Estimating the survival benefits gained from providing national cancer genetic services to women with a family history of breast cancer

The aim of this paper is to compare a service offering genetic testing and presymptomatic surveillance to women at increased risk of developing breast cancer with its predecessor of no service at all in terms of survival and quality-adjusted survival (QALYs) by means of a Markov cohort chain simulation model. Genetic assessment and presymptomatic care provided between 0.07-1.61 mean additional life years and 0.05-1.67 mean QALYs over no services. Prophylactic surgery and surveillance extended mean life expectancy by 0.41-1.61 and 0.32-0.99 years, respectively over no services for high-risk women. Model outcomes were sensitive to all the parameters varied in the sensitivity analysis. Providing cancer genetic services increase survival and as long as services do not induce adverse psychological effects they also provide more QALYs. The greatest survival and QALY benefits were found for women with identified mutations. As more cancer genes are identified, the survival and cost-effectiveness of genetic services will improve. Although mastectomy provided most additional life years, when quality of life was accounted for oophorectomy was the optimal strategy. Delayed entry into coordinated genetic services was found to diminish the average survival and QALY gains for a woman utilising these services

Phenotypic Signatures Arising from Unbalanced Bacterial Growth

Fluctuations in the growth rate of a bacterial culture during unbalanced growth are generally considered undesirable in quantitative studies of bacterial physiology. Under well-controlled experimental conditions, however, these fluctuations are not random but instead reflect the interplay between intra-cellular networks underlying bacterial growth and the growth environment. Therefore, these fluctuations could be considered quantitative phenotypes of the bacteria under a specific growth condition. Here, we present a method to identify “phenotypic signatures” by time-frequency analysis of unbalanced growth curves measured with high temporal resolution. The signatures are then applied to differentiate amongst different bacterial strains or the same strain under different growth conditions, and to identify the essential architecture of the gene network underlying the observed growth dynamics. Our method has implications for both basic understanding of bacterial physiology and for the classification of bacterial strains

GC-Rich Sequence Elements Recruit PRC2 in Mammalian ES Cells

Polycomb proteins are epigenetic regulators that localize to developmental loci in the early embryo where they mediate lineage-specific gene repression. In Drosophila, these repressors are recruited to sequence elements by DNA binding proteins associated with Polycomb repressive complex 2 (PRC2). However, the sequences that recruit PRC2 in mammalian cells have remained obscure. To address this, we integrated a series of engineered bacterial artificial chromosomes into embryonic stem (ES) cells and examined their chromatin. We found that a 44 kb region corresponding to the Zfpm2 locus initiates de novo recruitment of PRC2. We then pinpointed a CpG island within this locus as both necessary and sufficient for PRC2 recruitment. Based on this causal demonstration and prior genomic analyses, we hypothesized that large GC-rich elements depleted of activating transcription factor motifs mediate PRC2 recruitment in mammals. We validated this model in two ways. First, we showed that a constitutively active CpG island is able to recruit PRC2 after excision of a cluster of activating motifs. Second, we showed that two 1 kb sequence intervals from the Escherichia coli genome with GC-contents comparable to a mammalian CpG island are both capable of recruiting PRC2 when integrated into the ES cell genome. Our findings demonstrate a causal role for GC-rich sequences in PRC2 recruitment and implicate a specific subset of CpG islands depleted of activating motifs as instrumental for the initial localization of this key regulator in mammalian genomes.Burroughs Wellcome FundCharles E. Culpeper FoundationMassachusetts General HospitalBroad Institute of MIT and Harvar

Use of DNA technology in forensic dentistry

The established importance of Forensic Dentistry for human identification, mainly when there is little remaining material to perform such identification (e.g., in fires, explosions, decomposing bodies or skeletonized bodies), has led dentists working with forensic investigation to become more familiar with the new molecular biology techniques. The currently available DNA tests have high reliability and are accepted as legal proofs in courts. This article presents a literature review referring to the main studies on Forensic Dentistry that involve the use of DNA for human identification, and makes an overview of the evolution of this technology in the last years, highlighting the importance of molecular biology in forensic sciences

Role of Mitofusin 2 in the Renal Stress Response

The role of mitofusin 2 (MFN2), a key regulator of mitochondrial morphology and function in the renal stress response is unknown. To assess its role, the MFN2 floxed gene was conditionally deleted in the kidney of mice (MFN2 cKO) by Pax2 promoter driven Cre expression (Pax2Cre). MFN2 cKO caused severe mitochondrial fragmentation in renal epithelial cells that are critical for normal kidney tubular function. However, despite a small (20%) decrease in nephron number, newborn cKO pups had organ or tubular function that did not differ from littermate Cre-negative pups. MFN2 deficiency in proximal tubule epithelial cells in primary culture induced mitochondrial fragmentation but did not significantly alter ATP turnover, maximal mitochondrial oxidative reserve capacity, or the low level of oxygen consumption during cyanide exposure. MFN2 deficiency also did not increase apoptosis of tubule epithelial cells under non-stress conditions. In contrast, metabolic stress caused by ATP depletion exacerbated mitochondrial outer membrane injury and increased apoptosis by 80% in MFN2 deficient vs. control cells. Despite similar stress-induced Bax 6A7 epitope exposure in MFN2 deficient and control cells, MFN2 deficiency significantly increased mitochondrial Bax accumulation and was associated with greater release of both apoptosis inducing factor and cytochrome c. In conclusion, MFN2 deficiency in the kidney causes mitochondrial fragmentation but does not affect kidney or tubular function during development or under non-stress conditions. However, MFN2 deficiency exacerbates renal epithelial cell injury by promoting Bax-mediated mitochondrial outer membrane injury and apoptosis

Nuclear Pore Proteins Nup153 and Megator Define Transcriptionally Active Regions in the Drosophila Genome

Transcriptional regulation is one of the most important processes for modulating gene expression. Though much of this control is attributed to transcription factors, histones, and associated enzymes, it is increasingly apparent that the spatial organization of chromosomes within the nucleus has a profound effect on transcriptional activity. Studies in yeast indicate that the nuclear pore complex might promote transcription by recruiting chromatin to the nuclear periphery. In higher eukaryotes, however, it is not known whether such regulation has global significance. Here we establish nucleoporins as a major class of global regulators for gene expression in Drosophila melanogaster. Using chromatin-immunoprecipitation combined with microarray hybridisation, we show that Nup153 and Megator (Mtor) bind to 25% of the genome in continuous domains extending 10 kb to 500 kb. These Nucleoporin-Associated Regions (NARs) are dominated by markers for active transcription, including high RNA polymerase II occupancy and histone H4K16 acetylation. RNAi–mediated knock-down of Nup153 alters the expression of ∼5,700 genes, with a pronounced down-regulatory effect within NARs. We find that nucleoporins play a central role in coordinating dosage compensation—an organism-wide process involving the doubling of expression of the male X chromosome. NARs are enriched on the male X chromosome and occupy 75% of this chromosome. Furthermore, Nup153-depletion abolishes the normal function of the male-specific dosage compensation complex. Finally, by extensive 3D imaging, we demonstrate that NARs contribute to gene expression control irrespective of their sub-nuclear localization. Therefore, we suggest that NAR–binding is used for chromosomal organization that enables gene expression control

Fluorescence activated cell sorting followed by small RNA sequencing reveals stable microRNA expression during cell cycle progression.

BACKGROUND: Previously, drug-based synchronization procedures were used for characterizing the cell cycle dependent transcriptional program. However, these synchronization methods result in growth imbalance and alteration of the cell cycle machinery. DNA content-based fluorescence activated cell sorting (FACS) is able to sort the different cell cycle phases without perturbing the cell cycle. MiRNAs are key transcriptional regulators of the cell cycle, however, their expression dynamics during cell cycle has not been explored. METHODS: Following an optimized FACS, a complex initiative of high throughput platforms (microarray, Taqman Low Density Array, small RNA sequencing) were performed to study gene and miRNA expression profiles of cell cycle sorted human cells originating from different tissues. Validation of high throughput data was performed using quantitative real time PCR. Protein expression was detected by Western blot. Complex statistics and pathway analysis were also applied. RESULTS: Beyond confirming the previously described cell cycle transcriptional program, cell cycle dependently expressed genes showed a higher expression independently from the cell cycle phase and a lower amplitude of dynamic changes in cancer cells as compared to untransformed fibroblasts. Contrary to mRNA changes, miRNA expression was stable throughout the cell cycle. CONCLUSIONS: Cell cycle sorting is a synchronization-free method for the proper analysis of cell cycle dynamics. Altered dynamic expression of universal cell cycle genes in cancer cells reflects the transformed cell cycle machinery. Stable miRNA expression during cell cycle progression may suggest that dynamical miRNA-dependent regulation may be of less importance in short term regulations during the cell cycle

Clotrimazole Preferentially Inhibits Human Breast Cancer Cell Proliferation, Viability and Glycolysis

BACKGROUND: Clotrimazole is an azole derivative with promising anti-cancer effects. This drug interferes with the activity of glycolytic enzymes altering their cellular distribution and inhibiting their activities. The aim of the present study was to analyze the effects of clotrimazole on the growth pattern of breast cancer cells correlating with their metabolic profiles. METHODOLOGY/PRINCIPAL FINDINGS: Three cell lines derived from human breast tissue (MCF10A, MCF-7 and MDA-MB-231) that present increasingly aggressive profiles were used. Clotrimazole induces a dose-dependent decrease in glucose uptake in all three cell lines, with K(i) values of 114.3±11.7, 77.1±7.8 and 37.8±4.2 µM for MCF10A, MCF-7 and MDA-MB-231, respectively. Furthermore, the drug also decreases intracellular ATP content and inhibits the major glycolytic enzymes, hexokinase, phosphofructokinase-1 and pyruvate kinase, especially in the highly metastatic cell line, MDA-MB-231. In this last cell lineage, clotrimazole attenuates the robust migratory response, an effect that is progressively attenuated in MCF-7 and MCF10A, respectively. Moreover, clotrimazole reduces the viability of breast cancer cells, which is more pronounced on MDA-MB-231. CONCLUSIONS/SIGNIFICANCE: Clotrimazole presents deleterious effects on two human breast cancer cell lines metabolism, growth and migration, where the most aggressive cell line is more affected by the drug. Moreover, clotrimazole presents little or no effect on a non-tumor human breast cell line. These results suggest, at least for these three cell lines studied, that the more aggressive the cell is the more effective clotrimazole is