Active eukaryotic regulatory sites are characterized by open chromatin, and yeast promoters and transcription factor binding sites (TFBSs) typically have low intrinsic nucleosome occupancy. Here, we ...show that in contrast to yeast, DNA at human promoters, enhancers, and TFBSs generally encodes high intrinsic nucleosome occupancy. In most cases we examined, these elements also have high experimentally measured nucleosome occupancy in vivo. These regions typically have high G+C content, which correlates positively with intrinsic nucleosome occupancy, and are depleted for nucleosome-excluding poly-A sequences. We propose that high nucleosome preference is directly encoded at regulatory sequences in the human genome to restrict access to regulatory information that will ultimately be utilized in only a subset of differentiated cells.
Previous studies in Saccharomyces cerevisiae established that depletion of histone H4 results in the genome-wide transcriptional de-repression of hundreds of genes. To probe the mechanism of this ...transcriptional de-repression, we depleted nucleosomes in vivo by conditional repression of histone H3 transcription. We then measured the resulting changes in transcription by RNA-seq and in chromatin organization by MNase-seq. This experiment also bears on the degree to which trans-acting factors and DNA-encoded elements affect nucleosome position and occupancy in vivo. We identified ∼60,000 nucleosomes genome wide, and we classified ∼2,000 as having preferentially reduced occupancy following H3 depletion and ∼350 as being preferentially retained. We found that the in vivo influence of DNA sequences that favor or disfavor nucleosome occupancy increases following histone H3 depletion, demonstrating that nucleosome density contributes to moderating the influence of DNA sequence on nucleosome formation in vivo. To identify factors important for influencing nucleosome occupancy and position, we compared our data to 40 existing whole-genome data sets. Factors associated with promoters, such as histone acetylation and H2A.z incorporation, were enriched at sites of nucleosome loss. Nucleosome retention was linked to stabilizing marks such as H3K36me2. Notably, the chromatin remodeler Isw2 was uniquely associated with retained occupancy and altered positioning, consistent with Isw2 stabilizing histone-DNA contacts and centering nucleosomes on available DNA in vivo. RNA-seq revealed a greater number of de-repressed genes (∼2,500) than previous studies, and these genes exhibited reduced nucleosome occupancy in their promoters. In summary, we identify factors likely to influence nucleosome stability under normal growth conditions and the specific genomic locations at which they act. We find that DNA-encoded nucleosome stability and chromatin composition dictate which nucleosomes will be lost under conditions of limiting histone protein and that this, in turn, governs which genes are susceptible to a loss of regulatory fidelity.
The sequence specificity of DNA-binding proteins is the primary mechanism by which the cell recognizes genomic features. Here, we describe systematic determination of yeast transcription factor ...DNA-binding specificities. We obtained binding specificities for 112 DNA-binding proteins representing 19 distinct structural classes. One-third of the binding specificities have not been previously reported. Several binding sequences have striking genomic distributions relative to transcription start sites, supporting their biological relevance and suggesting a role in promoter architecture. Among these are Rsc3 binding sequences, containing the core CGCG, which are found preferentially ∼100 bp upstream of transcription start sites. Mutation of
RSC3 results in a dramatic increase in nucleosome occupancy in hundreds of proximal promoters containing a Rsc3 binding element, but has little impact on promoters lacking Rsc3 binding sequences, indicating that Rsc3 plays a broad role in targeting nucleosome exclusion at yeast promoters.
Mg²⁺ shares a distinctive relationship with RNA, playing important and specific roles in the folding and function of essentially all large RNAs. Here we use theory and experiment to evaluate Fe²⁺ in ...the absence of free oxygen as a replacement for Mg²⁺ in RNA folding and catalysis. We describe both quantum mechanical calculations and experiments that suggest that the roles of Mg²⁺ in RNA folding and function can indeed be served by Fe²⁺. The results of quantum mechanical calculations show that the geometry of coordination of Fe²⁺ by RNA phosphates is similar to that of Mg²⁺. Chemical footprinting experiments suggest that the conformation of the Tetrahymena thermophila Group I intron P4-P6 domain RNA is conserved between complexes with Fe²⁺ or Mg²⁺. The catalytic activities of both the L1 ribozyme ligase, obtained previously by in vitro selection in the presence of Mg²⁺, and the hammerhead ribozyme are enhanced in the presence of Fe²⁺ compared to Mg²⁺. All chemical footprinting and ribozyme assays in the presence of Fe²⁺ were performed under anaerobic conditions. The primary motivation of this work is to understand RNA in plausible early earth conditions. Life originated during the early Archean Eon, characterized by a non-oxidative atmosphere and abundant soluble Fe²⁺. The combined biochemical and paleogeological data are consistent with a role for Fe²⁺ in an RNA World. RNA and Fe²⁺ could, in principle, support an array of RNA structures and catalytic functions more diverse than RNA with Mg²⁺ alone.
Satellite tobacco mosaic virus (STMV) is a T = 1 icosahedral virus with a single-stranded RNA genome. It is widely accepted that the RNA genome plays an important structural role during assembly of ...the STMV virion. While the encapsidated form of the RNA has been extensively studied, less is known about the structure of the free RNA, aside from a purported tRNA-like structure at the 3' end. Here we use selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) analysis to examine the secondary structure of in vitro transcribed STMV RNA. The predicted secondary structure is unusual in the sense that it is highly extended, which could be significant for protecting the RNA from degradation. The SHAPE data are also consistent with the previously predicted tRNA-like fold at the 3' end of the molecule, which is also known to hinder degradation. Our data are not consistent with the secondary structure proposed for the encapsidated RNA by Schroeder et al., suggesting that, if the Schroeder structure is correct, either the RNA is packaged as it emerges from the replication complex, or the RNA undergoes extensive refolding upon encapsidation. We also consider the alternative, i.e., that the structure of the encapsidated STMV RNA might be the same as the in vitro structure presented here, and we examine how this structure might be organized in the virus. This possibility is not rigorously ruled out by the available data, so it remains open to examination by experiment.
Background
To evaluate the landscape of early extubation, and identify factors associated with early extubation (≤ 24 h) after superior cavopulmonary connection (stage 2 operation) among children ...with single ventricle anatomy.
Methods
Patients undergoing stage 2 operation after Norwood operation from the Pediatric Heart Network Single Ventricle Reconstruction (SVR) trial public‐use dataset were included. Elastic net regularized logistic regression models were fitted to evaluate the factors associated with early extubation after stage 2 operation.
Results
In total, 390 patients from 15 North American centers qualified for inclusion. Of these, 42 patients (10.8%) were extubated in operating room, 151 patients (38.7%) were extubated outside the operating room within the first 24 h after stage 2 operation, and the remaining 197 patients (50.5%) required mechanical ventilation for > 24 h. In adjusted models, factors associated with early extubation after stage 2 operation were elective timing of stage 2 operation, lower incidence of post‐Norwood complications, shorter CPB duration for stage 2 operation, and no cardiac catheterization after Stage 2 operation. We also performed multiple other alternative analyses to identify factors associated with early extubation that demonstrated same associations as the primary model. The mean hospital length of stay after Stage 2 operation was 20% shorter among patients with early extubation.
Conclusions
Data from this large multicenter study demonstrate that approximately one‐half of the patients undergoing operation for superior cavopulmonary connection are extubated within 24 h after heart operation. Furthermore, early extubation is associated with shorter hospital length of stay.
The objective of this study was to determine the evolution of obesity status (OS) in a longitudinal cohort of low birth weight preterm (LBWPT) infants to an age of 8 years, and to determine whether ...rapid weight gain in the first year of life independently predicts 8-year OS.
In total, 985 infants (birth weight ≤2500 g, gestation age ≤37 weeks) were recruited from the nursery in an eight-site intervention research program and were evaluated at an age of 3, 5, 6.5 and 8 years. Weight and height were measured by standard protocol at each visit and body mass index was calculated. Obesity status is ≥95% for age and sex. Multiple logistic analyses were performed on 8-year OS with predictor variables including infant race, gender, small for gestational age status, birth weight category, neonatal health index, treatment group and first-year weight gain; maternal education and weight status before conception; and HOME Inventory.
Overall, 2.3% were OS at an age of 3 years, 6.1% at an age of 5 years, 7.7% at age 6.5 years and 8.7% at an age 8 years. OS varied by birth weight category at each visit. The infants born ≤1500 g had the lowest prevalence of OS at each age. In the logistic regression, maternal race (Hispanic) (adjusted odds ratio=2.8, confidence interval=1.2 to 6.8), maternal obese status (adjusted odds ratio 3.4, confidence interval=1.5 to 7.8) and first-year weight gain (adjusted odds ratio=2.7, confidence interval=1.9 to 3.9), significantly predicted 8-year OS.
OS is common in LBWPT infants during childhood, and prevalence varies by birth weight category. High weight gain in the first year of life is an important predictor of the development of OS in LBWPT children.
Tetrachloroethene is a prominent groundwater pollutant that can be reductively dechlorinated by mixed anaerobic microbial populations to the nontoxic product ethene. Strain 195, a coccoid bacterium ...that dechlorinates tetrachloroethene to ethene, was isolated and characterized. Growth of strain 195 with H2 and tetrachloroethene as the electron donor and acceptor pair required extracts from mixed microbial cultures. Growth of strain 195 was resistant to ampicillin and vancomycin; its cell wall did not react with a peptidoglycan-specific lectin and its ultrastructure resembled S-layers of Archaea. Analysis of the 16S ribosomal DNA sequence of strain 195 indicated that it is a eubacterium without close affiliation to any known groups
DNA can be used as a structural component in the process of making conductive polymers called nanowires. Accurate molecular models could lead to a better understanding of how to prepare these types ...of materials. Here we present a computational tool that allows potential DNA-linked polymer designs to be screened and evaluated. The approach involves an iterative procedure that adjusts the positions of DNA-linked monomers in order to obtain reasonable molecular geometry compatible with normal DNA conformations and with the properties of the polymer being formed. This procedure has been used to evaluate designs already reported experimentally, as well as to suggest a new design based on pyrrylene vinylene (PV) monomers.
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