Genome-wide fetalization of enhancer architecture in heart disease.

Heart disease is associated with re-expression of key transcription factors normally active only during prenatal development of the heart. However, the impact of this reactivation on the regulatory landscape in heart disease is unclear. Here, we use RNA-seq and ChIP-seq targeting a histone modification associated with active transcriptional enhancers to generate genome-wide enhancer maps from left ventricle tissue from up to 26 healthy controls, 18 individuals with idiopathic dilated cardiomyopathy (DCM), and five fetal hearts. Healthy individuals have a highly reproducible epigenomic landscape, consisting of more than 33,000 predicted heart enhancers. In contrast, we observe reproducible disease-associated changes in activity at 6,850 predicted heart enhancers. Combined analysis of adult and fetal samples reveals that the heart disease epigenome and transcriptome both acquire fetal-like characteristics, with 3,400 individual enhancers sharing fetal regulatory properties. We also provide a comprehensive data resource (http://heart.lbl.gov) for the mechanistic exploration of DCM etiology.

Hyperspectral time series datasets of maize during the grain filling period.

OBJECTIVES: Remotely sensed hyperspectral data are increasingly being used to assess crop development and growth throughout the growing season. Large datasets capturing key growth stages can be useful to researchers studying many physiological plant responses. A time series analysis of hyperspectral reflectance measurements taken during the grain filling period and published within a publicly accessible database are described herein. These datasets document the spectral reflectance pattern of the canopy within the visible and near-infrared portion of the electromagnetic spectrum during the late stages of the grain filling period as plants approach and reach physiological maturity. DATA DESCRIPTION: Included within the data repository are canopy-level hyperspectral datasets collected in 2017 and 2018. Data is included in its raw form, as well as with several manipulations to smooth and standardize the raw data. Data are released as comma separated value spreadsheets as well as Microsoft Excel open XLSX spreadsheets. These are accompanied by README text files which further describe the data and supplemental files that record hybrids used and plant phenology for each year of data collection.

Coaching Strategies Used to Support Interprofessional Teams in 3 Primary Care Centers.

PURPOSE/AIMS: Clinical nurse specialists and other advanced practice registered nurses use healthcare team coaching to foster interprofessional practice and enhance healthcare quality. Although coaching has been shown to support positive changes in healthcare, little is known about how coaching strategies are used in practice. The purpose of this study was to describe the strategies used by an experienced healthcare team coach tasked with advancing interprofessional care and teamwork in primary care clinics. METHODS: This qualitative description study was part of a larger project that included an objective to increase interprofessional practice in 3 primary care clinics in the midwestern United States. Data drawn from 35 audio-recorded and transcribed coaching telephone calls were analyzed using content analysis. RESULTS: Twelve coaching strategies were identified and divided into the following groups: (a) enhancing team development, (b) affirming the work of the team, (c) facilitating progress, (d) providing resources, and (e) connecting work to theoretical frameworks. CONCLUSIONS: The coaching strategies described in this study can inform the work of clinical nurse specialists and other advanced practice registered nurses charged with advancing interprofessional collaborative practice. Future research is recommended to examine the efficacy of strategies and develop a comprehensive model of healthcare team coaching.

Author Correction: An atlas of dynamic chromatin landscapes in mouse fetal development.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Supervised enhancer prediction with epigenetic pattern recognition and targeted validation.

Enhancers are important non-coding elements, but they have traditionally been hard to characterize experimentally. The development of massively parallel assays allows the characterization of large numbers of enhancers for the first time. Here, we developed a framework using Drosophila STARR-seq to create shape-matching filters based on meta-profiles of epigenetic features. We integrated these features with supervised machine-learning algorithms to predict enhancers. We further demonstrated that our model could be transferred to predict enhancers in mammals. We comprehensively validated the predictions using a combination of in vivo and in vitro approaches, involving transgenic assays in mice and transduction-based reporter assays in human cell lines (153 enhancers in total). The results confirmed that our model can accurately predict enhancers in different species without re-parameterization. Finally, we examined the transcription factor binding patterns at predicted enhancers versus promoters. We demonstrated that these patterns enable the construction of a secondary model that effectively distinguishes enhancers and promoters.

An atlas of dynamic chromatin landscapes in mouse fetal development.

The Encyclopedia of DNA Elements (ENCODE) project has established a genomic resource for mammalian development, profiling a diverse panel of mouse tissues at 8 developmental stages from 10.5 days after conception until birth, including transcriptomes, methylomes and chromatin states. Here we systematically examined the state and accessibility of chromatin in the developing mouse fetus. In total we performed 1,128 chromatin immunoprecipitation with sequencing (ChIP-seq) assays for histone modifications and 132 assay for transposase-accessible chromatin using sequencing (ATAC-seq) assays for chromatin accessibility across 72 distinct tissue-stages. We used integrative analysis to develop a unified set of chromatin state annotations, infer the identities of dynamic enhancers and key transcriptional regulators, and characterize the relationship between chromatin state and accessibility during developmental gene regulation. We also leveraged these data to link enhancers to putative target genes and demonstrate tissue-specific enrichments of sequence variants associated with disease in humans. The mouse ENCODE data sets provide a compendium of resources for biomedical researchers and achieve, to our knowledge, the most comprehensive view of chromatin dynamics during mammalian fetal development to date.

Pharmacokinetic Parameters and Estimated Milk Withdrawal Intervals for Domestic Goats (Capra Aegagrus Hircus) After Administration of Single and Multiple Intravenous and Subcutaneous Doses of Flunixin Meglumine.

Introduction: The study objectives were to estimate plasma flunixin (FLU) pharmacokinetic parameters and milk depletion profiles for FLU and its metabolite (5-hydroxy flunixin; 5-OH) after subcutaneous (SC) and intravenous (IV) administration of single and multiple flunixin meglumine (FM) doses to non-lactating (nulliparous and pregnant does) and lactating dairy goats. Analytical methods (ELISA and UPLC-MS/MS) for quantifying plasma FLU concentrations were compared. The final objective was to use regulatory (FDA and EMA) methods to estimate milk withdrawal intervals following extra-label drug use in goats. Methods: FM was administered IV and SC to commercial dairy goats at 1.1 mg/kg for single and multiple doses. Plasma and milk samples were analyzed for FLU and 5-OH via UPLC-MS/MS. Plasma samples were also analyzed for FLU concentrations via ELISA. Using statistical approaches recommended by regulatory agencies, milk withdrawal intervals were estimated following FM extra-label use. Results: Following IV administration of a single FM dose, clearances were 127, 199, and 365 ml/kg/h for non-lactating (NL) pregnant does, NL nulliparous does, and lactating dairy does, respectively. Following multiple SC doses, clearance/F was 199 ml/kg/h for lactating does. After IV administration of a single FM dose, terminal elimination half-lives were 4.08, 2.87, and 3.77 h for NL pregnant does, NL nulliparous does, and lactating dairy does, respectively. After multiple SC doses, the terminal elimination half-life was 3.03 h for lactating dairy does. No significant differences were noted for samples analyzed by UPLC-MS/MS or ELISA. Milk withdrawal intervals ranged from 36 to 60 h depending on the regulatory statistical method and dosage regimen. Conclusions: Subcutaneous administration of FM to goats results in similar plasma pharmacokinetic parameters as IV administration. ELISA analysis is an alternative method to UPLC-MS/MS for quantifying FLU concentrations in caprine plasma samples. Following FM extra-label administration to dairy goats, clinicians could consider 36-60 h milk withdrawal intervals.

Clinical features and radiographic findings in cats with eosinophilic, neutrophilic, and mixed airway inflammation (2011-2018).

BACKGROUND: Idiopathic inflammatory airway disease (IAD) in cats often is described as asthmatic (eosinophilic) or bronchitic (neutrophilic), but this designation requires collection of airway fluid and it fails to consider cats with mixed airway inflammation. OBJECTIVE: To identify clinical features that would differentiate inflammatory disease types. ANIMALS: Forty-nine cats with nonspecific airway inflammation identified by bronchoscopic bronchoalveolar lavage (BAL) between 2011 and 2018 were evaluated. METHODS: This is a retrospective study. Cats were categorized by BAL differential cytology as having eosinophilic (eosinophils >20% with neutrophils <14%, or eosinophils >50%), mixed (eosinophils 20%-50% and neutrophils >14% or discordant inflammation from 2 BAL sites), or neutrophilic (neutrophils >14% and eosinophils <20%) inflammation. Type and duration of presenting complaints, signalment, body condition score, respiratory rate, CBC results, bronchoscopy, BAL results (% recovery, total nucleated cell count, differential cell count), and radiographic findings were compared among groups. RESULTS: Idiopathic IAD was diagnosed in 49 cats, with BAL eosinophilic inflammation in 23, mixed inflammation in 14, and neutrophilic inflammation in 12. Cough was the predominant presenting complaint with no difference in duration of signs among groups (median, 5.5 months). Respiratory rate and effort also did not differ. Cats with eosinophilic inflammation were significantly younger (4.4 ± 3.3 years) than those with neutrophilic (8.0 ±5.6 years) or mixed inflammation (7.5 ± 4.0 years; P = .03). Results of CBC and interpretation of radiographic findings did not differ among groups. CONCLUSIONS AND CLINICAL IMPORTANCE: Substantial overlap exists in clinical and radiographic findings in cats with various forms of idiopathic airway inflammation.

Identification of a polymorphism within the Rosa multiflora muRdr1A gene linked to resistance to multiple races of Diplocarpon rosae W. in tetraploid garden roses (Rosa × hybrida).

KEY MESSAGE: A QTL for resistance to several races of black spot co-located with the known Rrd1 locus in Rosa. A polymorphism in muRdr1A linked to black spot resistance was identified and molecular markers were designed. Black spot, caused by Diplocarpon rosae, is one of the most serious foliar diseases of landscape roses that reduces the marketability and weakens the plants against winter survival. Genetic resistance to black spot (BS) exists and race-specific resistance is a good target to implement marker-assisted selection. High-density single nucleotide polymorphism-based genetic maps were created for the female parent of a tetraploid cross between 'CA60' and 'Singing in the Rain' using genotyping-by-sequencing following a two-way pseudo-testcross strategy. The female linkage map was generated based on 227 individuals and included 31 linkage groups, 1055 markers, with a length of 1980 cM. Race-specific resistance to four D. rosae races (5, 7, 10, 14) was evaluated using a detached leaf assay. BS resistance was also evaluated under natural infection in the field. Resistance to races 5, 10 and 14 of D. rosae and field resistance co-located on chromosome 1. A unique sequence of 32 bp in exon 4 of the muRdr1A gene was identified in 'CA60' that co-segregates with D. rosae resistance. Two diagnostic markers, a presence/absence marker and an INDEL marker, specific to this sequence were designed and validated in the mapping population and a backcross population derived from 'CA60.' Resistance to D. rosae race 7 mapped to a different location on chromosome 1.

Enhancer redundancy provides phenotypic robustness in mammalian development.

Distant-acting tissue-specific enhancers, which regulate gene expression, vastly outnumber protein-coding genes in mammalian genomes, but the functional importance of this regulatory complexity remains unclear. Here we show that the pervasive presence of multiple enhancers with similar activities near the same gene confers phenotypic robustness to loss-of-function mutations in individual enhancers. We used genome editing to create 23 mouse deletion lines and inter-crosses, including both single and combinatorial enhancer deletions at seven distinct loci required for limb development. Unexpectedly, none of the ten deletions of individual enhancers caused noticeable changes in limb morphology. By contrast, the removal of pairs of limb enhancers near the same gene resulted in discernible phenotypes, indicating that enhancers function redundantly in establishing normal morphology. In a genetic background sensitized by reduced baseline expression of the target gene, even single enhancer deletions caused limb abnormalities, suggesting that functional redundancy is conferred by additive effects of enhancers on gene expression levels. A genome-wide analysis integrating epigenomic and transcriptomic data from 29 developmental mouse tissues revealed that mammalian genes are very commonly associated with multiple enhancers that have similar spatiotemporal activity. Systematic exploration of three representative developmental structures (limb, brain and heart) uncovered more than one thousand cases in which five or more enhancers with redundant activity patterns were found near the same gene. Together, our data indicate that enhancer redundancy is a remarkably widespread feature of mammalian genomes that provides an effective regulatory buffer to prevent deleterious phenotypic consequences upon the loss of individual enhancers.

Ultraconserved Enhancers Are Required for Normal Development.

Non-coding "ultraconserved" regions containing hundreds of consecutive bases of perfect sequence conservation across mammalian genomes can function as distant-acting enhancers. However, initial deletion studies in mice revealed that loss of such extraordinarily constrained sequences had no immediate impact on viability. Here, we show that ultraconserved enhancers are required for normal development. Focusing on some of the longest ultraconserved sites genome wide, located near the essential neuronal transcription factor Arx, we used genome editing to create an expanded series of knockout mice lacking individual or combinations of ultraconserved enhancers. Mice with single or pairwise deletions of ultraconserved enhancers were viable and fertile but in nearly all cases showed neurological or growth abnormalities, including substantial alterations of neuron populations and structural brain defects. Our results demonstrate the functional importance of ultraconserved enhancers and indicate that remarkably strong sequence conservation likely results from fitness deficits that appear subtle in a laboratory setting.

Progressive Loss of Function in a Limb Enhancer during Snake Evolution.

The evolution of body shape is thought to be tightly coupled to changes in regulatory sequences, but specific molecular events associated with major morphological transitions in vertebrates have remained elusive. We identified snake-specific sequence changes within an otherwise highly conserved long-range limb enhancer of Sonic hedgehog (Shh). Transgenic mouse reporter assays revealed that the in vivo activity pattern of the enhancer is conserved across a wide range of vertebrates, including fish, but not in snakes. Genomic substitution of the mouse enhancer with its human or fish ortholog results in normal limb development. In contrast, replacement with snake orthologs caused severe limb reduction. Synthetic restoration of a single transcription factor binding site lost in the snake lineage reinstated full in vivo function to the snake enhancer. Our results demonstrate changes in a regulatory sequence associated with a major body plan transition and highlight the role of enhancers in morphological evolution. PAPERCLIP.

Changes in light quality alter physiological responses of soybean to thiamethoxam.

MAIN CONCLUSION: The interaction between neighboring weed-induced far-red enriched light and thiamethoxam can significantly alter soybean seedling morphology, nodulation, isoflavone levels, UV-absorbing phenolics, and carbon and nitrogen content. Neonicotinoid insecticides that are widely used on major crop plants can enhance plant growth and yield. Although the underlying mechanism of this enhanced growth and yield is not clear, recent studies suggest that neonicotinoids such as thiamethoxam (TMX) may exert their effects at least in part via signals that involve salicylic acid (SA) and jasmonic acid (JA). In the current research, effects of TMX on morphological and physiological responses of soybean have been compared under far-red-depleted (FR-D) and far-red-enriched (FR-E) light reflected by neighboring weeds. TMX significantly enhanced shoot and root growth but did not prevent stem elongation under FR-E light. Also, TMX did not prevent reductions in shoot carbon content and shoot carbon to nitrogen ratio under FR-E light. Despite similarities between these TMX effects in soybean and those known for SA and JA in other plant species, TMX significantly enhanced root-nodule numbers per plant and levels of root isoflavones malonyl-daidzin and malonyl-genistin under FR-E light only. These results suggest that the combined effect of FR-E light and TMX triggers a mechanism that operates concomitantly to enhance root isoflavones and nodulation in soybean.

Convergence in feeling, divergence in physiology: How culture influences the consequences of disgust suppression and amplification among European Americans and Asian Americans.

Much empirical work documents the downsides of suppressing emotions. Emerging research points to the need for a more sophisticated and culturally informed approach to understanding the consequences of emotion regulation. To that end, we employed behavioral, self-report, and psychophysiological measures to examine the consequences of two types of emotion regulation (suppression and amplification) in a sample of 28 Asian Americans and 31 European Americans. Participants were shown a neutral film and then a series of disgust-eliciting films during which they were asked to regulate their response by suppressing or amplifying their emotional behavior (counterbalanced). Despite self-reporting equal levels of disgust, European Americans showed greater skin conductance reactivity than Asian Americans in both regulation conditions, but not in response to a neutral film. These findings extend work on divergence in the consequences of emotion regulation across different cultural groups, which could help identify optimal emotion regulation strategies for health and well-being.

Multimeric states of starch phosphorylase determine protein-protein interactions with starch biosynthetic enzymes in amyloplasts.

Protein-protein interactions between starch phosphorylase (SP) and other starch biosynthetic enzymes were investigated using isolated maize endosperm amyloplasts and a recombinant maize enzyme. Plastidial SP is a stromal enzyme existing as a multimeric protein in amyloplasts. Biochemical analysis of the recombinant maize SP indicated that the tetrameric form was catalytically active in both glucan-synthetic and phosphorolytic directions. Protein-protein interaction experiments employing the recombinant SP as an affinity ligand with amyloplast extracts showed that the multimeric state of SP determined interactions with other enzymes of the starch biosynthetic pathway. The monomeric form of SP interacts with starch branching enzyme I (SBEI) and SBEIIb, whereas only SBEI interacts with the tetrameric form of SP. In all cases, protein-protein interactions were broken when amyloplast lysates were dephosphorylated in vitro, and enhanced following pre-treatment with ATP, suggesting a mechanism of protein complex formation regulated by protein phosphorylation. In vitro protein phosphorylation experiments with [γ-(32)P]-ATP show that SP is phosphorylated by a plastidial protein kinase. Evidence is presented which suggests SBEIIb modulates the catalytic activity of SP through the formation of a heteromeric protein complex.

Preliminary findings of the relationship of lower heart rate variability with military sexual trauma and presumed posttraumatic stress disorder.

Decreased heart rate variability (HRV) occurs with physical and psychological disorders and is a predictor of cardiac and all-cause mortality. This study was the first of which we are aware to examine and report the relationship between military sexual trauma (MST) and HRV measures. In a historical cohort study of female veterans with (n = 27) and without (n = 99) MST who received Holter and electrocardiogram evaluation at a Veteran Affairs medical center during 2007-2010, we examined the relationship between MST and the standard deviation of all R-R intervals (SDNN) and the square root of the mean of the sum of the squares of differences between adjacent R-R intervals (RMSSD). Female veterans with MST were younger, p = .002, frequently had a probable posttraumatic stress disorder diagnosis, 80% versus 15%, p = < .0001, and had lower SDNN, p = .0001, and RMSSD, p = .001, than those without MST. The SDNN and RMSSD of a 25-year-old female veteran with MST were comparable to that of female veterans aged 69 to 81 years without MST. Further research is needed to evaluate relationships between MST and HRV measures.

Characteristics of emergency pages using a computer-based anesthesiology paging system in children and adults undergoing procedures at a tertiary care medical center.

BACKGROUND: In our large academic supervisory practice, attending anesthesiologists concomitantly care for multiple patients. To manage communications within the procedural environment, we use a proprietary electronic computer-based anesthesiology visual paging system. This system can send an emergency page that instantly alerts the attending anesthesiologist and other available personnel that immediate help is needed. We analyzed the characteristics of intraoperative emergency pages in children and adults. METHODS: We identified all emergency page activations between January 1, 2005 and July 31, 2010 in our main operating rooms. Electronic medical records were reviewed for rates and characteristics of pages such as primary etiology, performed interventions, and outcomes. RESULTS: During the study period, 258,135 anesthetics were performed (n = 32,103 children, younger than 18 years) and 370 emergency pages (n = 309 adults, n = 61 children) were recorded (1.4 per 1000 cases; 95% confidence interval, 1.3-1.6). Infants had the highest rates (9.4 per 1000; 95% confidence interval, 5.7-14.4) of emergency page activations (P < 0.001 compared with each other age group). In adults, the most frequent causes were hemodynamic (55%), and in children respiratory and airway (60.7%) events. CONCLUSION: Emergency pages were rare in patients older than 2 years. Infants were more likely than children 1 to 2 years of age to have emergency page activation, despite both groups being cared for by pediatric fellowship trained anesthesiologists.

Glucan affinity of starch synthase IIa determines binding of starch synthase I and starch-branching enzyme IIb to starch granules.

The sugary-2 mutation in maize (Zea mays L.) is a result of the loss of catalytic activity of the endosperm-specific SS (starch synthase) IIa isoform causing major alterations to amylopectin architecture. The present study reports a biochemical and molecular analysis of an allelic variant of the sugary-2 mutation expressing a catalytically inactive form of SSIIa and sheds new light on its central role in protein-protein interactions and determination of the starch granule proteome. The mutant SSIIa revealed two amino acid substitutions, one being a highly conserved residue (Gly522→Arg) responsible for the loss of catalytic activity and the inability of the mutant SSIIa to bind to starch. Analysis of protein-protein interactions in sugary-2 amyloplasts revealed the same trimeric assembly of soluble SSI, SSIIa and SBE (starch-branching enzyme) IIb found in wild-type amyloplasts, but with greatly reduced activities of SSI and SBEIIb. Chemical cross-linking studies demonstrated that SSIIa is at the core of the complex, interacting with SSI and SBEIIb, which do not interact directly with each other. The sugary-2 mutant starch granules were devoid of amylopectin-synthesizing enzymes, despite the fact that the respective affinities of SSI and SBEIIb from sugary-2 for amylopectin were the same as observed in wild-type. The data support a model whereby granule-bound proteins involved in amylopectin synthesis are partitioned into the starch granule as a result of their association within protein complexes, and that SSIIa plays a crucial role in trafficking SSI and SBEIIb into the granule matrix.

Phenotypic plasticity and integration in the mangrove rivulus (Kryptolebias marmoratus): a prospectus.

The mangrove rivulus (Kryptolebias marmoratus) is a small fish native to mangrove ecosystems in Florida, the Caribbean, Central America, and South America. This species is one of only two self-fertilizing, hermaphroditic vertebrates capable of producing offspring that are genetically identical to both the parent and all siblings. Long bouts of selfing result in individuals with completely homozygous genotypes, effectively allowing for the production of "clones." Rivulus is also extremely sensitive to environmental change, both during development and adulthood. Life-history traits, behavior, physiology, morphology, and even sexual phenotype are shaped to a large extent by the interaction of genes with the environment, and many of these traits appear to co-vary. True reaction norms can be generated for this species in much the same way as has been done for clonally reproducing invertebrates and plants that have contributed immensely to our understanding of the evolution of phenotypic plasticity. That is, rivulus provides the opportunity to place individuals with identical genotypes in many different environments at any point during ontogeny or adulthood. In addition, rivulus populations are characterized by high genotypic diversity, a luxury not afforded by many clonal vertebrates, which allows us to evaluate variation among genotypes in the shape of reaction norms and in patterns of covariance among traits. We provide background information on phenotypic plasticity and phenotypic integration, coupled with a description of characteristics that we feel qualify rivulus as a potentially powerful model in which to study the evolution of reaction norms and covariance among traits.