eFORGE: A Tool for Identifying Cell Type-Specific Signal in Epigenomic Data

Epigenome-wide association studies (EWAS) provide an alternative approach for studying human disease through consideration of non-genetic variants such as altered DNA methylation. To advance the complex interpretation of EWAS, we developed eFORGE (http://eforge.cs.ucl.ac.uk/), a new standalone and web-based tool for the analysis and interpretation of EWAS data. eFORGE determines the cell type-specific regulatory component of a set of EWAS-identified differentially methylated positions. This is achieved by detecting enrichment of overlap with DNase I hypersensitive sites across 454 samples (tissues, primary cell types, and cell lines) from the ENCODE, Roadmap Epigenomics, and BLUEPRINT projects. Application of eFORGE to 20 publicly available EWAS datasets identified disease-relevant cell types for several common diseases, a stem cell-like signature in cancer, and demonstrated the ability to detect cell-composition effects for EWAS performed on heterogeneous tissues. Our approach bridges the gap between large-scale epigenomics data and EWAS-derived target selection to yield insight into disease etiology.C.E.B. was supported by a PhD fellowship from the EU-FP7 project EpiTrain (316758). J.H. was supported by the UCL Cancer Institute Research Trust. V.K.R. was supported by BLUEPRINT (282510). K.D. was funded as a HSST trainee by NHS Health Education England. M.F. was supported by the BHF Cambridge Centre of Excellence (RE/13/6/30180). Research in W.H.O.’s laboratory was supported by EU-FP7 project BLUEPRINT (282510) and by program grants from the National Institute for Health Research (NIHR, http://www.nihr.ac.uk) and the British Heart Foundation under numbers RP-PG-0310-1002 and RG/09/12/28096 (https://www.bhf.org.uk/). W.H.O.’s laboratory receives funding from NHS Blood and Transplant for facilities. We gratefully acknowledge the participation of all NIHR Cambridge BioResource volunteers. We thank the Cambridge BioResource staff for their help with volunteer recruitment. We thank members of the Cambridge BioResource SAB and Management Committee for their support of our study and the National Institute for Health Research Cambridge Biomedical Research Centre for funding. R.S. and his group were supported by the European Union in the framework of the BLUEPRINT Project (HEALTH-F5-2011-282510) and the German Ministry of Science and Education (BMBF) in the framework of the MMML-MYC-SYS project (036166B). We thank Deborah Winter (Weizmann Institute) for supplying a set of microglial enhancers from Lavin et al. (2014). Research in S.B.’s laboratory was supported by the Wellcome Trust (99148), Royal Society Wolfson Research Merit Award (WM100023), and EU-FP7 projects EpiTrain (316758), EPIGENESYS (257082), and BLUEPRINT (282510)

Rare Copy Number Deletions Predict Individual Variation in Intelligence

Phenotypic variation in human intellectual functioning shows substantial heritability, as demonstrated by a long history of behavior genetic studies. Many recent molecular genetic studies have attempted to uncover specific genetic variations responsible for this heritability, but identified effects capture little variance and have proven difficult to replicate. The present study, motivated an interest in “mutation load” emerging from evolutionary perspectives, examined the importance of the number of rare (or infrequent) copy number variations (CNVs), and the total number of base pairs included in such deletions, for psychometric intelligence. Genetic data was collected using the Illumina 1MDuoBeadChip Array from a sample of 202 adult individuals with alcohol dependence, and a subset of these (N = 77) had been administered the Wechsler Abbreviated Scale of Intelligence (WASI). After removing CNV outliers, the impact of rare genetic deletions on psychometric intelligence was investigated in 74 individuals. The total length of the rare deletions significantly and negatively predicted intelligence (r = −.30, p = .01). As prior studies have indicated greater heritability in individuals with relatively higher parental socioeconomic status (SES), we also examined the impact of ethnicity (Anglo/White vs. Other), as a proxy measure of SES; these groups did not differ on any genetic variable. This categorical variable significantly moderated the effect of length of deletions on intelligence, with larger effects being noted in the Anglo/White group. Overall, these results suggest that rare deletions (between 5% and 1% population frequency or less) adversely affect intellectual functioning, and that pleotropic effects might partly account for the association of intelligence with health and mental health status. Significant limitations of this research, including issues of generalizability and CNV measurement, are discussed

Application of the bacteriophage Mu-driven system for the integration/amplification of target genes in the chromosomes of engineered Gram-negative bacteria—mini review

The advantages of phage Mu transposition-based systems for the chromosomal editing of plasmid-less strains are reviewed. The cis and trans requirements for Mu phage-mediated transposition, which include the L/R ends of the Mu DNA, the transposition factors MuA and MuB, and the cis/trans functioning of the E element as an enhancer, are presented. Mini-Mu(LR)/(LER) units are Mu derivatives that lack most of the Mu genes but contain the L/R ends or a properly arranged E element in cis to the L/R ends. The dual-component system, which consists of an integrative plasmid with a mini-Mu and an easily eliminated helper plasmid encoding inducible transposition factors, is described in detail as a tool for the integration/amplification of recombinant DNAs. This chromosomal editing method is based on replicative transposition through the formation of a cointegrate that can be resolved in a recombination-dependent manner. (E-plus)- or (E-minus)-helpers that differ in the presence of the trans-acting E element are used to achieve the proper mini-Mu transposition intensity. The systems that have been developed for the construction of stably maintained mini-Mu multi-integrant strains of Escherichia coli and Methylophilus methylotrophus are described. A novel integration/amplification/fixation strategy is proposed for consecutive independent replicative transpositions of different mini-Mu(LER) units with “excisable” E elements in methylotrophic cells

Subcortical volumes across the lifespan: Data from 18,605 healthy individuals aged 3-90 years

Age has a major effect on brain volume. However, the normative studies available are constrained by small sample sizes, restricted age coverage and significant methodological variability. These limitations introduce inconsistencies and may obscure or distort the lifespan trajectories of brain morphometry. In response, we capitalized on the resources of the Enhancing Neuroimaging Genetics through Meta-Analysis (ENIGMA) Consortium to examine age-related trajectories inferred from cross-sectional measures of the ventricles, the basal ganglia (caudate, putamen, pallidum, and nucleus accumbens), the thalamus, hippocampus and amygdala using magnetic resonance imaging data obtained from 18,605 individuals aged 3-90 years. All subcortical structure volumes were at their maximum value early in life. The volume of the basal ganglia showed a monotonic negative association with age thereafter; there was no significant association between age and the volumes of the thalamus, amygdala and the hippocampus (with some degree of decline in thalamus) until the sixth decade of life after which they also showed a steep negative association with age. The lateral ventricles showed continuous enlargement throughout the lifespan. Age was positively associated with inter-individual variability in the hippocampus and amygdala and the lateral ventricles. These results were robust to potential confounders and could be used to examine the functional significance of deviations from typical age-related morphometric patterns

The impact of transposable element activity on therapeutically relevant human stem cells

Human stem cells harbor significant potential for basic and clinical translational research as well as regenerative medicine. Currently ~ 3000 adult and ~ 30 pluripotent stem cell-based, interventional clinical trials are ongoing worldwide, and numbers are increasing continuously. Although stem cells are promising cell sources to treat a wide range of human diseases, there are also concerns regarding potential risks associated with their clinical use, including genomic instability and tumorigenesis concerns. Thus, a deeper understanding of the factors and molecular mechanisms contributing to stem cell genome stability are a prerequisite to harnessing their therapeutic potential for degenerative diseases. Chemical and physical factors are known to influence the stability of stem cell genomes, together with random mutations and Copy Number Variants (CNVs) that accumulated in cultured human stem cells. Here we review the activity of endogenous transposable elements (TEs) in human multipotent and pluripotent stem cells, and the consequences of their mobility for genomic integrity and host gene expression. We describe transcriptional and post-transcriptional mechanisms antagonizing the spread of TEs in the human genome, and highlight those that are more prevalent in multipotent and pluripotent stem cells. Notably, TEs do not only represent a source of mutations/CNVs in genomes, but are also often harnessed as tools to engineer the stem cell genome; thus, we also describe and discuss the most widely applied transposon-based tools and highlight the most relevant areas of their biomedical applications in stem cells. Taken together, this review will contribute to the assessment of the risk that endogenous TE activity and the application of genetically engineered TEs constitute for the biosafety of stem cells to be used for substitutive and regenerative cell therapiesS.R.H. and P.T.R. are funded by the Government of Spain (MINECO, RYC-2016- 21395 and SAF2015–71589-P [S.R.H.]; PEJ-2014-A-31985 and SAF2015–71589- P [P.T.R.]). GGS is supported by a grant from the Ministry of Health of the Federal Republic of Germany (FKZ2518FSB403)

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe

The many meanings of evidence: a comparative analysis of the forms and roles of evidence within three health policy processes in Cambodia

Background Discussions within the health community routinely emphasise the importance of evidence in informing policy formulation and implementation. Much of the support for the evidence-based policy movement draws from concern that policy decisions are often based on inadequate engagement with high-quality evidence. In many such discussions, evidence is treated as differing only in quality, and assumed to improve decisions if it can only be used more. In contrast, political science scholars have described this as an overly simplistic view of the policy-making process, noting that research ‘use’ can mean a variety of things and rely on nuanced aspects of political systems. An approach more in recognition of how policy-making systems operate in practice can be to consider how institutions and ideas influence which pieces of evidence appear to be relevant for, and are used within, different policy processes. Methods Drawing on in-depth interviews undertaken in 2015/16 with key health sector stakeholders in Cambodia, we investigate the evidence perceived to be relevant to policy decisions for three contrasting health policy examples – tobacco control, HIV/AIDS and performance-based salary incentives. These cases allow us to examine the ways that policy relevant evidence may differ given the framing of the issue and the broader institutional context in which evidence is considered. Results The three health issues show few similarities in how pieces of evidence were used in various aspects of policy-making, despite all being discussed within a broad policy environment in which evidence-based policymaking is rhetorically championed. Instead, we find that evidence use can be better understood by mapping how these health policy issues differ in terms of the issue characteristics, and also in terms of the stakeholders structurally established as having dominant influence for each issue. Both of these have important implications for evidence use. Contrasting concerns of key stakeholders meant that evidence related to differing issues could be understood in terms of how it was policy relevant. The stakeholders involved, however, could further be seen to possess differing logics about how to go about achieving their various outcomes – logics that could further help explain the differences seen in evidence utilisation. Conclusion A comparative approach reiterates that evidence is not a uniform concept for which more is obviously better, but rather illustrates how different constructions and pieces of evidence become relevant in relation to the features of specific health policy decisions. An institutional approach that considers the structural position of stakeholders with differing core goals or objectives, as well as their logics related to evidence utilisation, can further help to understand some of the complexities of evidence use in health policymaking

Heterologous extracellular production of enterocin P from Enterococcus faecium P13 in the methylotrophic bacterium Methylobacterium extorquens.

Enterocin P (EntP), a strong antilisterial pediocin-like bacteriocin from Enterococcus faecium P13, was produced by Methylobacterium extorquens. For heterologous expression of EntP in the methylotrophic bacterium M. extorquens, a recombinant plasmid was constructed. The gene encoding the EntP structural gene (entP) was cloned into the plasmid vector pCM80, under control of the methanol dehydrogenase promoter (P(mxaF)), to generate plasmid pS25. When M. extorquens ATCC 55366 was transformed with pS25, EntP was detected and quantified in supernatants of the recombinant M. extorquens S25 strain by using specific anti-EntP antibodies and a non-competitive indirect enzyme-linked immunosorbent assay (NCI-ELISA). Purification of EntP by hydrophobic adsorption and reverse-phase (RP-FPLC) chromatographies, permitted recovery of active EntP from the supernatants of M. extorquens S25 grown in a synthetic defined medium

Procurement and Importing in New Product Projects of Brazilian Aerospace Program

New product development is a business process with many functional interactions in a company. The concurrency of these interactions must be managed in order to meet the preestablished schedule, budget and scope. The issue of procurement is central to a succesful project. When a new project belongs to an aerospace program this issue is even more crucial. And when the aerospace program belongs to a developing country such as Brazil, the core issue involves its budget and schedule planning. This article addresses the question of procurement in a small company designing a new satellite camera for the Brazilian Government. The procurement process was mapped, a monitoring structure was created and performance indicators were developed. The performance indicators are discussed to understand the leverage of each kind of purchased item and each process step on costs and schedule.Pages: 790-79