As the Drosophila embryo transitions from the use of maternal RNAs to zygotic transcription, domains of open chromatin, with relatively low nucleosome density and specific histone marks, are ...established at promoters and enhancers involved in patterned embryonic transcription. However it remains unclear how regions of activity are established during early embryogenesis, and if they are the product of spatially restricted or ubiquitous processes. To shed light on this question, we probed chromatin accessibility across the anterior-posterior axis (A-P) of early Drosophila melanogaster embryos by applying a transposon based assay for chromatin accessibility (ATAC-seq) to anterior and posterior halves of hand-dissected, cellular blastoderm embryos. We find that genome-wide chromatin accessibility is highly similar between the two halves, with regions that manifest significant accessibility in one half of the embryo almost always accessible in the other half, even for promoters that are active in exclusively one half of the embryo. These data support previous studies that show that chromatin accessibility is not a direct result of activity, and point to a role for ubiquitous factors or processes in establishing chromatin accessibility at promoters in the early embryo. However, in concordance with similar works, we find that at enhancers active exclusively in one half of the embryo, we observe a significant skew towards greater accessibility in the region of their activity, highlighting the role of patterning factors such as Bicoid in this process.
Background: Exposure to metalworking fluids has been previously associated with prostate cancer mortality in a cohort of autoworkers. Our objective was to further explore this finding in a study of ...prostate cancer incidence in the same cohort, with reduced misclassification of outcome. Methods: We conducted a nested case-control study in the General Motors cohort of autoworkers. Incident cases of prostate cancer (n = 872) were identified via the Michigan Cancer Registry from 1985 through 2000. Controls were selected using incidence-density sampling with 5:1 ratio. Using cumulative exposure (mg/m³-years) as the dose metric, we first examined varying lengths of lags (0-25 years). Then, we evaluated consecutive windows of exposure: 25 or more years before risk age, and fewer than 25 years. We used penalized splines to model the relative risk as a smooth function of exposure, and adjusted for race and calendar year of diagnosis in a Cox model. Results: Risk of prostate cancer increased with exposure to soluble and straight fluids 25 years or more before risk age but not with exposure in the last 25 years. The relationship with soluble fluids was piecewise linear, with a small increase in risk at lower exposures followed by a steeper rise. By contrast, the relationship with straight fluids was linear, with a relative risk of 1.12 per 10 mg/m³--years of exposure (95% confidence interval = 1.04-1.20). Conclusions: Exposure to oil-based fluids, soluble and straight, is modestly associated with prostate cancer risk among autoworkers, with a latency period of at least 25 years.
Remarkable advances in DNA sequencing technology have created a need for de novo genome assembly methods tailored to work with the new sequencing data types. Many such methods have been published in ...recent years, but assembling raw sequence data to obtain a draft genome has remained a complex, multi-step process, involving several stages of sequence data cleaning, error correction, assembly, and quality control. Successful application of these steps usually requires intimate knowledge of a diverse set of algorithms and software. We present an assembly pipeline called A5 (Andrew And Aaron's Awesome Assembly pipeline) that simplifies the entire genome assembly process by automating these stages, by integrating several previously published algorithms with new algorithms for quality control and automated assembly parameter selection. We demonstrate that A5 can produce assemblies of quality comparable to a leading assembly algorithm, SOAPdenovo, without any prior knowledge of the particular genome being assembled and without the extensive parameter tuning required by the other assembly algorithm. In particular, the assemblies produced by A5 exhibit 50% or more reduction in broken protein coding sequences relative to SOAPdenovo assemblies. The A5 pipeline can also assemble Illumina sequence data from libraries constructed by the Nextera (transposon-catalyzed) protocol, which have markedly different characteristics to mechanically sheared libraries. Finally, A5 has modest compute requirements, and can assemble a typical bacterial genome on current desktop or laptop computer hardware in under two hours, depending on depth of coverage.
We have purified and determined the complete primary structure of human stromelysin, a secreted metalloprotease with a wide range of substrate specificities. Human stromelysin is synthesized in a ...preproenzyme form with a calculated size of 53,977 Da and a 17-amino acid long signal peptide. Prostromelysin is secreted in two forms, with apparent molecular masses on NaDodSO4/PAGE of 60 and 57 kDa. The minor 60-kDa polypeptide is a glycosylated form of the major 57-kDa protein containing N-linked complex oligosaccharides. Zymogen activation by trypsin results in the removal of 84 amino acids from the amino terminus of the enzyme generating a 45-kDa active enzyme species. Human stromelysin is capable of degrading proteoglycan, fibronectin, laminin, and type IV collagen but not interstitial type I collagen. The enzyme is not capable of activating purified human fibroblast procollagenase. Analysis of its primary structure shows that stromelysin is in all likelihood the human analog of rat transin, which is an oncogene transformation-induced protease. The pattern of enzyme expression in normal and tumorigenic cells revealed that human skin fibroblasts in vitro secrete stromelysin constitutively (1-2 μ g per 106 cells per 24 hr). Human fetal lung fibroblasts transformed with simian virus 40, human bronchial epithelial cells transformed with the ras oncogene, fibrosarcoma cells (HT-1080), and a melanoma cell strain (A 2058), do not express this protease nor can the enzyme be induced in these cells by treatment with phorbol 12-myristate 13-acetate. Our data indicate that the expression and the possible involvement of secreted metalloproteases in tumorigenesis result from a specific interaction between the transforming factor and the target cell, which may vary in different species.
Objective: Identify most likely health effects of occupational exposure to engineered nanoparticles (ENP). Recommend analytic approaches to address epidemiologic challenges. Methods: Review air ...pollution and occupational literature on health effects of fine particulate matter (PM). Provide example of mortality study of exposure to PM composed of metalworking fluid. Apply standard Cox models and g-estimation to adjust for potential healthy worker survival effect (HWSE). Results: In contrast with standard methods, g-estimation suggests that exposure to PM may cause chronic heart and lung disease; longer exposure reduces survival. HWSE appears stronger for chronic disease than for cancer. Conclusions: We recommend hazard surveillance, short-term panel studies of biomarkers, and prospective cohort studies of cardiovascular and respiratory diseases. Building research capacity in g-estimation methods to reduce HWSE is necessary for future studies of chronic disease and ENP.
The C57BL/6 (B6) mouse is more sensitive to the effects of a high-fat diet than the A/J strain. The B6 mouse develops severe obesity, hyperglycemia, and hyperinsulinemia when fed this dietary ...regimen. This study was conducted to determine the effects of dietary fat and sucrose concentrations on body composition and intestinal sucrase (EC 3.2.1.48) and maltase (EC 3.2.1.20) activity in these two mouse strains. High-fat diets, regardless of sucrose content, resulted in significant weight gain, higher body fat, and lower body protein and water content in both strains of mice. The shift toward higher body fat and lower protein and water content was far greater in the B6 strain. Low-fat, high-sucrose diets resulted in lower body weight in both strains, as well as significantly greater body protein content in B6 mice. Analysis of intestinal sucrase showed that the enzyme was less active in B6 mice when the diet was high in sucrose. Both sucrase and maltase had lower activity in the presence of high dietary fat in both mouse strains. The percent reduction of intestinal enzyme activity due to dietary fat was similar in both strains. The B6 mouse exhibits disproportionate weight gain and altered body composition on a high-fat diet. This coupled with the reduced body weight and increased body protein on a low-fat, high-sucrose diet suggests that factors-relative to fat metabolism rather than sucrose metabolism are responsible for obesity.
Over 3000 microbial (bacterial and archaeal) genomes have been made publically available to date, providing an unprecedented opportunity to examine evolutionary genomic trends and offering valuable ...reference data for a variety of other studies such as metagenomics. The utility of these genome sequences is greatly enhanced when we have an understanding of how they are phylogenetically related to each other. Therefore, we here describe our efforts to reconstruct the phylogeny of all available bacterial and archaeal genomes. We identified 24, single-copy, ubiquitous genes suitable for this phylogenetic analysis. We used two approaches to combine the data for the 24 genes. First, we concatenated alignments of all genes into a single alignment from which a Maximum Likelihood (ML) tree was inferred using RAxML. Second, we used a relatively new approach to combining gene data, Bayesian Concordance Analysis (BCA), as implemented in the BUCKy software, in which the results of 24 single-gene phylogenetic analyses are used to generate a "primary concordance" tree. A comparison of the concatenated ML tree and the primary concordance (BUCKy) tree reveals that the two approaches give similar results, relative to a phylogenetic tree inferred from the 16S rRNA gene. After comparing the results and the methods used, we conclude that the current best approach for generating a single phylogenetic tree, suitable for use as a reference phylogeny for comparative analyses, is to perform a maximum likelihood analysis of a concatenated alignment of conserved, single-copy genes.
Objectives Water-based soluble and synthetic metalworking fluids (MWF) used in auto manufacturing may be contaminated by endotoxin from Gram-negative bacteria, a possible anticarcinogen via increased ...immunosurveillance. The effectiveness of biocide, generally added to limit bacterial growth is unknown. We investigated whether an inverse relationship between lung cancer and synthetic MWF and biocide - as surrogates of endotoxin exposure -persisted in an extended follow-up of autoworkers. Methods A nested case-control analysis was performed within a retrospective cohort study of 46 399 auto manufacturing workers. Follow-up began in 1941 and was extended from 1985-1995. Mortality rate ratios (MRR) were estimated in Cox regression models for lung cancer as discrete and smoothed functions of cumulative exposure to synthetic MWF (mg/m 3 per year) and years exposed to biocide with both synthetic and soluble MWF. The analysis was also restricted to the subcohort hired on or after 1941 and stratified by follow-up period. Results The splines suggested a non-linear inverse exposure-response for lung cancer mortality with increasing endotoxin exposure. Overall, the greatest reduction in mortality was observed among those with the highest exposure MRR 0.63, 95% confidence interval (95% CI) 0.39-0.98 at the 99th percentile of exposure (15.8 mg/m³ per year). Evidence for an inverse effect was limited to the earlier follow-up period. Effect modification by biocide was marginally significant (P= 0.07); the protective effect of synthetic MWF was observed only for those who were co-exposed. Conclusions The protective effect of synthetic MWF against lung cancer mortality persisted through the extended period of follow-up, although attenuated, and was observed only among workers with co-exposure to biocide and synthetic MWF.
Patients with health conditions associated with impaired vascular function and inflammation may be more susceptible to the adverse health effects of fine particulate (particulate matter with a mass ...median aerodynamic diameter of ≤2.5 μm (PM2.5)) exposure. In 2006, the authors conducted a panel study to investigate directly whether vascular function and inflammation (assessed by C-reactive protein) modify PM2.5-associated reductions in heart rate variability among 23 young male workers (mean age, 40 years) from Massachusetts. Concurrent 24-hour ambulatory electrocardiogram and personal PM2.5 exposure information was collected over a total of 36 person-days, including either or both welding and nonwelding days. Linear mixed models were used to examine the 5-minute standard deviation of normal-to-normal intervals (SDNN) in relation to the moving PM2.5 averages in the preceding 1–4 hours. C-reactive protein levels and 3 measures of vascular function (augmentation index, mean arterial pressure, and pulse pressure) were determined at baseline. The authors observed an inverse association between the 1-hour PM2.5 and 5-minute SDNN. Greater SDNN declines were observed among those with C-reactive protein (Pinteraction < 0.001) and augmentation index (P = 0.06) values at or above the 75th percentile and pulse pressure values below the 75th percentile (P < 0.001). Systemic inflammation and poorer vascular function appear to aggravate particle-related declines in heart rate variability among workers.
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