Employing BAC-reporter constructs in the sea anemone Nematostella vectensis

Author Posting. © The Author(s), 2013. This is the author's version of the work. It is posted here by permission of Oxford University Press for personal use, not for redistribution. The definitive version was published in Integrative and Comparative Biology 53 (2013): 832-846, doi:10.1093/icb/ict091.Changes in the expression and function of genes drive evolutionary change. Comparing how genes are regulated in different species is therefore becoming an important part of evo-devo studies. A key tool for investigating the regulation of genes is represented by Bacterial Artificial Chromosomes-reporter constructs (BAC). BACs are large insert libraries, often >>100 kb, which thus capture the genomic sequences surrounding a gene of interest, including all, or nearly all, of the elements underpinning regulation. Recombinant BACs, containing a reporter gene in place of the endogenous coding sequence of genes, can be utilized to drive the expression of reporter genes under the regulatory control of the gene of interest while still embedded within its genomic context. Systematic deletions within the BAC reporter construct can be used to identify the minimal reporter in an unbiased way, avoiding the risk of overlooking regulatory elements that may be many kilobases away from the transcription start-site. Nematostella vectensis (Edwardsiidae, Anthozoa, Cnidaria) has become an important model in regenerative biology, ecology, and especially in studies of evo-devo and gene-regulatory networks due to its interesting phylogenetic position and amenability to molecular techniques. The increasing interest in this rising model system also led to a demand for methods that can be used to study the regulation of genes in Nematostella. Here we present our progress in employing BAC reporter constructs to visualize gene-expression in Nematostella. Using a new Nematostella-specific recombination cassette, we made nine different BAC reporter constructs. Although five BAC recombinants gave variable effects, three constructs, namely Nv-bra:eGFP::L10 BAC, Nv-dpp:eGFP::L10 BAC, and Nv-grm:eGFP::L10 BAC, delivered promising results. We show that these three constructs express the reporter gene eGFP in 10.4% – 17.2% of all analyzed larvae, out of which 26.2 – 41.9% express GFP in a mosaic fashion within the expected domain. In addition to the expression within the known domains, we also observed cases of misexpression of eGFP and examples that could represent actual expression outside the described domain. Furthermore, we deep-sequenced and assembled five different BACs containing Nv-chordin, Nv-foxa, Nv-dpp, Nv-wnta, and Nvwnt1, to improve assembly around these genes. The use of BAC reporter constructs will foster cis-regulatory analyses in Nematostella and thus help to improve our understanding of the regulatory network in this cnidarian system. Ultimately, this will advance the comparison of gene-regulation across species and lead to a much better understanding of evolutionary changes and novelties.2014-08-1

SeaBase : a multispecies transcriptomic resource and platform for gene network inference

Author Posting. © The Author(s), 2014. This is the author's version of the work. It is posted here by permission of Oxford University Press for personal use, not for redistribution. The definitive version was published in Integrative and Comparative Biology 54 (2014): 250-263, doi: 10.1093/icb/icu065.Marine and aquatic animals are extraordinarily useful as models for identifying mechanisms of development and evolution, regeneration, resistance to cancer, longevity and symbiosis, among many other areas of research. This is due to the great diversity of these organisms and their wide-ranging capabilities. Genomics tools are essential for taking advantage of these “free lessons” of nature. However, genomics and transcriptomics are challenging in emerging model systems. Here, we present SeaBase, a tool for helping to meet these needs. Specifically, SeaBase provides a platform for sharing and searching transcriptome data. More importantly, SeaBase will support a growing number of tools for inferring gene network mechanisms. The first dataset available on SeaBase is a developmental transcriptome profile of the sea anemone Nematostella vectensis (Anthozoa, Cnidaria). Additional datasets are currently being prepared and we are aiming to expand SeaBase to include user-supplied data for any number of marine and aquatic organisms, thereby supporting many potentially new models for gene network studies.2015-06-0

Instances and connectors : issues for a second generation process language

This work is supported by UK EPSRC grants GR/L34433 and GR/L32699Over the past decade a variety of process languages have been defined, used and evaluated. It is now possible to consider second generation languages based on this experience. Rather than develop a second generation wish list this position paper explores two issues: instances and connectors. Instances relate to the relationship between a process model as a description and the, possibly multiple, enacting instances which are created from it. Connectors refers to the issue of concurrency control and achieving a higher level of abstraction in how parts of a model interact. We believe that these issues are key to developing systems which can effectively support business processes, and that they have not received sufficient attention within the process modelling community. Through exploring these issues we also illustrate our approach to designing a second generation process language.Postprin

New genetic loci implicated in fasting glucose homeostasis and their impact on type 2 diabetes risk.

Levels of circulating glucose are tightly regulated. To identify new loci influencing glycemic traits, we performed meta-analyses of 21 genome-wide association studies informative for fasting glucose, fasting insulin and indices of beta-cell function (HOMA-B) and insulin resistance (HOMA-IR) in up to 46,186 nondiabetic participants. Follow-up of 25 loci in up to 76,558 additional subjects identified 16 loci associated with fasting glucose and HOMA-B and two loci associated with fasting insulin and HOMA-IR. These include nine loci newly associated with fasting glucose (in or near ADCY5, MADD, ADRA2A, CRY2, FADS1, GLIS3, SLC2A2, PROX1 and C2CD4B) and one influencing fasting insulin and HOMA-IR (near IGF1). We also demonstrated association of ADCY5, PROX1, GCK, GCKR and DGKB-TMEM195 with type 2 diabetes. Within these loci, likely biological candidate genes influence signal transduction, cell proliferation, development, glucose-sensing and circadian regulation. Our results demonstrate that genetic studies of glycemic traits can identify type 2 diabetes risk loci, as well as loci containing gene variants that are associated with a modest elevation in glucose levels but are not associated with overt diabetes

Impact of Metabolic Regulators on the Expression of the Obesity Associated Genes FTO and NAMPT in Human Preadipocytes and Adipocytes

FTO and NAMPT/PBEF/visfatin are thought to play a role in obesity but their transcriptional regulation in adipocytes is not fully understood. In this study, we evaluated the transcriptional regulation of FTO and NAMPT in preadipocytes and adipocytes by metabolic regulators.We assessed FTO mRNA expression during human adipocyte differentiation of Simpson-Golabi-Behmel syndrome (SGBS) cells and primary subcutaneous preadipocytes in vitro and evaluated the effect of the metabolic regulators glucose, insulin, dexamethasone, IGF-1 and isoproterenol on FTO and NAMPT mRNA expression in SGBS preadipocytes and adipocytes. FTO mRNA levels were not significantly modulated during adipocyte differentiation. Also, metabolic regulators had no impact on FTO expression in preadipocytes or adipocytes. In SGBS preadipocytes NAMPT expression was more than 3fold induced by dexamethasone and isoproterenol and 1.6fold by dexamethasone in adipocytes. Complete glucose restriction caused an increase in NAMPT mRNA expression by more than 5fold and 1.4fold in SGBS preadipocytes and adipocytes, respectively.FTO mRNA expression is not significantly affected by differentiation or metabolic regulators in human adipocytes. The stimulation of NAMPT expression by dexamethasone, isoproterenol and complete glucose restriction may indicate a regulation of NAMPT by metabolic stress, which was more pronounced in preadipocytes compared to mature adipocytes

Pleiotropic Benefit of Monomeric and Oligomeric Flavanols on Vascular Health - A Randomized Controlled Clinical Pilot Study

BACKGROUND: Cardiovascular diseases are expanding to a major social-economic burden in the Western World and undermine man's deep desire for healthy ageing. Epidemiological studies suggest that flavanol-rich foods (e.g. grapes, wine, chocolate) sustain cardiovascular health. For an evidenced-based application, however, sound clinical data on their efficacy are strongly demanded. METHODS: In a double-blind, randomized, placebo-controlled intervention study we supplemented 28 male smokers with 200 mg per day of monomeric and oligomeric flavanols (MOF) from grape seeds. At baseline, after 4 and 8 weeks we measured macro- and microvascular function and a cluster of systemic biomarkers for major pathological processes occurring in the vasculature: disturbances in lipid metabolism and cellular redox balance, and activation of inflammatory cells and platelets. RESULTS: In the MOF group serum total cholesterol and LDL decreased significantly (P ≤ 0.05) by 5% (n = 11) and 7% (n = 9), respectively in volunteers with elevated baseline levels. Additionally, after 8 weeks the ratio of glutathione to glutathione disulphide in erythrocytes rose from baseline by 22% (n = 15, P<0.05) in MOF supplemented subjects. We also observed that MOF supplementation exerts anti-inflammatory effects in blood towards ex vivo added bacterial endotoxin and significantly reduces expression of inflammatory genes in leukocytes. Conversely, alterations in macro- and microvascular function, platelet aggregation, plasma levels of nitric oxide surrogates, endothelin-1, C-reactive protein, fibrinogen, prostaglandin F2alpha, plasma antioxidant capacity and gene expression levels of antioxidant defense enzymes did not reach statistical significance after 8 weeks MOF supplementation. However, integrating all measured effects into a global, so-called vascular health index revealed a significant improvement of overall vascular health by MOF compared to placebo (P ≤ 0.05). CONCLUSION: Our integrative multi-biomarker approach unveiled the pleiotropic vascular health benefit of an 8 weeks supplementation with 200 mg/d MOF in humans. TRIAL REGISTRATION: ClinicalTrials.gov NCT00742287

Framework and baseline examination of the German National Cohort (NAKO)

The German National Cohort (NAKO) is a multidisciplinary, population-based prospective cohort study that aims to investigate the causes of widespread diseases, identify risk factors and improve early detection and prevention of disease. Specifically, NAKO is designed to identify novel and better characterize established risk and protection factors for the development of cardiovascular diseases, cancer, diabetes, neurodegenerative and psychiatric diseases, musculoskeletal diseases, respiratory and infectious diseases in a random sample of the general population. Between 2014 and 2019, a total of 205,415 men and women aged 19–74 years were recruited and examined in 18 study centres in Germany. The baseline assessment included a face-to-face interview, self-administered questionnaires and a wide range of biomedical examinations. Biomaterials were collected from all participants including serum, EDTA plasma, buffy coats, RNA and erythrocytes, urine, saliva, nasal swabs and stool. In 56,971 participants, an intensified examination programme was implemented. Whole-body 3T magnetic resonance imaging was performed in 30,861 participants on dedicated scanners. NAKO collects follow-up information on incident diseases through a combination of active follow-up using self-report via written questionnaires at 2–3 year intervals and passive follow-up via record linkages. All study participants are invited for re-examinations at the study centres in 4–5 year intervals. Thereby, longitudinal information on changes in risk factor profiles and in vascular, cardiac, metabolic, neurocognitive, pulmonary and sensory function is collected. NAKO is a major resource for population-based epidemiology to identify new and tailored strategies for early detection, prediction, prevention and treatment of major diseases for the next 30 years. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10654-022-00890-5

New genetic loci link adipose and insulin biology to body fat distribution.

Body fat distribution is a heritable trait and a well-established predictor of adverse metabolic outcomes, independent of overall adiposity. To increase our understanding of the genetic basis of body fat distribution and its molecular links to cardiometabolic traits, here we conduct genome-wide association meta-analyses of traits related to waist and hip circumferences in up to 224,459 individuals. We identify 49 loci (33 new) associated with waist-to-hip ratio adjusted for body mass index (BMI), and an additional 19 loci newly associated with related waist and hip circumference measures (P < 5 × 10(-8)). In total, 20 of the 49 waist-to-hip ratio adjusted for BMI loci show significant sexual dimorphism, 19 of which display a stronger effect in women. The identified loci were enriched for genes expressed in adipose tissue and for putative regulatory elements in adipocytes. Pathway analyses implicated adipogenesis, angiogenesis, transcriptional regulation and insulin resistance as processes affecting fat distribution, providing insight into potential pathophysiological mechanisms

The FLUXNET2015 dataset and the ONEFlux processing pipeline for eddy covariance data

The FLUXNET2015 dataset provides ecosystem-scale data on CO2, water, and energy exchange between the biosphere and the atmosphere, and other meteorological and biological measurements, from 212 sites around the globe (over 1500 site-years, up to and including year 2014). These sites, independently managed and operated, voluntarily contributed their data to create global datasets. Data were quality controlled and processed using uniform methods, to improve consistency and intercomparability across sites. The dataset is already being used in a number of applications, including ecophysiology studies, remote sensing studies, and development of ecosystem and Earth system models. FLUXNET2015 includes derived-data products, such as gap-filled time series, ecosystem respiration and photosynthetic uptake estimates, estimation of uncertainties, and metadata about the measurements, presented for the first time in this paper. In addition, 206 of these sites are for the first time distributed under a Creative Commons (CC-BY 4.0) license. This paper details this enhanced dataset and the processing methods, now made available as open-source codes, making the dataset more accessible, transparent, and reproducible.Peer reviewe