Nucleosomes represent mechanical and energetic barriers that RNA Polymerase II (Pol II) must overcome during transcription. A high-resolution description of the barrier topography, its modulation by epigenetic modifications, and their effects on Pol II nucleosome crossing dynamics, is still missing. Here, we obtain topographic and transcriptional (Pol II residence time) maps of canonical, H2A.Z, and monoubiquitinated H2B (uH2B) nucleosomes at near base-pair resolution and accuracy. Pol II crossing dynamics are complex, displaying pauses at specific loci, backtracking, and nucleosome hopping between wrapped states. While H2A.Z widens the barrier, uH2B heightens it, and both modifications greatly lengthen Pol II crossing time. Using the dwell times of Pol II at each nucleosomal position we extract the energetics of the barrier. The orthogonal barrier modifications of H2A.Z and uH2B, and their effects on Pol II dynamics rationalize their observed enrichment in +1 nucleosomes and suggest a mechanism for selective control of gene expression.

Public health interventions should be designed with scale in mind, and researchers and implementers must plan for scale-up at an early stage. Yet, there is limited awareness among researchers of the critical value of considering scalability and relatively limited empirical evidence on assessing scalability, despite emerging methodological guidance. We aimed to integrate scalability considerations in the design of a study to evaluate a multi-component intervention to reduce unnecessary caesarean sections in low- and middle-income countries. First, we reviewed and synthesized existing scale up frameworks to identify relevant dimensions and available scalability assessment tools. Based on these, we defined our scalability assessment process and adapted existing tools for our study. Here, we document our experience and the methodological challenges we encountered in integrating a scalability assessment in our study protocol. These include: achieving consensus on the purpose of a scalability assessment; and identifying the optimal timing of such an assessment, moving away from the concept of a one-off assessment at the start of a project. We also encountered tensions between the need to establish the proof of principle, and the need to design an innovation that would be fit-for-scale. Particularly for complex interventions, scaling up may warrant rigorous research to determine an efficient and effective scaling-up strategy. We call for researchers to better incorporate scalability considerations in pragmatic trials through greater integration of impact and process evaluation, more stringent definition and measurement of scale-up objectives and outcome evaluation plans that allow for comparison of effects at different stages of scale-up.

The reversible oxidation of methionine residues in proteins has emerged as a biologically important post-translational modification. However, detection and quantitation of methionine sulfoxide in proteins is difficult. Our aim is to develop a method for specifically derivatizing methionine sulfoxide residues. We report a Pummerer rearrangement of methionine sulfoxide treated sequentially with trimethylsilyl chloride and then 2-mercaptoimidazole or pyridine-2-thiol to produce a dithioacetal product. This derivative is stable to standard mass spectrometry conditions, and its formation identified oxidized methionine residues. The scope and requirements of dithioacetal formation are reported for methionine sulfoxide and model substrates. The reaction intermediates have been investigated by computational techniques and by 13 C NMR spectroscopy. These provide evidence for an α-chlorinated intermediate. The derivatization allows for detection and quantitation of methionine sulfoxide in proteins by mass spectrometry and potentially by immunochemical methods.

A library of poly(2-oxazoline)s functionalized with controllable amounts of alendronate, hydroxyl, and carboxylic acid side groups was successfully synthesized to create novel polymers with tunable affinity for calcium cations. The affinity of alendronate-containing polymers for calcium cations was quantified using isothermal titration calorimetry. Thermodynamic measurements revealed that the Ca2+-binding affinity of these polymers increased linearly with the amount of alendronate functionalization, up to values (KCa2+ = 2.4 × 105 M-1) that were about 120-fold higher than those for previously reported polymers. The calcium-binding capacity of alendronate-functionalized poly(2-oxazoline)s was exploited to form robust hydrogel networks cross-linked using reversible physical bonds. Oscillatory rheology showed that these hydrogels recovered more than 100% of their initial storage modulus after severe network destruction. The versatile synthesis of alendronate-functionalized polymers and their strong and tunable affinity for calcium cations render these polymers promising candidates for various biomedical applications.

Kinetic parameters kex (s-1) and kex/Kd (M-1 s-1) are reported for exchange for deuterium in D2O of the C-6 hydrogen of 5-fluororotidine 5'-monophosphate (FUMP) catalyzed by the Q215A, Y217F, and Q215A/Y217F variants of yeast orotidine 5'-monophosphate decarboxylase (ScOMPDC) at pD 8.1, and by the Q215A variant at pD 7.1-9.3. The pD rate profiles for wildtype ScOMPDC and the Q215A variant are identical, except for a 2.5 log unit downward displacement in the profile for the Q215A variant. The Q215A, Y217F and Q215A/Y217F substitutions cause 1.3-2.0 kcal/mol larger increases in the activation barrier for wildtype ScOMPDC-catalyzed deuterium exchange compared with decarboxylation, because of the stronger apparent side chain interaction with the transition state for the deuterium exchange reaction. The stabilization of the transition state for the OMPDC-catalyzed deuterium exchange reaction of FUMP is ca. 19 kcal/mol smaller than the transition state for decarboxylation of OMP, and ca. 8 kcal/mol smaller than for OMPDC-catalyzed deprotonation of FUMP to form the vinyl carbanion intermediate common to OMPDC-catalyzed reactions OMP/FOMP and UMP/FUMP. We propose that ScOMPDC shows similar stabilizing interactions with the common portions of decarboxylation and deprotonation transition states that lead to formation of this vinyl carbanion intermediate, and that there is a large ca. (19-8) = 11 kcal/mol stabilization of the former transition state from interactions with the nascent CO2 of product. The effects of Q215A and Y217F substitutions on kcat/Km for decarboxylation of OMP are expressed mainly as an increase in Km for the reactions catalyzed by the variant enzymes, while the effects on kex/Kd for deuterium exchange are expressed mainly as an increase in kex. This shows that the Q215 and Y217 side chains stabilize the Michaelis complex to OMP for the decarboxylation reaction, compared with the complex to FUMP for the deuterium exchange reaction. These results provide strong support for the conclusion that interactions which stabilize the transition state for ScOMPDC-catalyzed decarboxylation at a nonpolar enzyme active site dominate over interactions that destabilize the ground-state Michaelis complex.

Oncology-associated adverse drug/device reactions can be fatal. Some clinicians who treat single patients with severe oncology-associated toxicities have researched case series and published this information. We investigated motivations and experiences of select individuals leading such efforts. Clinicians treating individual patients who developed oncology-associated serious adverse drug events were asked to participate. Inclusion criteria included having index patient information, reporting case series, and being collaborative with investigators from two National Institutes of Health funded pharmacovigilance networks. Thirty-minute interviews addressed investigational motivation, feedback from pharmaceutical manufacturers, FDA personnel, and academic leadership, and recommendations for improving pharmacovigilance. Responses were analyzed using constant comparative methods of qualitative analysis. Overall, 18 clinicians met inclusion criteria and 14 interviewees are included. Primary motivations were scientific curiosity, expressed by six clinicians. A less common theme was public health related (three clinicians). Six clinicians received feedback characterized as supportive from academic leaders, while four clinicians received feedback characterized as negative. Three clinicians reported that following the case series publication they were invited to speak at academic institutions worldwide. Responses from pharmaceutical manufacturers were characterized as negative by 12 clinicians. One clinician's wife called the post-reporting time the "Maalox month," while another clinician reported that the manufacturer collaboratively offered to identify additional cases of the toxicity. Responses from FDA employees were characterized as collaborative for two clinicians, neutral for five clinicians, unresponsive for negative by six clinicians. Three clinicians endorsed developing improved reporting mechanisms for individual physicians, while 11 clinicians endorsed safety activities that should be undertaken by persons other than a motivated clinician who personally treats a patient with a severe adverse drug/device reaction. Our study provides some of the first reports of clinician motivations and experiences with reporting serious or potentially fatal oncology-associated adverse drug or device reactions. Overall, it appears that negative feedback from pharmaceutical manufacturers and mixed feedback from the academic community and/or the FDA were reported. Big data, registries, Data Safety Monitoring Boards, and pharmacogenetic studies may facilitate improved pharmacovigilance efforts for oncology-associated adverse drug reactions. These initiatives overcome concerns related to complacency, indifference, ignorance, and system-level problems as barriers to documenting and reporting adverse drug events- barriers that have been previously reported for clinician reporting of serious adverse drug reactions.

Variation in synonymous codon usage is abundant across multiple levels of organization: between codons of an amino acid, between genes in a genome, and between genomes of different species. It is now well understood that variation in synonymous codon usage is influenced by mutational bias coupled with both natural selection for translational efficiency and genetic drift, but how these processes shape patterns of codon usage bias across entire lineages remains unexplored. To address this question, we used a rich genomic data set of 327 species that covers nearly one third of the known biodiversity of the budding yeast subphylum Saccharomycotina. We found that, while genome-wide relative synonymous codon usage (RSCU) for all codons was highly correlated with the GC content of the third codon position (GC3), the usage of codons for the amino acids proline, arginine, and glycine was inconsistent with the neutral expectation where mutational bias coupled with genetic drift drive codon usage. Examination between genes' effective numbers of codons and their GC3 contents in individual genomes revealed that nearly a quarter of genes (381,174/1,683,203; 23%), as well as most genomes (308/327; 94%), significantly deviate from the neutral expectation. Finally, by evaluating the imprint of translational selection on codon usage, measured as the degree to which genes' adaptiveness to the tRNA pool were correlated with selective pressure, we show that translational selection is widespread in budding yeast genomes (264/327; 81%). These results suggest that the contribution of translational selection and drift to patterns of synonymous codon usage across budding yeasts varies across codons, genes, and genomes; whereas drift is the primary driver of global codon usage across the subphylum, the codon bias of large numbers of genes in the majority of genomes is influenced by translational selection.

Cervicitis is one of the major health problems amongst women caused by infection of various pathogens including Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV) as well as human papillomavirus (HPV), and persistent cervical inflammation is one of the etiologic agents of cervical cancer. Toll-like receptors (TLRs) play an important role in the recognition and subsequent elimination of these pathogens. Variations in the Toll-like receptor genes influence susceptibility to pathogens as well as disease progression independently.

Ultrasonic vocalizations (USV) are emitted by mice under certain developmental, social and behavioral conditions. The analysis of USV can be used as a reliable measure of the general affective state, for testing the efficacy of pharmacological compounds and for investigating communication in mutant mice with predicted social or communication deficits. Social and communication studies in mice have focused mainly on the investigation of USV emitted by neonatal pups after separation from the dam and during social interaction between adult males and females. Longitudinal USV analysis among the different developmental states remained uninvestigated. In our study, we first recorded USV from three inbred mouse strains C57BL/6N, DBA/2 and FVB/N during the neonatal stages after separation from the littermates and then during a reunion with one littermate. Our results revealed significant strain-specific differences in the numbers and categories of USV calls. In addition, the USV profiles seemed to be sensitive to small developmental progress during infancy. By following these mice to the adolescent stage and measuring USV in the three-chamber social test, we found that USV profiles still showed significant differences between these strains in the different trials of the test. To study the effects of social context on USV characteristics, we measured USV emitted by another cohort of adolescent mice during the direct social interaction test. To this end, this study provides a strategy for evaluating novel mouse mutants in behavioral questions relevant to disorders with deficits in communication and sociability and emphasizes the important contribution of genetics and experimental contexts on the behavioral outcome.

The development of cancer mouse models is still needed for the identification and preclinical validation of novel therapeutic targets in colorectal cancer, which is the third leading cause of cancer-related deaths in Europe. The purpose of this study was to determine the most accurate tumour cell injection method to obtain suitable peritoneal metastasis (PM) for subsequent therapeutic treatments. Here, we grafted murine colon carcinoma CT-26 cells expressing luciferase into immunocompetent BALB-c mice by intravenous injection (IV group), subcutaneous injection (SC group), intraperitoneal injection after peritoneal scratching (A group) or intraperitoneal injection alone (IP group). Tumour growth was monitored by bioluminescence during the first 15 days post-grafting. The peritoneal carcinomatosis index was evaluated macroscopically, histology, immunohistochemistry and multiphoton microscopy were performed in peritoneal tumour tissue. Upon implantation, no tumour growth was observed in the IV group, similar to the non-injected group. Both the IP and SC groups showed intermediate growth rates, but the SC group produced only a single subcutaneous nodule. The A group exhibited the highest tumour growth at 15 days post-surgery. Anatomic and histologic analyses corroborated the existence of various tumour nodules, and multiphoton microscopy was used to evaluate tumour fibrosis-infiltrating cells in a non-pathologic peritoneum. In conclusion, limited PM was obtained by IP injection, whereas IP injection after peritoneal scratching led to an extensive PM murine model for evaluating new therapeutics.

Rain cracking severely affects the commercial production of many fleshy-fruit species, including of sweet cherries. The objectives were to investigate how the gaping macroscopic cracks (macrocracks) of a rain-cracked fruit can develop from microscopic cracks in the cuticle (microcracks). Incubating fruit in deionized water is well known to cause significant macrocracking. We found that after a lag phase of 2 h, the numbers and lengths of macrocracks increased. Macrocrack number approached an asymptote at 12 h, whereas macrocrack length continued to increase. The rate of macrocrack propagation (extension at the crack tip) was initially 10.8 mm h-1 but then decreased to a near-constant 0.5 mm h-1. Light microscopy revealed three characteristic zones along a developing macrocrack. In zone I (ahead of the crack), the cuticle was intact, the epidermal cells were unbroken and their cell walls were thin. In zone II, the cuticle was fractured, the first epidermal cells died and their cell walls began to thicken (swell). In zone III, most epidermal cells had died, their cell walls were swollen and cell:cell separation began along the middle lamellae. The thickness of the anticlinal epidermal cell walls and the percentage of intact living cells along a crack were closely and negatively related. Cracks were stained by calcofluor white, but there was no binding of monoclonal antibodies (mAbs) specific for hemicelluloses (LM11, LM21, LM25). Strong binding was obtained with the anti-homogalacturonan mAb (LM19), indicating the presence of unesterified homogalacturonans on the crack surface. We conclude that macrocrack propagation is related to cell death and to cell wall swelling. Cell wall swelling weakens the cell:cell adhesion between neighbouring epidermal cells, which separate along their middle lamellae. The skin macrocrack propagates like a 'run' in a fine, knitted fabric.

In this research work, carbon nanofibers (CNFs) were synthesized on honeycomb monolith substrates using injection chemical vapor deposition (ICVD) technique. The effect of various wash-coated materials and catalyst promoter on the growth rate of CNFs on monolith substrates were examined. The characteristics of the synthesized CNFs-coated monolith composites were examined using Raman spectroscopy, Brunauer-Emmett-Teller (BET), thermogravimetric analysis (TGA), field emission scanning electron microscopy (FE-SEM), and Transmission electron microscopy (TEM) techniques. According to the textural characterization study, the specific surface area and pore volume of CNFs-coated monolith composites were significantly improved as compared to bare monolith which might be attributed to the growth of highly pure and aligned CNFs over monolith substrate. Besides that, the synthesized CNFs-coated monolith possessed extremely well thermal stability up to the temperature of 550 °C which was corresponded to the strong attachment of highly graphitized CNFs over monolith substrates.

To elicit patterns in pathogenic biofilm composition we characterized the oral microbiome present in patients with dentin caries in comparison to healthy subjects.

Pathogens can impact host survival, fecundity, and population dynamics even when no obvious disease is observed. Few baseline data on pathogen prevalence and diversity of caribou are available, which hampers our ability to track changes over time and evaluate impacts on caribou health. Archived blood samples collected from ten migratory caribou herds in Canada and two in Greenland were used to test for exposure to pathogens that have the potential to effect population productivity, are zoonotic or are emerging. Relationships between seroprevalence and individual, population, and other health parameters were also examined. For adult caribou, the highest overall seroprevalence was for alphaherpesvirus (49%, n = 722), pestivirus (49%, n = 572) and Neospora caninum (27%, n = 452). Lower seroprevalence was found for parainfluenza virus type 3 (9%, n = 708), Brucella suis (2%, n = 758), and Toxoplasma gondii (2%, n = 706). No animal tested positive for antibodies against West Nile virus (n = 418) or bovine respiratory syncytial virus (n = 417). This extensive multi-pathogen survey of migratory caribou herds provides evidence that caribou are exposed to pathogens that may have impacts on herd health and revealed potential interactions between pathogens as well as geographical differences in pathogen exposure that could be linked to the bio-geographical history of caribou. Caribou are a keystone species and the socio-economic cornerstone of many indigenous cultures across the North. The results from this study highlight the urgent need for a better understanding of pathogen diversity and the impact of pathogens on caribou health.

Deep learning algorithms have achieved human-equivalent performance in image recognition. However, the majority of clinical data within electronic health records is inherently in a non-image format. Therefore, creating visual representations of clinical data could facilitate using cutting-edge deep learning models for predicting outcomes such as in-hospital mortality, while enabling clinician interpretability. The objective of this study was to develop a framework that first transforms longitudinal patient data into visual timelines and then utilizes deep learning to predict in-hospital mortality.

The reliability of MyotonPRO that can monitor the mechanical properties of tissues is still unclear. This study aimed to analyze the within-day inter-operator and between-day intra-operator reliability of MyotonPRO for assessing tone and stiffness of quadriceps femoris and patellar tendon at different knee angles. The tone and stiffness of healthy participants (15 males and 15 females, aged 24.7±1.6 years) in the supine and resting position were measured using the MyotonPRO device. The measurements were quantified at 0°, 30°, 60°, and 90° of knee flexion. The intraclass correlation coefficient (ICC), standard error of measurement (SEM), and minimal detectable change (MDC) were calculated and a Bland-Altman analysis was conducted to estimate reliability. The results indicated excellent inter-operator reliability (ICC > 0.78) and good to excellent intra-operator reliability (ICC > 0.41). The inter-operator SEM measurements ranged between 0.1-0.9 Hz and 3.8-37.9 N/m, and intra-operator SEM ranged between 0.5-1.3 Hz and 7.9-52.0 N/m. The inter-operator MDC ranged between 0.3-2.5 Hz and 10.5-105.1 N/m, and intra-operator SEM ranged between 1.1-3.3 Hz and 21.9-144.1 N/m. The agreement of inter-operator was better than that of intra-operator. The study concluded that MyotonPRO is a reliable device to detect the tone and stiffness of quadriceps femoris and patellar tendon.

Beyond subject matter learning, primary science teaching aims at encouraging positive attitudes toward, and lasting interest in, science. This study tested whether the learning and science commitment of 8- to 10-year old pupils was affected by extracurricular learning opportunities involving repeated interactions with free-living northern bald ibis (Geronticus eremita; an endangered bird species) and professional researchers. To examine the project's efficacy, pupils' learning progress was monitored by repeated measures of knowledge (i) about northern bald ibises in specific, and (ii) of bird diversity in general. In addition, the children's attitudes toward science were monitored and their dynamic mental models of northern bald ibis' morphology and size were assessed from drawings. A total of 55 pupils from two schools were tested for eight months (before, during and after the experience). Control groups went through regular (curricular) science lessons with similar content and time investment. The extracurricular experience produced a clear learning progress with large effect sizes, which was particularly evident on a long timescale. In addition to subject matter knowledge, the project group pupils could name more bird species and expressed their views about the importance of scientific research for society in a higher proportion than control groups. Differences between schools suggest that project participation also changed the teacher's own interest in northern bald ibises, which affected learning in the control group taught by the same teacher. Beyond the pupils' language and reading skills, learning progress was also visible by scoring ibis-typical features in drawings; their mental models of relative size were not changed due to project participation, however. The results are discussed in reference to inducing and maintaining pupils' attitude and interest in a topic. The study adds empirical evidence for the potentials of advancing primary science education e. g. by fostering the collaboration of schools with scientists.

Brain-derived neurotrophic factor (BDNF) is widely expressed in the central nervous system and prolongs the survival of dopaminergic neurons in the substantia nigra. Several studies have recently investigated the association between BDNF G196A (Val66Met), a single nucleotide polymorphism influencing cognitive processes, and cognitive impairment in Parkinson's disease (PD), but with contradictory findings. Thus, this meta-analysis was performed to clarify the possible association. Relevant studies were identified by a systematic search of PubMed, Embase, and China National Knowledge Infrastructure (CNKI) databases. The strength of the association was evaluated using crude odds ratios and 95% confidence interval. Finally, six studies involving 532 cases and 802 controls were included. Our analyses suggested the G196A (Val66Met) polymorphism was significantly associated with cognitive impairment in PD, especially in Caucasian populations. In conclusion, BDNF G196A (Val66Met) is confirmed to be a risk factor for cognitive impairment in PD.

Plant-parasitic nematodes secrete numerous effectors to facilitate parasitism, but detailed functions of nematode effectors and their plant targets remain largely unknown. Here, we characterized four macrophage migration inhibitory factors (MIFs) in Meloidogyne incognita resembling the MIFs secreted by human and animal parasites. Transcriptional data showed MiMIFs are up-regulated in parasitism. Immunolocalization provided evidence that MiMIF proteins are secreted from the nematode hypodermis to the parasite surface, detected in plant tissues and giant cells. In planta MiMIFs RNA interference in Arabidopsis decreased infection and nematode reproduction. Transient expression of MiMIF-2 could suppress Bax- and RBP1/Gpa2-induced cell death. MiMIF-2 ectopic expression led to higher levels of Arabidopsis susceptibility, suppressed immune responses triggered by flg22, and impaired [Ca2+]cyt influx induced by H2O2. The immunoprecipitation of MiMIF-2-interacting proteins, followed by co-immunoprecipitation and bimolecular fluorescence complementation validations, revealed specific interactions between MiMIF-2 and two Arabidopsis annexins, AnnAt1 and AnnAt4, involved in the transport of calcium ions, stress responses, and signal transduction. Suppression of expression or overexpression of these annexins modified nematode infection. Our results provide functional evidence that nematode effectors secreted from hypodermis to the parasite cuticle surface target host proteins and M. incognita uses MiMIFs to promote parasitism by interfering with the annexin-mediated plant immune responses.

Skeletal muscle cell (myofiber) atrophy is a detrimental component of aging and cancer that primarily results from muscle protein degradation via the proteasome and ubiquitin ligases. Transcriptional upregulation of some ubiquitin ligases contributes to myofiber atrophy, but little is known about the role that most other ubiquitin ligases play in this process. To address this question, we have used RNAi screening in Drosophila to identify the function of > 320 evolutionarily conserved ubiquitin ligases in myofiber size regulation in vivo. We find that whereas RNAi for some ubiquitin ligases induces myofiber atrophy, loss of others (including the N-end rule ubiquitin ligase UBR4) promotes hypertrophy. In Drosophila and mouse myofibers, loss of UBR4 induces hypertrophy via decreased ubiquitination and degradation of a core set of target proteins, including the HAT1/RBBP4/RBBP7 histone-binding complex. Together, this study defines the repertoire of ubiquitin ligases that regulate myofiber size and the role of UBR4 in myofiber hypertrophy.