TherMos: Estimating protein-DNA binding energies from in vivo binding profiles

Accurately characterizing transcription factor (TF)-DNA affinity is a central goal of regulatory genomics. Although thermodynamics provides the most natural language for describing the continuous range of TF-DNA affinity, traditional motif discovery algorithms focus instead on classification paradigms that aim to discriminate 'bound' and 'unbound' sequences. Moreover, these algorithms do not directly model the distribution of tags in ChIP-seq data. Here, we present a new algorithm named Thermodynamic Modeling of ChIP-seq (TherMos), which directly estimates a positionspecific binding energy matrix (PSEM) from ChIPseq/exo tag profiles. In cross-validation tests on seven genome-wide TF-DNA binding profiles, one of which we generated via ChIP-seq on a complex developing tissue, TherMos predicted quantitative TF-DNA binding with greater accuracy than five well-known algorithms. We experimentally validated TherMos binding energy models for Klf4 and Esrrb, using a novel protocol to measure PSEMs in vitro. Strikingly, our measurements revealed strong nonadditivity at multiple positions within the two PSEMs. Among the algorithms tested, only TherMos was able to model the entire binding energy landscape of Klf4 and Esrrb. Our study reveals new insights into the energetics of TF-DNA binding in vivo and provides an accurate first-principles approach to binding energy inference from ChIP-seq and ChIP-exo data. © 2013 The Author(s).Link_to_subscribed_fulltex

Digital Signal Processing

Contains research objectives and summary of research on seven research projects.Joint Services Electronics Program (Contract DAAB07-76-C-1400)U. S. Navy - Office of Naval Research (Contract N00014-75-C-0951-NR 049-308)National Science Foundation (Grant ENG71-02319-AO2

An initial event in insect innate immune response: structural and biological studies of interactions between β-1,3-glucan and the N-terminal domain of β-1,3-glucan recognition protein

In response to invading microorganisms, insect β-1,3-glucan recognition protein (βGRP), a soluble receptor in the hemolymph, binds to the surfaces of bacteria and fungi and activates serine protease cascades that promote destruction of pathogens by means of melanization or expression of antimicrobial peptides. Here we report on the NMR solution structure of the N-terminal domain of βGRP (N-βGRP) from Indian meal moth (Plodia interpunctella), which is sufficient to activate the prophenoloxidase (proPO) pathway resulting in melanin formation. NMR and isothermal calorimetric titrations of N-βGRP with laminarihexaose, a glucose hexamer containing β-1,3 links, suggest a weak binding of the ligand. However, addition of laminarin, a glucose polysaccharide (~ 6 kDa) containing β-1,3 and β-1,6 links that activates the proPO pathway, to N-βGRP results in the loss of NMR cross-peaks from the backbone 15N-1H groups of the protein, suggesting the formation of a large complex. Analytical ultra centrifugation (AUC) studies of formation of N-βGRP:laminarin complex show that ligand-binding induces sel-fassociation of the protein:carbohydrate complex into a macro structure, likely containing six protein and three laminarin molecules (~ 102 kDa). The macro complex is quite stable, as it does not undergo dissociation upon dilution to sub-micromolar concentrations. The structural model thus derived from the present studies for N-βGRP:laminarin complex in solution differs from the one in which a single N-βGRP molecule has been proposed to bind to a triple helical form of laminarin on the basis of an X-ray crystallographic structure of N-βGRP:laminarihexaose complex [Kanagawa, M., Satoh, T., Ikeda, A., Adachi, Y., Ohno, N., and Yamaguchi, Y. (2011) J. Biol. Chem. 286, 29158-29165]. AUC studies and phenoloxidase activation measurements carried out with the designed mutants of N-βGRP indicate that electrostatic interactions involving Asp45, Arg54, and Asp68 between the ligand-bound protein molecules contribute in part to the stability of N-βGRP:laminarin macro complex and that a decreased stability is accompanied by a reduced activation of the proPO pathway. Increased β-1,6 branching in laminarin also results in destabilization of the macro complex. These novel findings suggest that ligand-induced self-association of βGRP:β-1,3-glucan complex may form a platform on a microbial surface for recruitment of downstream proteases, as a means of amplification of the initial signal of pathogen recognition for the activation of the proPO pathway

Exposure to Mycobacterium remodels alveolar macrophages and the early innate response to Mycobacterium tuberculosis infection

Alveolar macrophages (AMs) play a critical role during Mycobacterium tuberculosis (Mtb) infection as the first cells in the lung to encounter bacteria. We previously showed that AMs initially respond to Mtb in vivo by mounting a cell-protective, rather than pro-inflammatory response. However, the plasticity of the initial AM response was unknown. Here, we characterize how previous exposure to Mycobacterium, either through subcutaneous vaccination with Mycobacterium bovis (scBCG) or through a contained Mtb infection (coMtb) that mimics aspects of concomitant immunity, impacts the initial response by AMs. We find that both scBCG and coMtb accelerate early innate cell activation and recruitment and generate a stronger pro-inflammatory response to Mtb in vivo by AMs. Within the lung environment, AMs from scBCG vaccinated mice mount a robust interferon-associated response, while AMs from coMtb mice produce a broader inflammatory response that is not dominated by Interferon Stimulated Genes. Using scRNAseq, we identify changes to the frequency and phenotype of airway-resident macrophages following Mycobacterium exposure, with enrichment for both interferon-associated and pro-inflammatory populations of AMs. In contrast, minimal changes were found for airway-resident T cells and dendritic cells after exposures. Ex vivo stimulation of AMs with Pam3Cys, LPS and Mtb reveal that scBCG and coMtb exposures generate stronger interferon-associated responses to LPS and Mtb that are cell-intrinsic changes. However, AM profiles that were unique to each exposure modality following Mtb infection in vivo are dependent on the lung environment and do not emerge following ex vivo stimulation. Overall, our studies reveal significant and durable remodeling of AMs following exposure to Mycobacterium, with evidence for both AM-intrinsic changes and contributions from the altered lung microenvironments. Comparisons between the scBCG and coMtb models highlight the plasticity of AMs in the airway and opportunities to target their function through vaccination or host-directed therapies

Genetic origins of honey bees (Apis mellifera) on Kangaroo Island and Norfolk Island (Australia) and the Kingdom of Tonga

International audienceAbstractWe examine the origin of honey bee (Apis mellifera) populations in Kangaroo Island (Australia), Norfolk Island (Australia) and the Kingdom of Tonga using a highly polymorphic mitochondrial DNA region and a panel of 37 single nucleotide polymorphisms that assigns ancestry to three evolutionary lineages: Eastern Europe, Western Europe and Africa. We also examine inbreeding coefficients and genetic variation using microsatellites and mitochondrial sequencing. The honey bees of Kangaroo Island have a high proportion of Eastern European ancestry (90.2%), consistent with claims that they are of the subspecies A. m. ligustica. The honey bees of Norfolk Island also had a majority of ancestry from Eastern Europe (73.1%) with some contribution from Western Europe (21.2%). The honey bees of Tonga are mainly of Western European (70.3%) origin with some Eastern European ancestry (27.4%). Despite the suspected severe bottlenecks experienced by these island population, inbreeding coefficients were low

Bounds on the width, mass difference and other properties of X(3872) --> pi+pi-J/psi decays

We present results from a study of X(3872) --> pi+pi- J/psi decays produced via exclusive B--> K X(3872) decays. We determine the mass to be M_X(3872)= (3871.84\pm 0.27 (stat)\pm 0.19 (syst)) MeV, a 90% CL upper limit on the natural width of Gamma_X(3872) K+X(3872))xBf(X(3872)-->pi+pi-J/psi)=(8.61 \pm 0.82(stat) \pm 0.52 (syst)) x10^{-6}, and a ratio of branching fractions Bf(B0--> K0 X(3872))/BF(B+--> K+ X(3872))=0.50\pm 0.14(stat)\pm0.04(syst). The difference in mass between the X(3872)-->pi+pi-J/psi signals in B+ and B0 decays is Delta M_{X(3872)= (-0.69 \pm 0.97 (stat)} \pm 0.19 (syst)) MeV. A search for a charged partner of the X(3872) in the decays Bbar0-->K- X+ or B+-->K0X+, X+-->pi+pi0 J/psi resulted in upper limits on the product branching fractions for these processes that are well below expectations for the case that the X(3872) is the neutral member of an isospin triplet. In addition, we examine possible J^{PC} quantum number assignments for the X(3872) based on comparisons of angular correlations between final state particles in X(3872)-->pi+pi-J/psi decays with simulated data for J^{PC} values of 1^{++} and 2^{-+}. We examine the influence of rho-omega interference in the M(pi+pi-) spectrum. The analysis is based on a 711fb^{-1} data sample that contains 772 million BBbar meson pairs collected at the Upsilon(4S) resonance in the Belle detector at the KEKB e+e- collider.Comment: 15 pages, 10 figures and 6 tables. Submitted to Physical Review

Management of asthma in pregnant women by general practitioners: A cross sectional survey

<p>Abstract</p> <p>Background</p> <p>Poorly controlled asthma can lead to maternal and fetal complications. Despite the known risks of poorly controlled asthma during pregnancy and the need for stepping up therapy when appropriate, there are concerns that management is suboptimal in primary care.</p> <p>Our objective was to investigate the management of asthma during pregnancy by general practitioners providing shared maternity care.</p> <p>Methods</p> <p>A pre-piloted, anonymous mail survey was sent to all general practitioners (n = 842) involved in shared maternity care at six maternity hospitals in Victoria, Australia. Respondents were asked about their perceived safety of individual asthma medications during pregnancy. Approach to asthma management during pregnancy was further explored using scenarios of pregnant women with stable and deteriorating asthma and poor medication adherence.</p> <p>Results</p> <p>Inhaled corticosteroids (ICS) were perceived to be the safest and were the preferred preventive medication in first trimester (74.1%), whilst leukotriene receptor antagonists were the least preferred (2.9%). A quarter (25.8%) of respondents would stop or decrease patients' ICS doses during pregnancy, even when their asthma was well controlled by current therapy. In addition, 12.1% of respondents were not sure how to manage deteriorating asthma during pregnancy and opted to refer to another health professional. Almost half the respondents (48.9%) reported encountering medication nonadherence during pregnancy.</p> <p>Conclusion</p> <p>A lack of confidence and/or knowledge among general practitioners in managing deteriorating asthma in pregnancy was observed despite a good understanding of the safety of asthma medications during pregnancy, compliance with evidence-based guidelines in the selection of preventive medications, and self reported good asthma knowledge.</p

Evolutionary Toggling of Vpx/Vpr Specificity Results in Divergent Recognition of the Restriction Factor SAMHD1

SAMHD1 is a host restriction factor that blocks the ability of lentiviruses such as HIV-1 to undergo reverse transcription in myeloid cells and resting T-cells. This restriction is alleviated by expression of the lentiviral accessory proteins Vpx and Vpr (Vpx/Vpr), which target SAMHD1 for proteasome-mediated degradation. However, the precise determinants within SAMHD1 for recognition by Vpx/Vpr remain unclear. Here we show that evolution of Vpx/Vpr in primate lentiviruses has caused the interface between SAMHD1 and Vpx/Vpr to alter during primate lentiviral evolution. Using multiple HIV-2 and SIV Vpx proteins, we show that Vpx from the HIV-2 and SIVmac lineage, but not Vpx from the SIVmnd2 and SIVrcm lineage, require the C-terminus of SAMHD1 for interaction, ubiquitylation, and degradation. On the other hand, the N-terminus of SAMHD1 governs interactions with Vpx from SIVmnd2 and SIVrcm, but has little effect on Vpx from HIV-2 and SIVmac. Furthermore, we show here that this difference in SAMHD1 recognition is evolutionarily dynamic, with the importance of the N- and C-terminus for interaction of SAMHD1 with Vpx and Vpr toggling during lentiviral evolution. We present a model to explain how the head-to-tail conformation of SAMHD1 proteins favors toggling of the interaction sites by Vpx/Vpr during this virus-host arms race. Such drastic functional divergence within a lentiviral protein highlights a novel plasticity in the evolutionary dynamics of viral antagonists for restriction factors during lentiviral adaptation to its hosts. © 2013 Fregoso et al