Exploring the loblolly pine (Pinus taeda L.) genome by BAC sequencing and Cot analysis.

Loblolly pine (LP; Pinus taeda L.) is an economically and ecologically important tree in the southeastern U.S. To advance understanding of the loblolly pine (LP; Pinus taeda L.) genome, we sequenced and analyzed 100 BAC clones and performed a Cot analysis. The Cot analysis indicates that the genome is composed of 57, 24, and 10% highly-repetitive, moderately-repetitive, and single/low-copy sequences, respectively (the remaining 9% of the genome is a combination of fold back and damaged DNA). Although single/low-copy DNA only accounts for 10% of the LP genome, the amount of single/low-copy DNA in LP is still 14 times the size of the Arabidopsis genome. Since gene numbers in LP are similar to those in Arabidopsis, much of the single/low-copy DNA of LP would appear to be composed of DNA that is both gene- and repeat-poor. Macroarrays prepared from a LP bacterial artificial chromosome (BAC) library were hybridized with probes designed from cell wall synthesis/wood development cDNAs, and 50 of the "targeted" clones were selected for further analysis. An additional 25 clones were selected because they contained few repeats, while 25 more clones were selected at random. The 100 BAC clones were Sanger sequenced and assembled. Of the targeted BACs, 80% contained all or part of the cDNA used to target them. One targeted BAC was found to contain fungal DNA and was eliminated from further analysis. Combinations of similarity-based and ab initio gene prediction approaches were utilized to identify and characterize potential coding regions in the 99 BACs containing LP DNA. From this analysis, we identified 154 gene models (GMs) representing both putative protein-coding genes and likely pseudogenes. Ten of the GMs (all of which were specifically targeted) had enough support to be classified as intact genes. Interestingly, the 154 GMs had statistically indistinguishable (α = 0.05) distributions in the targeted and random BAC clones (15.18 and 12.61 GM/Mb, respectively), whereas the low-repeat BACs contained significantly fewer GMs (7.08 GM/Mb). However, when GM length was considered, the targeted BACs had a significantly greater percentage of their length in GMs (3.26%) when compared to random (1.63%) and low-repeat (0.62%) BACs. The results of our study provide insight into LP evolution and inform ongoing efforts to produce a reference genome sequence for LP, while characterization of genes involved in cell wall production highlights carbon metabolism pathways that can be leveraged for increasing wood production

Informational Gene Phylogenies Do Not Support a Fourth Domain of Life for Nucleocytoplasmic Large DNA Viruses

Mimivirus is a nucleocytoplasmic large DNA virus (NCLDV) with a genome size (1.2 Mb) and coding capacity ( 1000 genes) comparable to that of some cellular organisms. Unlike other viruses, Mimivirus and its NCLDV relatives encode homologs of broadly conserved informational genes found in Bacteria, Archaea, and Eukaryotes, raising the possibility that they could be placed on the tree of life. A recent phylogenetic analysis of these genes showed the NCLDVs emerging as a monophyletic group branching between Eukaryotes and Archaea. These trees were interpreted as evidence for an independent “fourth domain” of life that may have contributed DNA processing genes to the ancestral eukaryote. However, the analysis of ancient evolutionary events is challenging, and tree reconstruction is susceptible to bias resulting from non-phylogenetic signals in the data. These include compositional heterogeneity and homoplasy, which can lead to the spurious grouping of compositionally-similar or fast-evolving sequences. Here, we show that these informational gene alignments contain both significant compositional heterogeneity and homoplasy, which were not adequately modelled in the original analysis. When we use more realistic evolutionary models that better fit the data, the resulting trees are unable to reject a simple null hypothesis in which these informational genes, like many other NCLDV genes, were acquired by horizontal transfer from eukaryotic hosts. Our results suggest that a fourth domain is not required to explain the available sequence data

Protective Immunity Induced with the RTS,S/AS Vaccine Is Associated with IL-2 and TNF-α Producing Effector and Central Memory CD4+ T Cells

A phase 2a RTS,S/AS malaria vaccine trial, conducted previously at the Walter Reed Army Institute of Research, conferred sterile immunity against a primary challenge with infectious sporozoites in 40% of the 80 subjects enrolled in the study. The frequency of Plasmodium falciparum circumsporozoite protein (CSP)-specific CD4+ T cells was significantly higher in protected subjects as compared to non-protected subjects. Intrigued by these unique vaccine-related correlates of protection, in the present study we asked whether RTS,S also induced effector/effector memory (TE/EM) and/or central memory (TCM) CD4+ T cells and whether one or both of these sub-populations is the primary source of cytokine production. We showed for the first time that PBMC from malaria-non-exposed RTS,S-immunized subjects contain both TE/EM and TCM cells that generate strong IL-2 responses following re-stimulation in vitro with CSP peptides. Moreover, both the frequencies and the total numbers of IL-2-producing CD4+ TE/EM cells and of CD4+ TCM cells from protected subjects were significantly higher than those from non-protected subjects. We also demonstrated for the first time that there is a strong association between the frequency of CSP peptide-reactive CD4+ T cells producing IL-2 and the titers of CSP-specific antibodies in the same individual, suggesting that IL-2 may be acting as a growth factor for follicular Th cells and/or B cells. The frequencies of CSP peptide-reactive, TNF-α-producing CD4+ TE/EM cells and of CD4+ TE/EM cells secreting both IL-2 and TNF-α were also shown to be higher in protected vs. non-protected individuals. We have, therefore, demonstrated that in addition to TNF-α, IL-2 is also a significant contributing factor to RTS,S/AS vaccine induced immunity and that both TE/EM and TCM cells are major producers of IL-2

Blood Signature of Pre-Heart Failure: A Microarrays Study

International audienceBACKGROUND: The preclinical stage of systolic heart failure (HF), known as asymptomatic left ventricular dysfunction (ALVD), is diagnosed only by echocardiography, frequent in the general population and leads to a high risk of developing severe HF. Large scale screening for ALVD is a difficult task and represents a major unmet clinical challenge that requires the determination of ALVD biomarkers. METHODOLOGY/PRINCIPAL FINDINGS: 294 individuals were screened by echocardiography. We identified 9 ALVD cases out of 128 subjects with cardiovascular risk factors. White blood cell gene expression profiling was performed using pangenomic microarrays. Data were analyzed using principal component analysis (PCA) and Significant Analysis of Microarrays (SAM). To build an ALVD classifier model, we used the nearest centroid classification method (NCCM) with the ClaNC software package. Classification performance was determined using the leave-one-out cross-validation method. Blood transcriptome analysis provided a specific molecular signature for ALVD which defined a model based on 7 genes capable of discriminating ALVD cases. Analysis of an ALVD patients validation group demonstrated that these genes are accurate diagnostic predictors for ALVD with 87% accuracy and 100% precision. Furthermore, Receiver Operating Characteristic curves of expression levels confirmed that 6 out of 7 genes discriminate for left ventricular dysfunction classification. CONCLUSIONS/SIGNIFICANCE: These targets could serve to enhance the ability to efficiently detect ALVD by general care practitioners to facilitate preemptive initiation of medical treatment preventing the development of HF

Rationale and methods of the European Study on Cardiovascular Risk Prevention and Management in Daily Practice (EURIKA)

<p>Abstract</p> <p>Background</p> <p>The EURIKA study aims to assess the status of primary prevention of cardiovascular disease (CVD) across Europe. Specifically, it will determine the degree of control of cardiovascular risk factors in current clinical practice in relation to the European guidelines on cardiovascular prevention. It will also assess physicians' knowledge and attitudes about CVD prevention as well as the barriers impeding effective risk factor management in clinical practice.</p> <p>Methods/Design</p> <p>Cross-sectional study conducted simultaneously in 12 countries across Europe. The study has two components: firstly at the physician level, assessing eight hundred and nine primary care and specialist physicians with a daily practice in CVD prevention. A physician specific questionnaire captures information regarding physician demographics, practice settings, cardiovascular prevention beliefs and management. Secondly at the patient level, including 7641 patients aged 50 years or older, free of clinical CVD and with at least one classical risk factor, enrolled by the participating physicians. A patient-specific questionnaire captures information from clinical records and patient interview regarding sociodemographic data, CVD risk factors, and current medications. Finally, each patient provides a fasting blood sample, which is sent to a central laboratory for measuring serum lipids, apolipoproteins, hemoglobin-A1c, and inflammatory biomarkers.</p> <p>Discussion</p> <p>Primary prevention of CVD is an extremely important clinical issue, with preventable circulatory diseases remaining the leading cause of major disease burden. The EURIKA study will provide key information to assess effectiveness of and attitudes toward primary prevention of CVD in Europe. A transnational study creates opportunities for benchmarking good clinical practice across countries and improving outcomes. (ClinicalTrials.gov number, NCT00882336.)</p

Armadillo 1.1: An Original Workflow Platform for Designing and Conducting Phylogenetic Analysis and Simulations

In this paper we introduce Armadillo v1.1, a novel workflow platform dedicated to designing and conducting phylogenetic studies, including comprehensive simulations. A number of important phylogenetic and general bioinformatics tools have been included in the first software release. As Armadillo is an open-source project, it allows scientists to develop their own modules as well as to integrate existing computer applications. Using our workflow platform, different complex phylogenetic tasks can be modeled and presented in a single workflow without any prior knowledge of programming techniques. The first version of Armadillo was successfully used by professors of bioinformatics at Université du Quebec à Montreal during graduate computational biology courses taught in 2010–11. The program and its source code are freely available at: <http://www.bioinfo.uqam.ca/armadillo>

Induction of Plasmodium falciparum-Specific CD4+ T Cells and Memory B Cells in Gabonese Children Vaccinated with RTS,S/AS01E and RTS,S/AS02D

The recombinant circumsporozoite protein (CS) based vaccine, RTS,S, confers protection against Plasmodium falciparum infection in controlled challenge trials and in field studies. The RTS,S recombinant antigen has been formulated with two adjuvant systems, AS01 and AS02, which have both been shown to induce strong specific antibody responses and CD4 T cell responses in adults. As infants and young children are particularly susceptible to malaria infection and constitute the main target population for a malaria vaccine, we have evaluated the induction of adaptive immune responses in young children living in malaria endemic regions following vaccination with RTS,S/AS01(E) and RTS,S/AS02(D). Our data show that a CS-specific memory B cell response is induced one month after the second and third vaccine dose and that CS-specific antibodies and memory B cells persist up to 12 months after the last vaccine injection. Both formulations also induced low but significant amounts of CS-specific IL-2(+) CD4(+) T cells one month after the second and third vaccine dose, upon short-term in vitro stimulation of whole blood cells with peptides covering the entire CS derived sequence in RTS,S. These results provide evidence that both RTS,S/AS01(E) and RTS,S/AS02(D) induced adaptive immune responses including antibodies, circulating memory B cells and CD4(+) T cells directed against P. falciparum CS protein.ClinicalTrials.gov NCT00307021

Induction of GADD34 Is Necessary for dsRNA-Dependent Interferon-β Production and Participates in the Control of Chikungunya Virus Infection

Nucleic acid sensing by cells is a key feature of antiviral responses, which generally result in type-I Interferon production and tissue protection. However, detection of double-stranded RNAs in virus-infected cells promotes two concomitant and apparently conflicting events. The dsRNA-dependent protein kinase (PKR) phosphorylates translation initiation factor 2-alpha (eIF2α) and inhibits protein synthesis, whereas cytosolic DExD/H box RNA helicases induce expression of type I-IFN and other cytokines. We demonstrate that the phosphatase-1 cofactor, growth arrest and DNA damage-inducible protein 34 (GADD34/Ppp1r15a), an important component of the unfolded protein response (UPR), is absolutely required for type I-IFN and IL-6 production by mouse embryonic fibroblasts (MEFs) in response to dsRNA. GADD34 expression in MEFs is dependent on PKR activation, linking cytosolic microbial sensing with the ATF4 branch of the UPR. The importance of this link for anti-viral immunity is underlined by the extreme susceptibility of GADD34-deficient fibroblasts and neonate mice to Chikungunya virus infection

The human phylome

The human phylome, which includes evolutionary relationships of all human proteins and their homologs among thirty-nine fully sequenced eukaryotes, is reconstructed