysis using the pERK assay on mutant E264Q demonstrated that mutation of glutamic acid E264 leads to an increase in the DUSP5 catalytic activity. The role of the secondary binding site in assembling the DUSP5–pERK pre-reactive complex was further demonstrated by molecular dynamics simulations that showed that the remote C197–C219 disulfide linkage controls the structure of the secondary binding pocket based on its redox state (i.e., disulfide/dithiol) and, in turn, the enzymatic activity of DUSP5

Method for Monitoring Quality of Extension Programs: A Dashboard Construction Process

We describe a process for construction and use of a dashboard for monitoring the quality of in-person nonformal education programs. We followed the process to develop a 4-H special interest (SPIN) club dashboard, but the steps are applicable to any education program that includes instruction over multiple sessions. The dashboard construction process comprises selecting performance measures, choosing data collection strategies, designing the content and layout of the dashboard, collecting data, and populating the dashboard with performance metrics. Although these process steps are intuitive, specific decision options within each step can be complex. Our article includes discussion of these complexities in the context of our SPIN club application

Serendipitous Discovery of Light-Induced \u3cem\u3e(In Situ)\u3c/em\u3e Formation of An Azo-Bridged Dimeric Sulfonated Naphthol as a Potent PTP1B Inhibito

Background Protein tyrosine phosphatases (PTPs) like dual specificity phosphatase 5 (DUSP5) and protein tyrosine phosphatase 1B (PTP1B) are drug targets for diseases that include cancer, diabetes, and vascular disorders such as hemangiomas. The PTPs are also known to be notoriously difficult targets for designing inihibitors that become viable drug leads. Therefore, the pipeline for approved drugs in this class is minimal. Furthermore, drug screening for targets like PTPs often produce false positive and false negative results. Results Studies presented herein provide important insights into: (a) how to detect such artifacts, (b) the importance of compound re-synthesis and verification, and (c) how in situ chemical reactivity of compounds, when diagnosed and characterized, can actually lead to serendipitous discovery of valuable new lead molecules. Initial docking of compounds from the National Cancer Institute (NCI), followed by experimental testing in enzyme inhibition assays, identified an inhibitor of DUSP5. Subsequent control experiments revealed that this compound demonstrated time-dependent inhibition, and also a time-dependent change in color of the inhibitor that correlated with potency of inhibition. In addition, the compound activity varied depending on vendor source. We hypothesized, and then confirmed by synthesis of the compound, that the actual inhibitor of DUSP5 was a dimeric form of the original inhibitor compound, formed upon exposure to light and oxygen. This compound has an IC50 of 36 μM for DUSP5, and is a competitive inhibitor. Testing against PTP1B, for selectivity, demonstrated the dimeric compound was actually a more potent inhibitor of PTP1B, with an IC50 of 2.1 μM. The compound, an azo-bridged dimer of sulfonated naphthol rings, resembles previously reported PTP inhibitors, but with 18-fold selectivity for PTP1B versus DUSP5. Conclusion We report the identification of a potent PTP1B inhibitor that was initially identified in a screen for DUSP5, implying common mechanism of inhibitory action for these scaffolds

Line Surge Arresters Applications On The Compact Transmission Lines

This paper presents application of line surge arresters on the compact transmission lines. Single and double circuit compact transmission lines are considered. Line lightning performance is computed using sigma slp simulation software. Different line surge arrester installation configurations are considered. Line lightning performance is computed for different tower footing resistance. Line performance before and after line surge arrester installation are compared. Line lightning performance of the unshielded line with line surge arresters is compared with the performance of the shielded line without line surge arresters. For double circuit shielded compact lines, double circuit outage rate is computed. Influence of the tower footing resistance on the double circuit outage rate is presented. KEYWORD

Monitoring Structured Experiences during Youth Programs: Development of Brief Measures of Perceived Value and Engagement

A brief questionnaire that can be used to routinely monitor the quality of structured experiences for youth is discussed in this paper. Structured experiences are discrete periods of time in which youth gather for activity under the supervision of adult or youth leaders.  Four-item measures of perceived value and engagement were created.  A questionnaire including these measures along with items from the 4-H Common Measures was administered to 219 youth from 11 4-H clubs.  Data were analyzed for evidence of reliability and validity.  Alpha reliability estimates were .82 and .71 for the two multiple-item monitoring instruments.  Correlations (validity coefficients) ranged from .48 to .61. Multiple regression results were consistent with predictions.  Significant relations were found between perceived value, engagement, meaningfulness and supportiveness of social climate and safety of social climate.  Results thus suggest that these instruments may be appropriate for use in monitoring structured experiences for youth

Can current farmland landscapes feed declining steppe birds? Evaluating arthropod abundance for the endangered little bustard (Tetrax tetrax) in cereal farmland during the chick-rearing period: Variations between habitats and localities

Agriculture intensification threatens farmland bird populations because, among other reasons, it reduces the availability of food resources required to rear their offspring. In our study, we sampled and analyzed total arthropod abundance, biomass and richness, and orthopteran and coleopteran abundance and biomass in different agricultural habitats (alfalfa fields, stubble fields, grazed fields, and field margins) across 4 study localities with different levels of agriculture abandonment–intensification, comparing between areas used and not used by one of the most threatened farmland birds in Europe, the little bustard (Tetrax tetrax), during the chick-rearing season. Field margins were the taxonomically richest habitat, while alfalfa fields presented significantly higher total arthropod abundance and biomass than other habitats. All arthropod variables were the highest in the localities with clear conservation-focused agrarian management, and the lowest in the most intensive one. Areas used by little bustards had higher orthopteran and coleopteran abundance and biomass than nonused areas, except for coleopteran biomass in grazed fields. These results highlight the relevance of these arthropods for the species, the importance of dry alfalfa fields as food reservoirs in this critical time of year, the food scarcity in sites where agrarian management disregards farmland bird conservation, and the role of stubbles as providers of food resources during the chick-rearing season in areas used by the species. The adequate management of alfalfa fields and stubbles to provide those key resources seems crucial to improve little bustard breeding success.We thank the International Fund for Houbara Conservation (IFHC) and Fundación Patrimonio Natural (FPN) de Castilla y León in the persons of Cédric Ferlat (IFHC), David Cubero (Dirección General de Medio Natural, Junta de CyL), and Francisco Jiménez (FPN) for their financial, administrative, and logistic support. This paper also contributes to project S2018/EMT-4338 REMEDINAL TE-CM from Comunidad de Madrid. We are also thankful to the officers responsible for the Reserve of Villafáfila, Mariano Rodíguez y Jesús Palacios. Little bustard spring survey data were kindly provided by SEO-BirdLife.Peer reviewe

Cytotoxicity of apigenin on leukemia cell lines: implications for prevention and therapy

Natural-food-based compounds show substantial promise for prevention and biotherapy of cancers including leukemia. In general, their mechanism of action remains unclear, hampering rational use of these compounds. Herein we show that the common dietary flavonoid apigenin has anticancer activity, but also may decrease chemotherapy sensitivity, depending on the cell type. We analyzed the molecular consequences of apigenin treatment in two types of leukemia, the myeloid and erythroid subtypes. Apigenin blocked proliferation in both lineages through cell-cycle arrest in G2/M phase for myeloid HL60 and G0/G1 phase for erythroid TF1 cells. In both cell lines the JAK/STAT pathway was one of major targets of apigenin. Apigenin inhibited PI3K/PKB pathway in HL60 and induced caspase-dependent apoptosis. In contrast, no apoptosis was detected in TF1 cells, but initiation of autophagy was observed. The block in cell cycle and induction of autophagy observed in this erythroleukemia cell line resulted in a reduced susceptibility toward the commonly used therapeutic agent vincristine. Thus, this study shows that although apigenin is a potential chemopreventive agent due to the induction of leukemia cell-cycle arrest, caution in dietary intake of apigenin should be taken during disease as it potentially interferes with cancer treatment

Landscape determinants of European roller foraging habitat: implications for the definition of agri-environmental measures for species conservation

Across much of Europe, farmland birds are declining more than those in other habitats. From a conservation perspective, identifying the primary preferred habitats could help improve the foraging conditions of target species and, consequently, enhance their breeding success and survival. Here, we investigated the ranging behaviour and foraging habitat selection of the European roller (Coracias garrulus) during the breeding season in an agricultural landscape of South Iberia. The occurrence of foraging rollers was predicted to gradually increase with decreasing distance from the nest and increasing availability of perches, such as fences and electric wires. Traditional olive groves and stubble fields were positively and negatively associated with the occurrence of rollers, respectively. Additionally, analysis of hunting strikes showed that rollers highly prefer foraging in fallows rather than cereal or stubble fields. Prey surveys revealed that fallows had the highest abundance of grasshoppers, rollers’ preferred prey during chick-rearing. Pair home-ranges, obtained from 95% fixed Kernel estimators averaged 70.9 ha (range = 34–118 ha) and most foraging trips (80%) occurred in the close vicinity of the nest (<500 m). Number of chicks fledged was not affected by mean foraging distances travelled during the chick-rearing period. Overall, our results suggest that traditional extensive practices of cereal cultivation, with large areas of low-intensity grazed fallows, represent a high-quality foraging habitat for rollers and should be promoted through agri-environmental schemes within at least 1-km radius from the nest. These recommendations are targeted at the roller, but have been shown to apply broadly to several other steppe-bird species

Social Competitiveness and Plasticity of Neuroendocrine Function in Old Age: Influence of Neonatal Novelty Exposure and Maternal Care Reliability

Early experience is known to have a profound impact on brain and behavioral function later in life. Relatively few studies, however, have examined whether the effects of early experience remain detectable in the aging animal. Here, we examined the effects of neonatal novelty exposure, an early stimulation procedure, on late senescent rats' ability to win in social competition. During the first 3 weeks of life, half of each litter received daily 3-min exposures to a novel environment while the other half stayed in the home cage. At 24 months of age, pairs of rats competed against each other for exclusive access to chocolate rewards. We found that novelty-exposed rats won more rewards than home-staying rats, indicating that early experience exerts a life-long effect on this aspect of social dominance. Furthermore, novelty-exposed but not home-staying rats exhibited habituation of corticosterone release across repeated days of social competition testing, suggesting that early experience permanently enhances plasticity of the stress response system. Finally, we report a surprising finding that across individual rat families, greater effects of neonatal novelty exposure on stress response plasticity were found among families whose dams provided more reliable, instead of a greater total quantity of, maternal care