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19 pags., 8 figs., 4 tabs. -- This article belongs to the Special Issue The Amazing World of IDPs in Human Diseases ; Methyl-CpG binding protein 2 (MeCP2) is a transcriptional regulator and a chromatin-binding protein involved in neuronal development and maturation. Loss-of-function mutations in MeCP2 result in Rett syndrome (RTT), a neurodevelopmental disorder that is the main cause of mental retardation in females. MeCP2 is an intrinsically disordered protein (IDP) constituted by six domains. Two domains are the main responsible elements for DNA binding (methyl-CpG binding domain, MBD) and recruitment of gene transcription/silencing machinery (transcription repressor domain, TRD). These two domains concentrate most of the RTT-associated mutations. R106W and R133C are associated with severe and mild RTT phenotype, respectively. We have performed a comprehensive characterization of the structural and functional impact of these substitutions at molecular level. Because we have previously shown that the MBD-flanking disordered domains (N-terminal domain, NTD, and intervening domain, ID) exert a considerable influence on the structural and functional features of the MBD (Claveria-Gimeno, R. et al. Sci Rep. 2017, 7, 41635), here we report the biophysical study of the influence of the protein scaffold on the structural and functional effect induced by these two RTT-associated mutations. These results represent an example of how a given mutation may show different effects (sometimes opposing effects) depending on the molecular context. ; This research was funded by the Spanish Ministry of Economy and Competitiveness and European ERDF Funds (MCIU/AEI/FEDER, EU) (BFU2016-78232-P to A.V.C.; BES-2017-080739 to D.O.A.); Miguel Servet Program from Instituto de Salud Carlos III (CPII13/00017 to O.A.); Fondo de Investigaciones Sanitarias from Instituto de Salud Carlos III and European Union (ERDF/ESF, "Investing in your future") (PI15/00663 and PI18/00349 to O.A.); Diputación General de Aragón (Protein Targets and Bioactive Compounds Group E45_17R to A.V.C. and Digestive Pathology Group B25_17R to O.A.); and the Centro de Investigación Biomédica en Red en Enfermedades Hepáticas y Digestivas (CIBERehd). ; Peer reviewed

15 p.-5 fig.-3 tab. ; Protein modification by lipid derived reactive species or lipoxidation is increased during oxidative stress, a common feature observed in many pathological conditions. Biochemical and functional consequences of lipoxidation include changes in the conformation and assembly of the target proteins, altered recognition of ligands and/or cofactors, changes in the interactions with DNA or protein-protein interactions, modifications in membrane partitioning and binding and/or subcellular localization. These changes may impact, directly or indirectly, signaling pathways involved in the activation of cell defense mechanisms, but when these are overwhelmed they may lead to pathological outcomes. Mass spectrometry provides state of the art approaches for the identification and characterization of lipoxidized proteins/residues and the modifying species. Nevertheless, understanding the complexity of the functional effects of protein lipoxidation requires the use of additional methodologies. Herein, biochemical and biophysical methods used to detect and measure functional effects of protein lipoxidation at different levels of complexity, from in vitro and reconstituted cell-like systems to cells, are reviewed, focusing especially on macromolecular interactions. Knowledge generated through innovative and complementary technologies will contribute to comprehend the role of lipoxidation in pathophysiology and, ultimately, its potential as target for therapeutic intervention. ; This work was supported by the European Union's Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement number 675132 (http://cordis.europa.eu/project/rcn/198275_en.html), and by grants from the Spanish Ministerio de Economía y Competitividad (MINECO/FEDER, http://www.mineco.gob.es/portal/site/mineco/idi) SAF2015-68590-R to DPS and BFU2016-75471-C2-1-P to GR, and RETIC Aradyal from ISCIII/FEDER (RD16/0006/0021) to DPS. ; Peer reviewed

The rolling-circle replication is the most common mechanism for the replication of small plasmids carrying antibiotic resistance genes in Gram-positive bacteria. It is initiated by the binding and nicking of double-stranded origin of replication by a replication initiator protein (Rep). Duplex unwinding is then performed by the PcrA helicase, whose processivity is critically promoted by its interaction with Rep. How Rep and PcrA proteins interact to nick and unwind the duplex is not fully understood. Here, we have used magnetic tweezers to monitor PcrA helicase unwinding and its relationship with the nicking activity of Staphylococcus aureus plasmid pT181 initiator RepC. Our results indicate that PcrA is a highly processive helicase prone to stochastic pausing, resulting in average translocation rates of 30 bp s-1, while a typical velocity of 50 bp s-1 is found in the absence of pausing. Single-strand DNA binding protein did not affect PcrA translocation velocity but slightly increased its processivity. Analysis of the degree of DNA supercoiling required for RepC nicking, and the time between RepC nicking and DNA unwinding, suggests that RepC and PcrA form a protein complex on the DNA binding site before nicking. A comprehensive model that rationalizes these findings is presented. ; Ministerio de Economía y Competitividad (MINECO) [BFU2017-83794-P (AEI/FEDER, UE) to F.M.-H.]; European Research Council (ERC) under the European Union Horizon 2020 research and innovation grant agreemen t[681299 to F.M.-H.]; National Science Foundation [toS.H.L.]. C.C. was supported by a 'Severo Ochoa' postdoc-toral contract from the National Center of Biotechnology (CNB-CSIC); Funding for open access charge: EuropeanResearch Council [681299]. ; Peer reviewed

Linking non-coding genetic variants associated with the risk of diseases or disease relevant traits to target genes is a crucial step to realize GWAS potential in the introduction of precision medicine. Here we set out to determine the mechanisms underpinning variant association with platelet quantitative traits using cell type matched epigenomic data and promoter long-range interactions. We identified potential regulatory functions for 423 of 565 (75%) non-coding platelet variants and highlighted, including ex vivo and genome editing proof of principle validation, the role of associated variants in super enhancers in controlling archetypical platelet functions. ; The work was funded by a grant from the European Commission 7th Framework Program (FP7/2007-2013, grant 282510, BLUEPRINT). F.A.C. is a Medical Research Council (MRC) clinical fellow (MR/K024043/1); K.D. is funded by NHS Health Education England; M.F. is supported by the British Heart Foundation (BHF) Cambridge Centre of Excellence (RE/13/6/30180); research in the W.H.O. laboratory is also supported by grants from Bristol Myers-Squibb, BHF, European Commission, MRC, NIHR (W.H.O. is NIHR Senior Investigator) and NHS Blood and Transplant (NHSBT). RP is supported by the European Union's Horizon 2020 research and innovation program under grant agreement No. 692041 (TrainMALTA, H2020-TWINN-2015). L.V. is funded by the ERC grant Relieve IMDs (ERC-2011-StG). P.M. and A-S.L. are funded by the NIHR Cambridge Biomedical Research Centre (BRC) hIPSCs core facility. B.M.J., P.Fr. and M.S. are supported by the MRC (MR/L007150/1) and Biotechnology and Biological Sciences Research Council (BB/J004480/1). K.F. is funded by FWO-Vlaanderen (G.0B17.13N) and BOF KULeuven (OT/14/098). Work at EMBL-EBI received additional support from the Wellcome Trust (WT095908) to P.Fl and from the European Molecular Biology Laboratory to L.C., M.K., P.Fl. and O.S. The MRC/BHF Cardiovascular Epidemiology receives core support from the MRC (G0800270), the BHF (SP/09/002), the NIHR and NIHR Cambridge BRC, as well as grants from the European Research Council (268834), the European Commission FP7 (HEALTH-F2-2012-279233), Merck and Pfizer. J.D. is a BHF Professor, European Research Council Senior Investigator, and NIHR Senior Investigator. The NIHR Blood and Transplant Research Unit in Donor Health and Genomics at the University of Cambridge is funded by NIHR and NHSBT.

The interactions between viruses and their microbial hosts play a central role in the control of microbial communities in nature. However, the study of such interactions within the uncultured majority is technically very challenging. Here, we review how microarray tools can be used to analyze the interactions between viruses and their microbial hosts in nature, away from laboratory pure culture-based models. We show examples of how DNA arrays have been used to study the expression of viral assemblages in natural samples, and to assign viruses to hosts within uncultured communities. Finally, we briefly discuss the possibilities of protein and glycan arrays to gain insight into the ways microbes interact with their viruses. ; Our current studies with viral microarrays are supported by projects CGL2012-39627-C03-01 (to Josefa Antón) and AYA2011-24803 (to Víctor Parro) of the Spanish Ministry of Science and Innovation, which are co-financed with FEDER support from the European Union. ; Peer reviewed ; Peer Reviewed

The field of genealogy has rapidly evolved with the rise of direct-to-consumer DNA testing, a digital technology with the potential to disrupt previously held understandings of family history. Recent theorisation in the sociology of personal life provides a useful lens to attune to potent sensations and affinities sparked by direct-to-consumer DNA testing. Our analysis focuses on the accounts of three donor-conceived people – Thomas, Simone and Amira – whose DNA discoveries significantly disrupted their familial, cultural and national identities. We explore three cascading flashes of insight derived from our analysis: realising and reordering the family tree – how DNA discoveries were experienced through a range of initial sensations from shock to stupidity to feeling cheated and duped; reflecting on resemblance – how DNA discoveries were felt through sensations of similarity and alterity; and re-evaluating relationships – how the sensations of DNA discoveries (re)shaped interactions with family. This article contributes to understanding donor-conceived people's relational practices following DNA discoveries and the role of sensations in these practices. We argue that such sensations impact understandings of self and familial relationships, raising questions about social and genetic inheritance and who counts as family.

During repair of DNA double-strand breaks, resection of DNA ends influences how these lesions will be repaired. If resection is activated, the break will be channeled through homologous recombination; if not, it will be simply ligated using the non-homologous end-joining machinery. Regulation of resection relies greatly on modulating CtIP, which can be done by modifying: i) its interaction partners, ii) its post-translational modifications, or iii) its cellular levels, by regulating transcription, splicing and/or protein stability/degradation. Here, we have analyzed the role of ALC1, a chromatin remodeler previously described as an integral part of the DNA damage response, in resection. Strikingly, we found that ALC1 affects resection independently of chromatin remodeling activity or its ability to bind damaged chromatin. In fact, it cooperates with the RNA-helicase eIF4A1 to help stabilize the most abundant splicing form of CtIP mRNA. This function relies on the presence of a specific RNA sequence in the 5′ UTR of CtIP. Therefore, we describe an additional layer of regulation of CtIP—at the level of mRNA stability through ALC1 and eIF4A1. ; This work was financed by an R+D+I grant from the Spanish Ministry of Economy and Competitivity (SAF2016-74855-P) and by the European Union Regional Funds (FEDER). FM-N was funded with an FPU fellowship from the Spanish Ministry of Education, and JR and GR-R were supported by the Regional Government of Andalucía (Junta de Andalucía) with a contract of the program "GARANTÍA JUVENIL EN LA UNIVERSIDAD DE SEVILLA". CABIMER is supported by the regional government of Andalucía (Junta de Andalucía). ; Peer reviewed

This paper examines an inflammatory subject - `DNA typing' (or `DNA fingerprinting' as it is popularly called) - to show how credible scientific knowledge is produced through the systematic erasure of uncertainty and random variation. This erasure occurs at the levels of measurement and of group processes. I use the history of DNA typing in the United States as a case study to develop a three-fold argument. First, that objective knowledge is achieved through interactions and social processes that erase the actions of the representing subject from the representations made of the natural world. Second, that this set of social processes and interactions includes normative judgements, but that these judgements do not pose a threat to objectivity: instead, they are what constitute objectivity. Objectivity is attained through personal judgements and evaluations of what is good enough to constitute an objective measurement, or judgements of how big or small a deviation must be to count for or against a particular theoretical interpretation. Third, I argue that measurements, the result of the translation of properties of the natural world into numbers, always have a gap or error between the theoretically predicted measurement and the empirically obtained measurement. At some point this error cannot be reduced any further. The final measurement is the outcome of negotiation and interaction between people and nature. I demonstrate that these judgements, evaluations and negotiations take place within a laboratory, when forensic workers construct estimates of measurement error, and also in negotiations among disparate social groups involved in calculating random match probabilities. Scientific and legal disputes about the construction of measurements and measurement standards for DNA profiling began in the late 1980s, and continued for the better part of a decade. In this paper, the protracted controversy provides especially rich materials for examining the social construction of measurement.

ABSTARCT Biotechnology has enabled the modification of agricultural materials in a very precise way. Crops have been modified through the insertion of new traits or the inhibition of existing gene functions, named Genetically Modified Organism (GMO), and resulted in improved tolerance of herbicide and/or increased resistance against pests, viruses and fungi. Commercial cultivation of GMO started in 1996 and increased rapidly in 2003 according to a recently released report by the International Service for the Acquisition of Agri-Biotech Applications (ISAAA), depicted continuing consumer resistance in Europe and other part of the world. Upon these developments, the European Union regulations mandated labeling of GMOs containing food and as a consequence, the labeling of GMO containing product in the case of exceeding the1% threshold of alien DNA is required. The aim of the study is to be able to detect and quantify the GMO from the mixture of natural food components. The surface plasmon resonance (SPR) technique combined with fluorescence was used for this purpose. During the presented studies, two key issues are addressed and tried to solve; what is the best strategy to design and built an interfacial architecture of a probe oligonucletide layer either on a two dimensional surface or on an array platform; and what is the best detection method allowing for a sensitive monitoring of the hybridisation events. The study includes two parts: first part includes characterization of different PNAs on a 2D planar surface by defining affinity constants using the very well established optical method "Surface Plasmon Fluorescence Spectroscopy"(SPFS) and on the array platform by "Surface Plasmon Fluorescence Microscopy" (SPFM), and at the end comparison of the sensitivity of these two techniques. The second part is composed of detection of alien DNA in food components by using DNA and PNA catcher probes on the array platform in real-time by SPFM.

14 p.-5 fig. ; Division ring formation at midcell is controlled by various mechanisms in Escherichia coli, one of them being the linkage between the chromosomal Ter macrodomain and the Z-ring mediated by MatP, a DNA binding protein that organizes this macrodomain and contributes to the prevention of premature chromosome segregation. Here we show that, during cell division, just before splitting the daughter cells, MatP seems to localize close to the cytoplasmic membrane, suggesting that this protein might interact with lipids. To test this hypothesis, we investigated MatP interaction with lipids in vitro. We found that, when encapsulated inside vesicles and microdroplets generated by microfluidics, MatP accumulates at phospholipid bilayers and monolayers matching the lipid composition in the E. coli inner membrane. MatP binding to lipids was independently confirmed using lipid-coated microbeads and biolayer interferometry assays, which suggested that the recognition is mainly hydrophobic. Interaction of MatP with the lipid membranes also occurs in the presence of the DNA sequences specifically targeted by the protein, but there is no evidence of ternary membrane/protein/DNA complexes. We propose that the association of MatP with lipids may modulate its spatiotemporal localization and its recognition of other ligands. ; IMPORTANCE The division of an E. coli cell into two daughter cells with equal genomic information and similar size requires duplication and segregation of the chromosome and subsequent scission of the envelope by a protein ring, the Z-ring. MatP is a DNA binding protein that contributes both to the positioning of the Z-ring at midcell and the temporal control of nucleoid segregation. Our integrated in vivo and in vitro analysis provides evidence that MatP can interact with lipid membranes reproducing the phospholipid mixture in the E. coli inner membrane, without concomitant recruitment of the short DNA sequences specifically targeted by MatP. This observation strongly suggests that the membrane may play a role in the regulation of the function and localization of MatP, which could be relevant for the coordination of the two fundamental processes in which this protein participates, nucleoid segregation and cell division. ; This work was supported by the Spanish government through grants BFU2014-52070-C2-2-P and BFU2016-75471-C2-1-P (to G. R.) and by the Dutch government through grant NWO ALW open program no. 822.02.019 (to N.Y.M.). M.R.-R was supported by the Agencia Estatal de Investigación and the European Social Fund through grant BES-2017-082003. ; Peer reviewed

The worldwide rapid declines in insect and plant abundance and diversity that have occurred in the past decades have gained public attention and demand for political actions to counteract these declines are growing. Rapid large-scale biomonitoring can aid in observing these changes and provide information for decisions for land management and species protection. Malaise traps have long been used for insect sampling and when insects are captured in these traps, they carry traces of plants they have visited on the body surface or as digested food material in the gut contents. Metabarcoding offers a promising method for identifying these plant traces, providing insight into the plants with which insects are directly interacting at a given time. To test the efficacy of DNA metabarcoding with these sample types, 79 samples from 21 sites across Germany were analysed with the ITS2 barcode. This study, to our knowledge, is the first examination of metabarcoding plant DNA traces from Malaise trap samples. Here, we report on the feasibility of sequencing these sample types, analysis of the resulting taxa, the usage of cultivated plants by insects near nature conservancy areas and the detection of rare and neophyte species. Due to the frequency of contamination and false positive reads, isolation and PCR negative controls should be used in every reaction. Metabarcoding has advantages in efficiency and resolution over microscopic identification of pollen and is the only possible identification method for the other plant traces from Malaise traps and could provide a broad utility for future studies of plant-insect interactions.

Systematic studies of cancer genomes have provided unprecedented insights into the molecular nature of cancer. Using this information to guide the development and application of therapies in the clinic is challenging. Here, we report how cancer-driven alterations identified in 11,289 tumors from 29 tissues (integrating somatic mutations, copy number alterations, DNA methylation, and gene expression) can be mapped onto 1,001 molecularly annotated human cancer cell lines and correlated with sensitivity to 265 drugs. We find that cell lines faithfully recapitulate oncogenic alterations identified in tumors, find that many of these associate with drug sensitivity/resistance, and highlight the importance of tissue lineage in mediating drug response. Logic-based modeling uncovers combinations of alterations that sensitize to drugs, while machine learning demonstrates the relative importance of different data types in predicting drug response. Our analysis and datasets are rich resources to link genotypes with cellular phenotypes and to identify therapeutic options for selected cancer sub-populations. ; This work was funded by the Wellcome Trust (086375 and 102696). F.I. was supported by the European Bioinformatics Institute and Wellcome Trust Sanger Institute post-doctoral (ESPOD) program. T.A.K. was supported by the National Cancer Institute (U24CA143835) and the Netherlands Organization for Scientific Research. D.T. was supported by the People Programme (Marie Curie Actions) of the 7th Framework Programme of the European Union (FP7/2007-2013; 600388) and the Agency of Competitiveness for Companies of the Government of Catalonia (ACCIO´ ). N.L.-B. was supported by La Fundacio ´ la Marato´ de TV3. M.E. was funded by the European Research Council (268626), the Ministerio de Ciencia e Innovacion (SAF2011-22803), the Institute of Health Carlos III (ISCIII) under the Integrated Project of Excellence (PIE13/00022), the Spanish Cancer Research Network (RD12/0036/0039), the Health and Science Departments of the Catalan Government Generalitat de Catalunya 2014-SGR 633, and the Cellex Foundation. U.M. was supported by a Cancer Research UK Clinician Scientist Fellowship. We thank Aiqing He for expression data and Ilya Shmulevich for assistance with the LOBICO framework. We thank P. Campbell, M. Ranzani, J. Brammeld, M. Petljak, F. Behan, C. Alsinet Armengol, H. Francies, V. Grinkevich, and A. ''Lilla'' Mupo for useful comments. P.R.-M., H.C., and H.d.S. are employees and shareholders of Bristol-Myers Squibb. Research in the M.J.G. lab is supported in part with funding from AstraZeneca.

Faithful segregation of bacterial chromosomes relies on the ParABS partitioning system and the SMC complex. In this work, we used single-molecule techniques to investigate the role of cytidine triphosphate (CTP) binding and hydrolysis in the critical interaction between centromere-like parS DNA sequences and the ParB CTPase. Using a combined optical tweezers confocal microscope, we observe the specific interaction of ParB with parS directly. Binding around parS is enhanced by the presence of CTP or the non-hydrolysable analogue CTPγS. However, ParB proteins are also detected at a lower density in distal non-specific DNA. This requires the presence of a parS loading site and is prevented by protein roadblocks, consistent with one-dimensional diffusion by a sliding clamp. ParB diffusion on non-specific DNA is corroborated by direct visualization and quantification of movement of individual quantum dot labelled ParB. Magnetic tweezers experiments show that the spreading activity, which has an absolute requirement for CTP binding but not hydrolysis, results in the condensation of parS-containing DNA molecules at low nanomolar protein concentrations. ; Work in the MSD lab was supported by the Wellcome Trust (New Investigator Award 100401/Z/12/Z to MSD) and the BBSRC (South West Biosciences Doctoral Training studentship to GLMF). FM-H acknowledges support from the European Research Council (ERC) under the European Union Horizon 2020 Research and Innovation Program (grant agreement 681299). Work in the Moreno-Herrero laboratory was also supported by Spanish Ministry of Economy and Competitiveness grant BFU2017-83794-P (AEI/FEDER, UE to FM-H) and Comunidad de Madrid grants Tec4-Bio – S2018/NMT-4443 and NanoBioCancer – Y2018/BIO-4747 (to FM-H).

Mammalian DNA methyltransferase 1 (DNMT1) is essential for maintaining DNA methylation patterns after cell division. Disruption of DNMT1 catalytic activity results in whole genome cytosine demethylation of CpG dinucleotides, promoting severe dysfunctions in somatic cells and during embryonic development. While these observations indicate that DNMT1-dependent DNA methylation is required for proper cell function, the possibility that DNMT1 has a role independent of its catalytic activity is a matter of controversy. Here, we provide evidence that DNMT1 can support cell functions that do not require the C-terminal catalytic domain. We report that PCNA and DMAP1 domains in the N-terminal region of DNMT1 are sufficient to modulate E-cadherin expression in the absence of noticeable changes in DNA methylation patterns in the gene promoters involved. Changes in E-cadherin expression are directly associated with regulation of β-catenin-dependent transcription. Present evidence suggests that the DNMT1 acts on E-cadherin expression through its direct interaction with the E-cadherin transcriptional repressor SNAIL1 ; MEC (SAF2008-00609 to J.E.); MEC (SAF2007-63051 and Consolider CSD2007-00017); EU (MRTN-CT-2004- 005428 to A.C.); MEC (SAF2004-07729, Consolider CSD2006-49); EU (FP7-CANCERDIP-2007-200620 to M.E.); MEC (SAF2010-19152 to J.R.). M.E. is an ICREA Research Professor. J.E. is a Ramon y Cajal Program researcher (MICIIN). Funding for open access charge: Consolider CSD2006-49 (Grant of the Spanish Science Governmental Department)

With the use of genetic technology, researchers have the potential to inform medical diagnoses and treatment in actionable ways. Accurate variant interpretation is a necessary condition for the utility of genetic technology to unfold. This relies on the ability to access large genomic datasets so that comparisons can be made between variants of interest. This can only be successful if DNA and medical data are donated by large numbers of people to 'research', including clinical, non-profit and for-profit research initiatives, in order to be accessed by scientists and clinicians worldwide. The objective of the 'Your DNA, Your Say' global survey is to explore public attitudes, values and opinions towards willingness to donate and concerns regarding the donation of one's personal data for use by others. Using a representative sample of 8967 English-speaking publics from the UK, the USA, Canada and Australia, we explore the characteristics of people who are unwilling (n = 1426) to donate their DNA and medical information, together with an exploration of their reasons. Understanding this perspective is important for making sense of the interaction between science and society. It also helps to focus engagement initiatives on the issues of concern to some publics. ; This work was supported by Wellcome grant [206194] paid to AM, LF, KIM, RM via Wellcome Genome Campus Society and Ethics Research Group, Connecting Science. We would like to thank the following people from GA4GH for their encouragement and infrastructure support: Peter Goodhand, Julia Wilson, Bartha Knoppers. This work was also supported by Global Alliance for Genomics and Health, with their funding delivered via Wellcome (GA4GH grant, with thanks to Audrey Duncansen). DV acknowledges the infrastructure funding received from the Victorian State Government through the Operational Infrastructure Support (OIS) Program.