The transcriptional repressor protein NsrR senses nitric oxide directly via a [2Fe-2S] cluster

The regulatory protein NsrR, a member of the Rrf2 family of transcription repressors, is specifically dedicated to sensing nitric oxide (NO) in a variety of pathogenic and non-pathogenic bacteria. It has been proposed that NO directly modulates NsrR activity by interacting with a predicted [Fe-S] cluster in the NsrR protein, but no experimental evidence has been published to support this hypothesis. Here we report the purification of NsrR from the obligate aerobe Streptomyces coelicolor. We demonstrate using UV-visible, near UV CD and EPR spectroscopy that the protein contains an NO-sensitive [2Fe-2S] cluster when purified from E. coli. Upon exposure of NsrR to NO, the cluster is nitrosylated, which results in the loss of DNA binding activity as detected by bandshift assays. Removal of the [2Fe-2S] cluster to generate apo-NsrR also resulted in loss of DNA binding activity. This is the first demonstration that NsrR contains an NO-sensitive [2Fe-2S] cluster that is required for DNA binding activity

Protein Architecture of the Human Kinetochore Microtubule Attachment Site

Centromeric chromatin – spindle microtubule interactions mediated by kinetochores drive chromosome segregation. We have developed a two-color fluorescence light microscopy method that measures average label separation, Delta, at < 5 nm accuracy — to elucidate the protein architecture of human metaphase kinetochores. Delta analysis, when correlated with tension states of spindle-attached sister kinetochore pairs, provided information on mechanical properties of protein linkages within kinetochores. Treatment with taxol—which suppresses microtubule dynamics, eliminates tension at kinetochores, and activates the spindle checkpoint—resulted in specific large-scale changes in kinetochore architecture. Cumulatively, Delta analysis revealed compliant linkages close to the centromeric chromatin, suggests a model for how the KMN (KNL1/Mis12 complex/Ndc80 complex) network provides microtubule attachment and generates pulling forces from depolymerization, and reveals architectural changes induced by taxol treatment. The methods described here should also be applicable to other intermediate-scale biological machines in cells

Association of Accelerometry-Measured Physical Activity and Cardiovascular Events in Mobility-Limited Older Adults: The LIFE (Lifestyle Interventions and Independence for Elders) Study.

BACKGROUND:Data are sparse regarding the value of physical activity (PA) surveillance among older adults-particularly among those with mobility limitations. The objective of this study was to examine longitudinal associations between objectively measured daily PA and the incidence of cardiovascular events among older adults in the LIFE (Lifestyle Interventions and Independence for Elders) study. METHODS AND RESULTS:Cardiovascular events were adjudicated based on medical records review, and cardiovascular risk factors were controlled for in the analysis. Home-based activity data were collected by hip-worn accelerometers at baseline and at 6, 12, and 24&nbsp;months postrandomization to either a physical activity or health education intervention. LIFE study participants (n=1590; age 78.9±5.2 [SD] years; 67.2% women) at baseline had an 11% lower incidence of experiencing a subsequent cardiovascular event per 500&nbsp;steps taken per day based on activity data (hazard ratio, 0.89; 95% confidence interval, 0.84-0.96; P=0.001). At baseline, every 30&nbsp;minutes spent performing activities ≥500&nbsp;counts per minute (hazard ratio, 0.75; confidence interval, 0.65-0.89 [P=0.001]) were also associated with a lower incidence of cardiovascular events. Throughout follow-up (6, 12, and 24&nbsp;months), both the number of steps per day (per 500&nbsp;steps; hazard ratio, 0.90, confidence interval, 0.85-0.96 [P=0.001]) and duration of activity ≥500&nbsp;counts per minute (per 30&nbsp;minutes; hazard ratio, 0.76; confidence interval, 0.63-0.90 [P=0.002]) were significantly associated with lower cardiovascular event rates. CONCLUSIONS:Objective measurements of physical activity via accelerometry were associated with cardiovascular events among older adults with limited mobility (summary score &gt;10 on the Short Physical Performance Battery) both using baseline and longitudinal data. CLINICAL TRIAL REGISTRATION:URL: http://www.clinicaltrials.gov. Unique identifier: NCT01072500

Inactivation of the transforming growth factor β type II receptor in human small cell lung cancer cell lines

Transforming growth factor β (TGF-β) exerts a growth inhibitory effect on many cell types through binding to two types of receptors, the type I and II receptors. Resistance to TGF-β due to lack of type II receptor (RII) has been described in some cancer types including small cell lung cancer (SCLC). The purpose of this study was to examine the cause of absent RII expression in SCLC cell lines. Northern blot analysis showed that RII RNA expression was very weak in 16 of 21 cell lines. To investigate if the absence of RII transcript was due to mutations, we screened the poly-A tract for mutations, but no mutations were detected. Additional screening for mutations of the RII gene revealed a GG to TT base substitution in one cell line, which did not express RII. This mutation generates a stop codon resulting in predicted synthesis of a truncated RII of 219 amino acids. The nature of the mutation, which has not previously been observed in RII, has been linked to exposure to benzo[a]-pyrene, a component of cigarette smoke. Since RII has been mapped to chromosome 3p22 and nearby loci are often hypermethylated in SCLC, it was examined whether the lack of RII expression was due to hypermethylation. Southern blot analysis of the RII promoter did not show altered methylation patterns. The restriction endonuclease pattern of the RII gene was altered in two SCLC cell lines when digested with Sma 1. However, treatment with 5-aza-2′-deoxycytidine did not induce expression of RII mRNA. Our results indicate that in SCLC lack of RII mRNA is not commonly due to mutations and inactivation of RII transcription was not due to hypermethylation of the RII promoter or gene. Thus, these data show that in most cases of the SCLC cell lines, the RII gene and promoter is intact in spite of absent RII expression. However, the nature of the mutation found could suggest that it was caused by cigarette smoking. © 1999 Cancer Research Campaig

Autism as a disorder of neural information processing: directions for research and targets for therapy

The broad variation in phenotypes and severities within autism spectrum disorders suggests the involvement of multiple predisposing factors, interacting in complex ways with normal developmental courses and gradients. Identification of these factors, and the common developmental path into which theyfeed, is hampered bythe large degrees of convergence from causal factors to altered brain development, and divergence from abnormal brain development into altered cognition and behaviour. Genetic, neurochemical, neuroimaging and behavioural findings on autism, as well as studies of normal development and of genetic syndromes that share symptoms with autism, offer hypotheses as to the nature of causal factors and their possible effects on the structure and dynamics of neural systems. Such alterations in neural properties may in turn perturb activity-dependent development, giving rise to a complex behavioural syndrome many steps removed from the root causes. Animal models based on genetic, neurochemical, neurophysiological, and behavioural manipulations offer the possibility of exploring these developmental processes in detail, as do human studies addressing endophenotypes beyond the diagnosis itself

Promoting higher added value to a finfish species rejected to sea

This project aimed to the development of the research and the technology necessary to promote higher added value to fishing activity. This is to be achieved by obtaining profit from a finfish species (“Rockcod”, Patagonotothen spp.) not known to consumers and currently discarded by the EU fishing fleet operating in the South West Atlantic, in order to supply the EU seafood industry with a good quality raw material for human food manufacturing. Use of this species, caught as a by-catch in the existing fisheries targeting hakes and cephalopods, should also increase the profitability of the fleet, contribute to maintaining employment and help to counterbalance the negative effects of fishing activity and discards in the ecosystem. The main scientific-technological objectives and expected achievements were the following: - Description of the fisheries - Improved knowledge of the biology of the species - Biomass assessment - Estimation of catches and discards - Analysis of the spatial and temporal distribution of the resource. Fishery forecasting and testing - Sensorial, Microbiological, Nutritional and Biochemical Evaluation of Rock cod - Development of the technical modifications on board commercial vessels - Development of new processed products from frozen Rock codEuropean Commission Cooperative Research (CRAFT

Efficacy and tolerability of lisdexamfetamine dimesylate in children with attention-deficit/hyperactivity disorder: sex and age effects and effect size across the day

<p>Abstract</p> <p>Background</p> <p>Efficacy and safety profiles by sex and age (6-9 vs 10-12 years) and magnitude and duration of effect by effect size overall and across the day of lisdexamfetamine dimesylate (LDX) vs placebo were assessed.</p> <p>Methods</p> <p>This study enrolled children (6-12 years) with attention-deficit/hyperactivity disorder (ADHD) in an open-label dose optimization with LDX (30-70 mg/d) followed by a randomized, double-blind, placebo-controlled, 2-way crossover phase. Post hoc analyses assessed interaction between sex or age and treatment and assessed effect sizes for Swanson, Kotkin, Agler, M-Flynn, and Pelham (SKAMP) and Permanent Product Measure of Performance (PERMP) scales and ADHD Rating Scale IV measures. No corrections for multiple testing were applied on time points and subgroup statistical comparisons.</p> <p>Results</p> <p>129 participants enrolled; 117 randomized. Both sexes showed improvement on all assessments at postdose time points; females showed less impairment than males for SKAMP and PERMP scores in treatment and placebo groups at nearly all times. Both age groups improved on all assessments at postdose time points. Children 10-12 years had less impairment in SKAMP ratings than those 6-9 years. Treatment-by-sex interactions were observed at time points for SKAMP-D, SKAMP total, and PERMP scores; no consistent pattern across scales or time points was observed. LDX demonstrated significant improvement vs placebo, by effect size, on SKAMP-D from 1.5-13 hours postdose. The overall LS mean (SE) SKAMP-D effect size was -1.73 (0.18). In the dose-optimization phase, common (≥2%) treatment-emergent adverse events (TEAEs) in males were upper abdominal pain, headache, affect lability, initial insomnia, and insomnia; in females were nausea and decreased weight. During the crossover phase for those taking LDX, higher incidence (≥2% greater) was observed in males for upper abdominal pain and insomnia and in females for nausea and headache. Overall incidence of TEAEs in age groups was similar.</p> <p>Conclusion</p> <p>Apparent differences in impairment level between sex and age groups were noted. However, these results support the efficacy of LDX from 1.5 hours to 13 hours postdose in boys and girls with medium to large effect sizes across the day with some variability in TEAE incidence by sex.</p> <p>Trial Registration Number</p> <p>ClinicalTrials.gov Identifier: <a href="http://clinicaltrials.gov/ct2/show/NCT00500149">NCT00500149</a>.</p

A cluster randomised trial of a telephone-based intervention for parents to increase fruit and vegetable consumption in their 3- to 5-year-old children: study protocol

Background: Inadequate fruit and vegetable consumption in childhood increases the risk of developing chronic disease. Despite this, a substantial proportion of children in developed nations, including Australia, do not consume sufficient quantities of fruits and vegetables. Parents are influential in the development of dietary habits of young children but often lack the necessary knowledge and skills to promote healthy eating in their children. The aim of this study is to assess the efficacy of a telephone-based intervention for parents to increase the fruit and vegetable consumption of their 3- to 5-year-old children. Methods/Design: The study, conducted in the Hunter region of New South Wales, Australia, employs a cluster randomised controlled trial design. Two hundred parents from 15 randomly selected preschools will be randomised to receive the intervention, which consists of print resources and four weekly 30-minute telephone support calls delivered by trained telephone interviewers. The calls will assist parents to increase the availability and accessibility of fruit and vegetables in the home, create supportive family eating routines and role-model fruit and vegetable consumption. A further two hundred parents will be randomly allocated to the control group and will receive printed nutrition information only. The primary outcome of the trial will be the change in the child's consumption of fruit and vegetables as measured by the fruit and vegetable subscale of the Children's Dietary Questionnaire. Pre-intervention and post-intervention parent surveys will be administered over the telephone. Baseline surveys will occur one to two weeks prior to intervention delivery, with follow-up data collection calls occurring two, six, 12 and 18 months following baseline data collection. Discussion: If effective, this telephone-based intervention may represent a promising public health strategy to increase fruit and vegetable consumption in childhood and reduce the risk of subsequent chronic disease. Trial registration: Australian Clinical Trials Registry ACTRN12609000820202

Mutations in the Mitochondrial Methionyl-tRNA Synthetase Cause a Neurodegenerative Phenotype in Flies and a Recessive Ataxia (ARSAL) in Humans

The study of Drosophila neurodegenerative mutants combined with genetic and biochemical analyses lead to the identification of multiple complex mutations in 60 patients with a novel form of ataxia/leukoencephalopathy

New insights into the genetic etiology of Alzheimer's disease and related dementias

Characterization of the genetic landscape of Alzheimer's disease (AD) and related dementias (ADD) provides a unique opportunity for a better understanding of the associated pathophysiological processes. We performed a two-stage genome-wide association study totaling 111,326 clinically diagnosed/'proxy' AD cases and 677,663 controls. We found 75 risk loci, of which 42 were new at the time of analysis. Pathway enrichment analyses confirmed the involvement of amyloid/tau pathways and highlighted microglia implication. Gene prioritization in the new loci identified 31 genes that were suggestive of new genetically associated processes, including the tumor necrosis factor alpha pathway through the linear ubiquitin chain assembly complex. We also built a new genetic risk score associated with the risk of future AD/dementia or progression from mild cognitive impairment to AD/dementia. The improvement in prediction led to a 1.6- to 1.9-fold increase in AD risk from the lowest to the highest decile, in addition to effects of age and the APOE ε4 allele