Smc5/6 coordinates formation and resolution of joint molecules with chromosome morphology to ensure meiotic divisions

During meiosis, Structural Maintenance of Chromosome (SMC) complexes underpin two fundamental features of meiosis: homologous recombination and chromosome segregation. While meiotic functions of the cohesin and condensin complexes have been delineated, the role of the third SMC complex, Smc5/6, remains enigmatic. Here we identify specific, essential meiotic functions for the Smc5/6 complex in homologous recombination and the regulation of cohesin. We show that Smc5/6 is enriched at centromeres and cohesin-association sites where it regulates sister-chromatid cohesion and the timely removal of cohesin from chromosomal arms, respectively. Smc5/6 also localizes to recombination hotspots, where it promotes normal formation and resolution of a subset of joint-molecule intermediates. In this regard, Smc5/6 functions independently of the major crossover pathway defined by the MutLγ complex. Furthermore, we show that Smc5/6 is required for stable chromosomal localization of the XPF-family endonuclease, Mus81-Mms4Eme1. Our data suggest that the Smc5/6 complex is required for specific recombination and chromosomal processes throughout meiosis and that in its absence, attempts at cell division with unresolved joint molecules and residual cohesin lead to severe recombination-induced meiotic catastroph

Expression of Regulatory Platelet MicroRNAs in Patients with Sickle Cell Disease

Background: Increased platelet activation in sickle cell disease (SCD) contributes to a state of hypercoagulability and confers a risk of thromboembolic complications. The role for post-transcriptional regulation of the platelet transcriptome by microRNAs (miRNAs) in SCD has not been previously explored. This is the first study to determine whether platelets from SCD exhibit an altered miRNA expression profile. Methods and Findings: We analyzed the expression of miRNAs isolated from platelets from a primary cohort (SCD = 19, controls = 10) and a validation cohort (SCD = 7, controls = 7) by hybridizing to the Agilent miRNA microarrays. A dramatic difference in miRNA expression profiles between patients and controls was noted in both cohorts separately. A total of 40 differentially expressed platelet miRNAs were identified as common in both cohorts (p-value 0.05, fold change>2) with 24 miRNAs downregulated. Interestingly, 14 of the 24 downregulated miRNAs were members of three families - miR-329, miR-376 and miR-154 - which localized to the epigenetically regulated, maternally imprinted chromosome 14q32 region. We validated the downregulated miRNAs, miR-376a and miR-409-3p, and an upregulated miR-1225-3p using qRT-PCR. Over-expression of the miR-1225-3p in the Meg01 cells was followed by mRNA expression profiling to identify mRNA targets. This resulted in significant transcriptional repression of 1605 transcripts. A combinatorial approach using Meg01 mRNA expression profiles following miR-1225-3p overexpression, a computational prediction analysis of miRNA target sequences and a previously published set of differentially expressed platelet transcripts from SCD patients, identified three novel platelet mRNA targets: PBXIP1, PLAGL2 and PHF20L1. Conclusions: We have identified significant differences in functionally active platelet miRNAs in patients with SCD as compared to controls. These data provide an important inventory of differentially expressed miRNAs in SCD patients and an experimental framework for future studies of miRNAs as regulators of biological pathways in platelets. © 2013 Jain et al

The chemopreventive polyphenol Curcumin prevents hematogenous breast cancer metastases in immunodeficient mice

Dissemination of metastatic cells probably occurs long before diagnosis of the primary tumor. Metastasis during early phases of carcinogenesis in high risk patients is therefore a potential prevention target. The plant polyphenol Curcumin has been proposed for dietary prevention of cancer. We therefore examined its effects on the human breast cancer cell line MDA-MB-231 in vitro and in a mouse metastasis model. Curcumin strongly induces apoptosis in MDA- MB- 231 cells in correlation with reduced activation of the survival pathway NF kappa B, as a consequence of diminished I kappa B and p65 phosphorylation. Curcumin also reduces the expression of major matrix metalloproteinases (MMPs) due to reduced NF kappa B activity and transcriptional downregulation of AP-1. NF kappa B/p65 silencing is sufficient to downregulate c-jun and MMP expression. Reduced NF kappa B/AP-1 activity and MMP expression lead to diminished invasion through a reconstituted basement membrane and to a significantly lower number of lung metastases in immunodeficient mice after intercardiac injection of 231 cells (p=0.0035). 68% of Curcumin treated but only 17% of untreated animals showed no or very few lung metastases, most likely as a consequence of down-regulation of NF kappa B/AP-1 dependent MMP expression and direct apoptotic effects on circulating tumor cells but not on established metastases. Dietary chemoprevention of metastases appears therefore feasible. Copyright (c) 2007 S. Karger AG, Basel

Quantification of Rapid Myosin Regulatory Light Chain Phosphorylation Using High-Throughput In-Cell Western Assays: Comparison to Western Immunoblots

Quantification of phospho-proteins (PPs) is crucial when studying cellular signaling pathways. Western immunoblotting (WB) is commonly used for the measurement of relative levels of signaling intermediates in experimental samples. However, WB is in general a labour-intensive and low-throughput technique. Because of variability in protein yield and phospho-signal preservation during protein harvesting, and potential loss of antigen during protein transfer, WB provides only semi-quantitative data. By comparison, the "in-cell western" (ICW) technique has high-throughput capacity and requires less extensive sample preparation. Thus, we compared the ICW technique to WB for measuring phosphorylated myosin regulatory light chain (PMLC(20)) in primary cultures of uterine myocytes to assess their relative specificity, sensitivity, precision, and quantification of biologically relevant responses.ICWs are cell-based microplate assays for quantification of protein targets in their cellular context. ICWs utilize a two-channel infrared (IR) scanner (Odyssey(R)) to quantify signals arising from near-infrared (NIR) fluorophores conjugated to secondary antibodies. One channel is dedicated to measuring the protein of interest and the second is used for data normalization of the signal in each well of the microplate. Using uterine myocytes, we assessed oxytocin (OT)-stimulated MLC(20) phosphorylation measured by ICW and WB, both using NIR fluorescence. ICW and WB data were comparable regarding signal linearity, signal specificity, and time course of phosphorylation response to OT.ICW and WB yield comparable biological data. The advantages of ICW over WB are its high-throughput capacity, improved precision, and reduced sample preparation requirements. ICW might provide better sensitivity and precision with low-quantity samples or for protocols requiring large numbers of samples. These features make the ICW technique an excellent tool for the study of phosphorylation endpoints. However, the drawbacks of ICW include the need for a cell culture format and the lack of utility where protein purification, concentration or stoichiometric analyses are required

Metallosupramolecular self-assembly of a universal 3-ravel

In the realm of supramolecular chemistry, a small number of intricately interwoven structures that bridge the boundaries between art and science have been reported. These motifs, which typically form on the nanometre scale, display both considerable beauty and complexity. However, the generation of new topologies of this type has remained a very significant synthetic challenge. Here, we describe the synthesis of a discrete highly intertwined metallosupramolecular assembly based on a universal 3-ravel motif—a topology as yet unprecedented in supramolecular chemistry. The exotic, 20-component, [Fe8L12] ravel entanglement may be considered as a 'branched knot', with individual molecules displaying either left- or right-handed chirality. The formation of this cluster was demonstrated by single-crystal and powder X-ray diffraction. The arrangement is stabilized by a favourable combination of π–π interactions and Nature's tendency to minimize voids in molecular architectures

Monogenic diabetes in children and young adults: Challenges for researcher, clinician and patient

Monogenic diabetes results from one or more mutations in a single gene which might hence be rare but has great impact leading to diabetes at a very young age. It has resulted in great challenges for researchers elucidating the aetiology of diabetes and related features in other organ systems, for clinicians specifying a diagnosis that leads to improved genetic counselling, predicting of clinical course and changes in treatment, and for patients to altered treatment that has lead to coming off insulin and injections with no alternative (Glucokinase mutations), insulin injections being replaced by tablets (e.g. low dose in HNFα or high dose in potassium channel defects -Kir6.2 and SUR1) or with tablets in addition to insulin (e.g. metformin in insulin resistant syndromes). Genetic testing requires guidance to test for what gene especially given limited resources. Monogenic diabetes should be considered in any diabetic patient who has features inconsistent with their current diagnosis (unspecified neonatal diabetes, type 1 or type 2 diabetes) and clinical features of a specific subtype of monogenic diabetes (neonatal diabetes, familial diabetes, mild hyperglycaemia, syndromes). Guidance is given by clinical and physiological features in patient and family and the likelihood of the proposed mutation altering clinical care. In this article, I aimed to provide insight in the genes and mutations involved in insulin synthesis, secretion, and resistance, and to provide guidance for genetic testing by showing the clinical and physiological features and tests for each specified diagnosis as well as the opportunities for treatment

Vitamins A, C and E and the risk of breast cancer: results from a case-control study in Greece

Although several dietary compounds are hypothesized to have anticarcinogenic properties, the role ofpecific micronutrients in the development of breast cancer remains unclear. To address this issue, we assessed intake of retinol, β-carotene, vitamin C and vitamin E in relation to breast cancer risk in a case–control study in Greece. Eight hundrednd twenty women with histologically confirmed breast cancer were compared with 1548 control women. Dietary data were collectedhrough a 115-item semiquantitative food frequency questionnaire. Data were modelled by logistic regression, with adjustment forotal energy intake and established breast cancer risk factors, as well as mutual adjustment among the micronutrients. Amongost-menopausal women, there was no association between any of the micronutrients evaluated and risk of breast cancer. Amongremenopausal women, β-carotene, vitamin C and vitamin E were each inversely associated with breast cancer risk, but afterutual adjustment among the three nutrients only β-carotene remained significant; the odds ratio (OR) for a one-quintilencrease in β-carotene intake was 0.84 (95% confidence interval 0.73–0.97). The inverse association observed with β-carotene intake, however, is slightly weaker than the association previously observed with vegetable intake in these data,aising the possibility that the observed β-carotene effect is accounted for by another component of vegetables. ©1999 Cancer Research Campaig

A meta-analytic review of stand-alone interventions to improve body image

Objective Numerous stand-alone interventions to improve body image have been developed. The present review used meta-analysis to estimate the effectiveness of such interventions, and to identify the specific change techniques that lead to improvement in body image. Methods The inclusion criteria were that (a) the intervention was stand-alone (i.e., solely focused on improving body image), (b) a control group was used, (c) participants were randomly assigned to conditions, and (d) at least one pretest and one posttest measure of body image was taken. Effect sizes were meta-analysed and moderator analyses were conducted. A taxonomy of 48 change techniques used in interventions targeted at body image was developed; all interventions were coded using this taxonomy. Results The literature search identified 62 tests of interventions (N = 3,846). Interventions produced a small-to-medium improvement in body image (d+ = 0.38), a small-to-medium reduction in beauty ideal internalisation (d+ = -0.37), and a large reduction in social comparison tendencies (d+ = -0.72). However, the effect size for body image was inflated by bias both within and across studies, and was reliable but of small magnitude once corrections for bias were applied. Effect sizes for the other outcomes were no longer reliable once corrections for bias were applied. Several features of the sample, intervention, and methodology moderated intervention effects. Twelve change techniques were associated with improvements in body image, and three techniques were contra-indicated. Conclusions The findings show that interventions engender only small improvements in body image, and underline the need for large-scale, high-quality trials in this area. The review identifies effective techniques that could be deployed in future interventions

A framework for monitoring the safety of water services: from measurements to security

The sustainable developments goals (SDGs) introduced monitoring of drinking water quality to the international development agenda. At present, Escherichia coli are the primary measure by which we evaluate the safety of drinking water from an infectious disease perspective. Here, we propose and apply a framework to reflect on the purposes of and approaches to monitoring drinking water safety. To deliver SDG 6.1, universal access to safe drinking water, a new approach to monitoring is needed. At present, we rely heavily on single measures of E. coli contamination to meet a normative definition of safety. Achieving and sustaining universal access to safe drinking water will require monitoring that can inform decision making on whether services are managed to ensure safety and security of access