Many chromatin features play critical roles in regulating gene expression. A complete understanding of gene regulation will require the mapping of specific chromatin features in small samples of ...cells at high resolution. Here we describe Cleavage Under Targets and Tagmentation (CUT&Tag), an enzyme-tethering strategy that provides efficient high-resolution sequencing libraries for profiling diverse chromatin components. In CUT&Tag, a chromatin protein is bound in situ by a specific antibody, which then tethers a protein A-Tn5 transposase fusion protein. Activation of the transposase efficiently generates fragment libraries with high resolution and exceptionally low background. All steps from live cells to sequencing-ready libraries can be performed in a single tube on the benchtop or a microwell in a high-throughput pipeline, and the entire procedure can be performed in one day. We demonstrate the utility of CUT&Tag by profiling histone modifications, RNA Polymerase II and transcription factors on low cell numbers and single cells.
Methods for quantifying gene expression
and chromatin accessibility
in single cells are well established, but single-cell analysis of chromatin regions with specific histone modifications has been ...technically challenging. In this study, we adapted the CUT&Tag method
to scalable nanowell and droplet-based single-cell platforms to profile chromatin landscapes in single cells (scCUT&Tag) from complex tissues and during the differentiation of human embryonic stem cells. We focused on profiling polycomb group (PcG) silenced regions marked by histone H3 Lys27 trimethylation (H3K27me3) in single cells as an orthogonal approach to chromatin accessibility for identifying cell states. We show that scCUT&Tag profiling of H3K27me3 distinguishes cell types in human blood and allows the generation of cell-type-specific PcG landscapes from heterogeneous tissues. Furthermore, we used scCUT&Tag to profile H3K27me3 in a patient with a brain tumor before and after treatment, identifying cell types in the tumor microenvironment and heterogeneity in PcG activity in the primary sample and after treatment.
An inexpensive and disposable paper-based lateral flow strip (PLFS) has been developed as an immunoassay, in which surface-enhanced Raman scattering (SERS) is utilized for sensing signal ...transduction. The Au nanostar@Raman Reporter@silica sandwich nanoparticles are developed as the SERS probes, which is the key to the high sensitivity of the device. Compared with a colorimetric PLFS, the SERS-PLFS exhibits superior performance in terms of sensitivity and limit of detection (LOD) in a blood plasma-containing sample matrix. In addition, the SERS-PLFS has been successfully used for detection of neuron-specific enolase (NSE), a traumatic brain injury (TBI) protein biomarker, in diluted blood plasma samples, achieving a LOD of 0.86 ng/mL. Moreover, the SERS-PLFS was successfully employed to measure the NSE level in clinical blood plasma samples taken from deidentified TBI patients. This work demonstrates that the SERS-PLFS has great potential in assisting screening of TBI patients in the point-of-care setting.
Acute myeloid and lymphoid leukemias often harbor chromosomal translocations involving the KMT2A gene, encoding the KMT2A lysine methyltransferase (also known as mixed-lineage leukemia-1), and ...produce in-frame fusions of KMT2A to other chromatin-regulatory proteins. Here we map fusion-specific targets across the genome for diverse KMT2A oncofusion proteins in cell lines and patient samples. By modifying CUT&Tag chromatin profiling for full automation, we identify common and tumor-subtype-specific sites of aberrant chromatin regulation induced by KMT2A oncofusion proteins. A subset of KMT2A oncofusion-binding sites are marked by bivalent (H3K4me3 and H3K27me3) chromatin signatures, and single-cell CUT&Tag profiling reveals that these sites display cell-to-cell heterogeneity suggestive of lineage plasticity. In addition, we find that aberrant enrichment of H3K4me3 in gene bodies is sensitive to Menin inhibitors, demonstrating the utility of automated chromatin profiling for identifying therapeutic vulnerabilities. Thus, integration of automated and single-cell CUT&Tag can uncover epigenomic heterogeneity within patient samples and predict sensitivity to therapeutic agents.
The biosynthesis of the unique cyanobacterial (oxyphotobacterial) indole-phenolic UVA sunscreen, scytonemin, is coded for in a conserved operon that contains both core metabolic genes and accessory, ...aromatic amino acid biosynthesis genes dedicated to supplying scytonemin's precursors. Comparative genomics shows conservation of this operon in many, but not all, cyanobacterial lineages. Phylogenetic analyses of the operon's aromatic amino acid genes indicate that five of them were recruited into the operon after duplication events of their respective housekeeping cyanobacterial cognates. We combined the fossil record of cyanobacteria and relaxed molecular clock models to obtain multiple estimates of these duplication events, setting a minimum age for the evolutionary advent of scytonemin at 2.1 ± 0.3 billion years. The same analyses were used to estimate the advent of cyanobacteria as a group (and thus the appearance of oxygenic photosynthesis), at 3.6 ± 0.2 billion years before present.
interpretation of 16S rRNA-based Bayesian analyses was consistent with these estimates. Because of physiological constraints on the use of UVA sunscreens in general, and the biochemical constraints of scytonemin in particular, scytonemin's age must postdate the time when Earth's atmosphere turned oxic, known as the Great Oxidation Event (GOE). Indeed, our biological estimate is in agreement with independent geochemical estimates for the GOE. The difference between the estimated ages of oxygenic photosynthesis and the GOE indicates the long span (on the order of a billion years) of the era of "oxygen oases," when oxygen was available locally but not globally.
The advent of cyanobacteria, with their invention of oxygenic photosynthesis, and the Great Oxidation Event are arguably among the most important events in the evolutionary history of life on Earth. Oxygen is a significant toxicant to all life, but its accumulation in the atmosphere also enabled the successful development and proliferation of many aerobic organisms, especially metazoans. The currently favored dating of the Great Oxidation Event is based on the geochemical rock record. Similarly, the advent of cyanobacteria is also often drawn from the same estimates because in older rocks paleontological evidence is scarce or has been discredited. Efforts to obtain molecular evolutionary alternatives have offered widely divergent estimates. Our analyses provide a novel means to circumvent these limitations and allow us to estimate the large time gap between the two events.
This article discusses the current state of contract theory and its usefulness for conceptualizing issues related to agricultural contracting. The paper will explore the limitations of existing ...theory for applied work, and what methodological improvements are needed to enhance the usefulness of the theory to agricultural economists. One pervasive problem is that the economic literature on contracts is rather fragmented and the various methodological strands are narrow in their focus. As such, agricultural economists should engage in methodological research to develop applied contracting models that can capture higher-order features of real-world agricultural contracts while delivering generalizable comparative statics predictions because contracting continues to expand along the entire modern food marketing channel. In the latter part of this article, a simple model is developed to illustrate how classic methodological approaches can be combined with recent developments in contract and game theory to construct applied theory models that are useful for capturing some important features of agricultural contracts.
bstract: Since originally isolated in 1899, the genus
has been demonstrated to predominate in the gut microbiota of breastfed infants and to benefit the host by accelerating maturation of the immune ...response, balancing the immune system to suppress inflammation, improving intestinal barrier function, and increasing acetate production. In particular,
subspecies
(
) is well adapted to the infant gut and has co-evolved with the mother-infant dyad and gut microbiome, in part due to its ability to consume complex carbohydrates found in human milk.
and its human host have a symbiotic relationship that protects the preterm or term neonate and nourishes a healthy gut microbiota prior to weaning. To provide benefits associated with
to all infants, a number of commercialized strains have been developed over the past decades. As new ingredients become available, safety and suitability must be assessed in preclinical and clinical studies. Consideration of the full clinical evidence for
use in pediatric nutrition is critical to better understand its potential impacts on infant health and development. Herein we summarize the recent clinical studies utilizing select strains of commercialized
.
We previously reported that the use of small stakes assessments in an undergraduate/graduate physiology class (PHSL3061/5061, Principles of Physiology) contributed to improved average test scores for ...physiology and biomedical engineering majors and reduced the number of non‐passing physiology majors (EB 2021). Our overall hypothesis is that using small stakes assessments could improve outcomes for all students.
In this current study, we examined physiology majors from 2018‐2021 by race, gender, first generation status, class (sophomore, junior, senior), and indicated financial need. A typical physiology major taking PHSL3061/5061 tended to be white (60%), female (67%), non‐first generation student (75%), and in their senior year (60%).
When small stakes assessments were given, the number of students achieving below 70% was reduced as previously reported. However, those students that achieved a final score below 70% tended to be black, indigenous, person of color (BIPOC) (60%), female (80%), first generation student (75%), in their senior year (60%), and indicated some financial need (60%). Moreover, when term GPAs versus final scores were plotted, this group tended to have a term GPA between 3.0‐3.5/4.0, revealing an achievement gap in PHSL3061 relative to their predicted performance based on GPA.
We also compared academic performance (final score and term GPA at the end of the course) by race, gender, first generation, and class. The most significant differences in final score and term GPA were found between first generation and non‐first generation students.
Overall, our data revealed a group of students that are not achieving scores relative to their GPAs. We believe that these analyses can identify achievement gaps sooner. Taken together, the use of small stakes assessments can still lead to improved outcomes for physiology students. Further analyses of physiology students by category could reveal other potential areas where there are achievement gaps.
Research on milk fat globule membrane (MFGM) is gaining traction. The interest is two-fold; on the one hand, it is a unique trilayer structure with specific secretory function. On the other hand, it ...is the basis for ingredients with the presence of phospho- and sphingolipids and glycoproteins, which are being used as food ingredients with valuable functionality, in particular, for use as a supplement in infant nutrition. This last application is at the center of this Review, which aims to contribute to understanding MFGM's function in the proper development of immunity, cognition, and intestinal trophism, in addition to other potential effects such as prevention of diseases including cardiovascular disease, impaired bone turnover and inflammation, skin conditions, and infections as well as age-associated cognitive decline and muscle loss. The phospholipid composition of MFGM from bovine milk is quite like human milk and, although there are some differences due to dairy processing, these do not result in a chemical change. The MFGM ingredients, as used to improve the formulation in different clinical studies, have indeed increased the presence of phospholipids, sphingolipids, glycolipids, and glycoproteins with the resulting benefits of different outcomes (especially immune and cognitive outcomes) with no reported adverse effects. Nevertheless, the precise mechanism(s) of action of MFGM remain to be elucidated and further basic investigation is warranted.
To evaluate neurodevelopment, growth, and health outcomes in infants receiving bovine milk fat globule membrane (MFGM) and lactoferrin in infant formula.
Healthy term infants were randomized to a ...cow's milk-based infant formula or MFGM + LF (a similar infant formula, with an added source of bovine milk fat globule membrane bMFGM; whey protein-lipid concentrate, 5 g/L and bovine lactoferrin 0.6 g/L) through 365 days of age. The Bayley Scales of Infant Development, 3rd edition cognitive composite score at day 365 was the primary outcome. Secondary outcomes included tolerance measures through day 365, additional neurodevelopmental and language outcomes, growth, and medically confirmed adverse events through day 545.
Of 451 infants enrolled (control, 228; MFGM + LF, 223), 291 completed study feeding and Bayley-III testing at day 365 (control, 148; MFGM + LF, 143). The mean cognitive (+8.7), language (+12.3), and motor (+12.6) scores were higher (P < .001) for the MFGM + LF group; no differences were observed at day 545. Global development scores from day 120 to day 275 and attention at day 365 were significantly improved. Few group differences in day 545 neurodevelopmental outcomes were detected, however scores of some subcategories of the MacArthur-Bates Communicative Development Inventories were higher (P < .05) in the MFGM + LF group. The overall incidence of respiratory-associated adverse events and diarrhea were significantly lower for the MFGM + LF group through day 545.
Infants receiving formula with added bovine MFGM and bovine lactoferrin had an accelerated neurodevelopmental profile at day 365 and improved language subcategories at day 545. Formulas were associated with age-appropriate growth and significantly fewer diarrhea and respiratory-associated adverse events through 545 days of age.
NCT02274883.
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