Aberrant DNA Damage Response Pathways May Predict the Outcome of Platinum Chemotherapy in Ovarian Cancer

Ovarian carcinoma (OC) is the most lethal gynecological malignancy. Despite the advances in the treatment of OC with combinatorial regimens, including surgery and platinum-based chemotherapy, patients generally exhibit poor prognosis due to high chemotherapy resistance. Herein, we tested the hypothesis that DNA damage response (DDR) pathways are involved in resistance of OC patients to platinum chemotherapy. Selected DDR signals were evaluated in two human ovarian carcinoma cell lines, one sensitive (A2780) and one resistant (A2780/C30) to platinum treatment as well as in peripheral blood mononuclear cells (PBMCs) from OC patients, sensitive (n = 7) or resistant (n = 4) to subsequent chemotherapy. PBMCs from healthy volunteers (n = 9) were studied in parallel. DNA damage was evaluated by immunofluorescence γH2AX staining and comet assay. Higher levels of intrinsic DNA damage were found in A2780 than in A2780/C30 cells. Moreover, the intrinsic DNA damage levels were significantly higher in OC patients relative to healthy volunteers, as well as in platinum-sensitive patients relative to platinum-resistant ones (all P<0.05). Following carboplatin treatment, A2780 cells showed lower DNA repair efficiency than A2780/C30 cells. Also, following carboplatin treatment of PBMCs ex vivo, the DNA repair efficiency was significantly higher in healthy volunteers than in platinum-resistant patients and lowest in platinum-sensitive ones (t_1/2 for loss of γH2AX foci: 2.7±0.5h, 8.8±1.9h and 15.4±3.2h, respectively; using comet assay, t_1/2 of platinum-induced damage repair: 4.8±1.4h, 12.9±1.9h and 21.4±2.6h, respectively; all P<0.03). Additionally, the carboplatin-induced apoptosis rate was higher in A2780 than in A2780/C30 cells. In PBMCs, apoptosis rates were inversely correlated with DNA repair efficiencies of these cells, being significantly higher in platinum-sensitive than in platinum-resistant patients and lowest in healthy volunteers (all P<0.05). We conclude that perturbations of DNA repair pathways as measured in PBMCs from OC patients correlate with the drug sensitivity of these cells and reflect the individualized response to platinum-based chemotherapy.     

A new computational method for the detection of horizontal gene transfer events

In recent years, the increase in the amounts of available genomic data has made it easier to appreciate the extent by which organisms increase their genetic diversity through horizontally transferred genetic material. Such transfers have the potential to give rise to extremely dynamic genomes where a significant proportion of their coding DNA has been contributed by external sources. Because of the impact of these horizontal transfers on the ecological and pathogenic character of the recipient organisms, methods are continuously sought that are able to computationally determine which of the genes of a given genome are products of transfer events. In this paper, we introduce and discuss a novel computational method for identifying horizontal transfers that relies on a gene's nucleotide composition and obviates the need for knowledge of codon boundaries. In addition to being applicable to individual genes, the method can be easily extended to the case of clusters of horizontally transferred genes. With the help of an extensive and carefully designed set of experiments on 123 archaeal and bacterial genomes, we demonstrate that the new method exhibits significant improvement in sensitivity when compared to previously published approaches. In fact, it achieves an average relative improvement across genomes of between 11 and 41% compared to the Codon Adaptation Index method in distinguishing native from foreign genes. Our method's horizontal gene transfer predictions for 123 microbial genomes are available online at http://cbcsrv.watson.ibm.com/HGT/.     

A structural constitutive model considering angular dispersion and waviness of collagen fibres of rabbit facial veins

Background  

Structural constitutive models of vascular wall integrate information on composition and structural arrangements of tissue. In blood vessels, collagen fibres are arranged in coiled and wavy bundles and the individual collagen fibres have a deviation from their mean orientation. A complete structural constitutive model for vascular wall should incorporate both waviness and orientational distribution of fibres. We have previously developed a model, for passive properties of vascular wall, which considers the waviness of collagen fibres. However, to our knowledge there is no structural model of vascular wall which integrates both these features.     

H3K27ac bookmarking promotes rapid post-mitotic activation of the pluripotent stem cell program without impacting 3D chromatin reorganization

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Genetic inactivation of the polycomb repressive complex 2 in T cell acute lymphoblastic leukemia

T cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we report the presence of loss-of-function mutations and deletions of the EZH2 and SUZ12 genes, which encode crucial components of the Polycomb repressive complex 2 (PRC2), in 25% of T-ALLs. To further study the role of PRC2 in T-ALL, we used NOTCH1-dependent mouse models of the disease, as well as human T-ALL samples, and combined locus-specific and global analysis of NOTCH1-driven epigenetic changes. These studies demonstrated that activation of NOTCH1 specifically induces loss of the repressive mark Lys27 trimethylation of histone 3 (H3K27me3) by antagonizing the activity of PRC2. These studies suggest a tumor suppressor role for PRC2 in human leukemia and suggest a hitherto unrecognized dynamic interplay between oncogenic NOTCH1 and PRC2 function for the regulation of gene expression and cell transformation.     

Microemulsion-based organogels as matrices for lipase immobilization

Organogels based on water-in-oil microemulsions can be formed using various natural polymers such as gelatin, agar or cellulose derivatives. Enzymes entrapped in the water core of the microemulsion can keep their activity and enhance their stability within the gel matrix. The importance of the microemulsion based organogels (MBGs) leans on their numerous potential biotechnological applications. An important example is the use of various lipase microemulsion systems for hydrolytic or synthetic reactions. In this review, several MBGs are being evaluated as immobilization matrices for various enzymes. The main subject focuses on the parameters that affect the use of MBGs as media for bioorganic reactions using lipases as catalysts.     

InteBac: An integrated bacterial and baculovirus expression vector suite

The successful production of recombinant protein for biochemical, biophysical, and structural biological studies critically depends on the correct expression organism. Currently, the most commonly used expression organisms for structural studies are Escherichia coli (~70% of all PDB structures) and the baculovirus/ insect cell expression system (~5% of all PDB structures). While insect cell expression is frequently successful for large eukaryotic proteins, it is relatively expensive and time‐consuming compared to E. coli expression. Frequently the decision to carry out a baculovirus project means restarting cloning from scratch. Here we describe an integrated system that allows simultaneous cloning into E. coli and baculovirus expression vectors using the same PCR products. The system offers a flexible array of N‐ and C‐terminal affinity, solubilization and utility tags, and the speed allows expression screening to be completed in E. coli, before carrying out time and cost‐intensive experiments in baculovirus. Importantly, we describe a means of rapidly generating polycistronic bacterial constructs based on the hugely successful biGBac system, making InteBac of particular interest for researchers working on recombinant protein complexes.