Antagonistic and cooperative AGO2-PUM interactions in regulating mRNAs.

Approximately 1500 RNA-binding proteins (RBPs) profoundly impact mammalian cellular function by controlling distinct sets of transcripts, often using sequence-specific binding to 3' untranslated regions (UTRs) to regulate mRNA stability and translation. Aside from their individual effects, higher-order combinatorial interactions between RBPs on specific mRNAs have been proposed to underpin the regulatory network. To assess the extent of such co-regulatory control, we took a global experimental approach followed by targeted validation to examine interactions between two well-characterized and highly conserved RBPs, Argonaute2 (AGO2) and Pumilio (PUM1 and PUM2). Transcriptome-wide changes in AGO2-mRNA binding upon PUM knockdown were quantified by CLIP-seq, and the presence of PUM binding on the same 3'UTR corresponded with cooperative and antagonistic effects on AGO2 occupancy. In addition, PUM binding sites that overlap with AGO2 showed differential, weakened binding profiles upon abrogation of AGO2 association, indicative of cooperative interactions. In luciferase reporter validation of candidate 3'UTR sites where AGO2 and PUM colocalized, three sites were identified to host antagonistic interactions, where PUM counteracts miRNA-guided repression. Interestingly, the binding sites for the two proteins are too far for potential antagonism due to steric hindrance, suggesting an alternate mechanism. Our data experimentally confirms the combinatorial regulatory model and indicates that the mostly repressive PUM proteins can change their behavior in a context-dependent manner. Overall, the approach underscores the importance of further elucidation of complex interactions between RBPs and their transcriptome-wide extent

Lump correction and identification in the combined thermal/epithermal neutron (CTEN) method

The authors present a model for self shielding in lumps of fissile material in active-neutron assays. The model combines the formula for self-attenuation of gamma-ray in lumpy sources with the multi-group analysis techniques used in neutron transport calculations. Models for thin foils and for spheres are examined in terms of error multiplication in determining lump corrections and the basic accuracy of the model

Defining Metrics for Short Term Success After LVAD Implant: An Analysis of the Society of Thoracic Surgeons Intermacs Registry

Purpose: While clinical trials evaluating left ventricular assist device (LVAD) technology typically use composite outcomes to assess efficacy, composite outcomes including patient reported outcomes (PROs) have not been utilized as benchmarks for LVAD implant center performance improvement initiatives or quality ranking. The objective of the study was to assess the feasibility of generating a patient composite outcome measure including PROs from a real world registry. Methods: Short term (ST, 180 days) adverse events (AEs) and mortality were tallied for Intermacs patients undergoing LVAD implant between 1/2012 and 12/2019. ST postoperative events included mortality on first device and frequencies of stroke, reoperation (device malfunction/other), right heart failure (RHF), prolonged respiratory failure, and/or dialysis on first device. Logistic regression was used to generate odds ratios for mortality for each AE. Separately, the EuroQOL visual analog scale (VAS) was assessed at baseline and 180 days in ST survivors. Results: Of 20,115 patients, 37% suffered at least one event, most commonly death, reoperation and stroke (Table, column A). Stroke, prolonged respiratory failure, and dialysis attributed the most to ST mortality (Table, column B). Of the 16725 patients alive at 180 days, 43% completed a VAS with 82.0% showing VAS improvement. Renal failure and RHF contributed most to failure to improve VAS (Figure). Conclusion: Assessment of a ST composite outcome metric after LVAD implant from a real world data source is feasible but limited by incomplete PRO reporting. ST adverse events display differential effects on mortality and PROs that can be used in development of global rank outcome scores. While reoperation is common, stroke, prolonged respiratory failure and renal failure conferred highest risks of ST deaths within Intermacs. Assessment of PROs should become a priority for LVAD centers to allow the field to generate a complete assessment of patient-centered outcomes

In-beam spectroscopy using the (t,p) reaction: recent results near A = 100

Charged particle spectroscopy using the (t,p) reaction has been employed for more than two decades to study the low-energy structure of nuclei. This reaction has contributed significantly to the elucidation of single-particle and collective phenomena for neutron rich nuclei in virtually every mass region. We have begun to use the (t,p) reaction in conjunctionuclei with in-beam ..gamma..-ray and conversion-electron spectroscopy to bring additional understanding to low-energy nuclear structure. In this report we briefly discuss the experimental considerations in using this reaction for in-beam spectroscopy, and present some results for nuclei with mass near 100

Methylation-sensitive linking libraries enhance gene-enriched sequencing of complex genomes and map DNA methylation domains

<p>Abstract</p> <p>Background</p> <p>Many plant genomes are resistant to whole-genome assembly due to an abundance of repetitive sequence, leading to the development of gene-rich sequencing techniques. Two such techniques are hypomethylated partial restriction (HMPR) and methylation spanning linker libraries (MSLL). These libraries differ from other gene-rich datasets in having larger insert sizes, and the MSLL clones are designed to provide reads localized to "epigenetic boundaries" where methylation begins or ends.</p> <p>Results</p> <p>A large-scale study in maize generated 40,299 HMPR sequences and 80,723 MSLL sequences, including MSLL clones exceeding 100 kb. The paired end reads of MSLL and HMPR clones were shown to be effective in linking existing gene-rich sequences into scaffolds. In addition, it was shown that the MSLL clones can be used for anchoring these scaffolds to a BAC-based physical map. The MSLL end reads effectively identified epigenetic boundaries, as indicated by their preferential alignment to regions upstream and downstream from annotated genes. The ability to precisely map long stretches of fully methylated DNA sequence is a unique outcome of MSLL analysis, and was also shown to provide evidence for errors in gene identification. MSLL clones were observed to be significantly more repeat-rich in their interiors than in their end reads, confirming the correlation between methylation and retroelement content. Both MSLL and HMPR reads were found to be substantially gene-enriched, with the <it>Sal</it>I MSLL libraries being the most highly enriched (31% align to an EST contig), while the HMPR clones exhibited exceptional depletion of repetitive DNA (to ~11%). These two techniques were compared with other gene-enrichment methods, and shown to be complementary.</p> <p>Conclusion</p> <p>MSLL technology provides an unparalleled approach for mapping the epigenetic status of repetitive blocks and for identifying sequences mis-identified as genes. Although the types and natures of epigenetic boundaries are barely understood at this time, MSLL technology flags both approximate boundaries and methylated genes that deserve additional investigation. MSLL and HMPR sequences provide a valuable resource for maize genome annotation, and are a uniquely valuable complement to any plant genome sequencing project. In order to make these results fully accessible to the community, a web display was developed that shows the alignment of MSLL, HMPR, and other gene-rich sequences to the BACs; this display is continually updated with the latest ESTs and BAC sequences.</p

Genetic Diversity of a Parasitic Weed, Striga hermonthica, on Sorghum and Pearl Millet in Mali

Eleven populations of witchweed, Striga hermonthica, were collected in four regions of Mali and investigated with 12 microsatellite markers. Extensive genetic diversity was observed, with most plants heterozygous for most markers. Allelic diversity was broadly distributed across populations with little genetic differentiation and large amounts of gene flow. Nearby fields of pearl millet and sorghum were found to have indistinguishable witchweed populations. Some population structure was apparent, but did not correlate with the local environment or host genotype, suggesting that seed transportation or other human-driven variables act to differentiate central Malian S. hermonthica populations from southern Malian populations

Kinase-activating and kinase-impaired cardio-facio-cutaneous syndrome alleles have activity during zebrafish development and are sensitive to small molecule inhibitors

The Ras/MAPK pathway is critical for human development and plays a central role in the formation and progression of most cancers. Children born with germ-line mutations in BRAF, MEK1 or MEK2 develop cardio-facio-cutaneous (CFC) syndrome, an autosomal dominant syndrome characterized by a distinctive facial appearance, heart defects, skin and hair abnormalities and mental retardation. CFC syndrome mutations in BRAF promote both kinase-activating and kinase-impaired variants. CFC syndrome has a progressive phenotype, and the availability of clinically active inhibitors of the MAPK pathway prompts the important question as to whether such inhibitors might be therapeutically effective in the treatment of CFC syndrome. To study the developmental effects of CFC mutant alleles in vivo, we have expressed a panel of 28 BRAF and MEK alleles in zebrafish embryos to assess the function of human disease alleles and available chemical inhibitors of this pathway. We find that both kinase-activating and kinase-impaired CFC mutant alleles promote the equivalent developmental outcome when expressed during early development and that treatment of CFC-zebrafish embryos with inhibitors of the FGF-MAPK pathway can restore normal early development. Importantly, we find a developmental window in which treatment with a MEK inhibitor can restore the normal early development of the embryo, without the additional, unwanted developmental effects of the drug

An evaluation of the effectiveness of PROMPT therapy in improving speech production accuracy in six children with cerebral palsy

This study evaluates perceptual changes in speech production accuracy in six children (3 – 11 years) with moderate-to-severe speech impairment associated with cerebral palsy before, during, and after participation in a motor-speech intervention program (Prompts for Restructuring Oral Muscular Phonetic Targets). An A1BCA2 single subject research design was implemented. Subsequent to the baseline phase (phase A1), phase B targeted each participant’s first intervention priority on the PROMPT motor-speech hierarchy. Phase C then targeted one level higher. Weekly speech probes were administered, containing trained and untrained words at the two levels of intervention, plus an additional level that served as a control goal. The speech probes were analysed for motor-speech-movement-parameters and perceptual accuracy. Analysis of the speech probe data showed all participants recorded a statistically significant change. Between phases A1 – B and B – C 6/6 and 4/6participants, respectively, recorded a statistically significant increase in performance level on the motor speech movement patterns targeted during the training of that intervention. The preliminary data presented in this study make a contribution to providing evidence that supports the use of a treatment approach aligned with dynamic systems theory to improve the motor-speech movement patterns and speech production accuracy in children with cerebral palsy

Global, regional, and national comparative risk assessment of 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks, 1990-2015: a systematic analysis for the Global Burden of Disease Study 2015

SummaryBackground The Global Burden of Diseases, Injuries, and Risk Factors Study 2015 provides an up-to-date synthesis of the evidence for risk factor exposure and the attributable burden of disease. By providing national and subnational assessments spanning the past 25 years, this study can inform debates on the importance of addressing risks in context. Methods We used the comparative risk assessment framework developed for previous iterations of the Global Burden of Disease Study to estimate attributable deaths, disability-adjusted life-years (DALYs), and trends in exposure by age group, sex, year, and geography for 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks from 1990 to 2015. This study included 388 risk-outcome pairs that met World Cancer Research Fund-defined criteria for convincing or probable evidence. We extracted relative risk and exposure estimates from randomised controlled trials, cohorts, pooled cohorts, household surveys, census data, satellite data, and other sources. We used statistical models to pool data, adjust for bias, and incorporate covariates. We developed a metric that allows comparisons of exposure across risk factors—the summary exposure value. Using the counterfactual scenario of theoretical minimum risk level, we estimated the portion of deaths and DALYs that could be attributed to a given risk. We decomposed trends in attributable burden into contributions from population growth, population age structure, risk exposure, and risk-deleted cause-specific DALY rates. We characterised risk exposure in relation to a Socio-demographic Index (SDI). Findings Between 1990 and 2015, global exposure to unsafe sanitation, household air pollution, childhood underweight, childhood stunting, and smoking each decreased by more than 25%. Global exposure for several occupational risks, high body-mass index (BMI), and drug use increased by more than 25% over the same period. All risks jointly evaluated in 2015 accounted for 57·8% (95% CI 56·6–58·8) of global deaths and 41·2% (39·8–42·8) of DALYs. In 2015, the ten largest contributors to global DALYs among Level 3 risks were high systolic blood pressure (211·8 million [192·7 million to 231·1 million] global DALYs), smoking (148·6 million [134·2 million to 163·1 million]), high fasting plasma glucose (143·1 million [125·1 million to 163·5 million]), high BMI (120·1 million [83·8 million to 158·4 million]), childhood undernutrition (113·3 million [103·9 million to 123·4 million]), ambient particulate matter (103·1 million [90·8 million to 115·1 million]), high total cholesterol (88·7 million [74·6 million to 105·7 million]), household air pollution (85·6 million [66·7 million to 106·1 million]), alcohol use (85·0 million [77·2 million to 93·0 million]), and diets high in sodium (83·0 million [49·3 million to 127·5 million]). From 1990 to 2015, attributable DALYs declined for micronutrient deficiencies, childhood undernutrition, unsafe sanitation and water, and household air pollution; reductions in risk-deleted DALY rates rather than reductions in exposure drove these declines. Rising exposure contributed to notable increases in attributable DALYs from high BMI, high fasting plasma glucose, occupational carcinogens, and drug use. Environmental risks and childhood undernutrition declined steadily with SDI; low physical activity, high BMI, and high fasting plasma glucose increased with SDI. In 119 countries, metabolic risks, such as high BMI and fasting plasma glucose, contributed the most attributable DALYs in 2015. Regionally, smoking still ranked among the leading five risk factors for attributable DALYs in 109 countries; childhood underweight and unsafe sex remained primary drivers of early death and disability in much of sub-Saharan Africa. Interpretation Declines in some key environmental risks have contributed to declines in critical infectious diseases. Some risks appear to be invariant to SDI. Increasing risks, including high BMI, high fasting plasma glucose, drug use, and some occupational exposures, contribute to rising burden from some conditions, but also provide opportunities for intervention. Some highly preventable risks, such as smoking, remain major causes of attributable DALYs, even as exposure is declining. Public policy makers need to pay attention to the risks that are increasingly major contributors to global burden. Funding Bill & Melinda Gates Foundation

MicroRNA-155 Deficiency Attenuates Liver Steatosis and Fibrosis without Reducing Inflammation in a Mouse Model of Steatohepatitis

BACKGROUND & AIM: MicroRNAs (miRs) regulate hepatic steatosis, inflammation and fibrosis. Fibrosis is the consequence of chronic tissue damage and inflammation. We hypothesized that deficiency of miR-155, a master regulator of inflammation, attenuates steatohepatitis and fibrosis. METHODS: Wild type (WT) and miR-155-deficient (KO) mice were fed methionine-choline-deficient (MCD) or -supplemented (MCS) control diet for 5 weeks. Liver injury, inflammation, steatosis and fibrosis were assessed. RESULTS: MCD diet resulted in steatohepatitis and increased miR-155 expression in total liver, hepatocytes and Kupffer cells. Steatosis and expression of genes involved in fatty acid metabolism were attenuated in miR-155 KO mice after MCD feeding. In contrast, miR-155 deficiency failed to attenuate inflammatory cell infiltration, nuclear factor kappa beta (NF-kappaB) activation and enhanced the expression of the pro-inflammatory cytokines tumor necrosis factor alpha (TNFalpha) and monocyte chemoattractant protein-1 (MCP1) in MCD diet-fed mice. We found a significant attenuation of apoptosis (cleaved caspase-3) and reduction in collagen and alpha smooth muscle actin (alphaSMA) levels in miR-155 KO mice compared to WTs on MCD diet. In addition, we found attenuation of platelet derived growth factor (PDGF), a pro-fibrotic cytokine; SMAD family member 3 (Smad3), a protein involved in transforming growth factor-beta (TGFbeta) signal transduction and vimentin, a mesenchymal marker and indirect indicator of epithelial-to-mesenchymal transition (EMT) in miR-155 KO mice. Nuclear binding of CCAAT enhancer binding protein beta (C/EBPbeta) a miR-155 target involved in EMT was significantly increased in miR-155 KO compared to WT mice. CONCLUSIONS: Our novel data demonstrate that miR-155 deficiency can reduce steatosis and fibrosis without decreasing inflammation in steatohepatitis