DNA-dependent Protein Kinase-mediated Phosphorylation of Protein Kinase B Requires a Specific Recognition Sequence in the C-terminal Hydrophobic Motif

DNA-dependent protein kinase (DNA-PK) has been implicated in a variety of nuclear processes including DNA double strand break repair, V(D)J recombination, and transcription. A recent study showed that DNA-PK is responsible for Ser-473 phosphorylation in the hydrophobic motif of protein kinase B (PKB/Akt) in genotoxic-stressed cells, suggesting a novel role for DNA-PK in cell signaling. Here, we report that DNA-PK activity toward PKB peptides is impaired in DNA-PK knock-out mouse embryonic fibroblast cells when compared with wild type. In addition, human glioblastoma cells expressing a mutant form of DNA-PK (M059J) displayed a lower DNA-PK activity when compared with glioblastoma cells expressing wild-type DNA- PK (M059K) when PKB peptide substrates were tested. DNA- PK preferentially phosphorylated PKB on Ser-473 when compared with its known in vitro substrate, p53. A consensus hydrophobic amino acid surrounding the Ser-473 phospho-acceptor site in PKB containing amino acids Phe at position +1 and +4 and Tyr at position -1 are critical for DNA- PK activity. Thus, these data define the specificity of DNA- PK action as a Ser-473 kinase for PKB in DNA repair signaling.

Confined Site Construction: A qualitative investigation of critical issues affecting management of Health and Safety

The construction industry is inherently risky, with a significant number of accidents and disasters occurring, particularly on confined construction sites. This research investigates and identifies the various issues affecting successful management of health and safety in confined construction sites. The rationale is that identifying the issues would assist the management of health and safety particularly in inner city centres which are mostly confined sites. Using empiricism epistemology, the methodology was based on qualitative research approach by means of multiple case studies in three different geographical locations of Ireland, UK and USA. Data on each case study were collected through individual interviews and focus group discussion with project participants. The findings suggest that three core issues are the underlying factors affecting management of health and safety on confined construction sites. It include, (i) lack of space, (ii) problem of co-ordination and management of site personnel, and (iii) overcrowding of workplace. The implication of this is that project teams and their organisations should see project processes from a holistic point of view, as a unified single system, where quick intervention in solving a particular issue should be the norm, so as not to adversely affect interrelated sequence of events in project operations. Proactive strategies should be devised to mitigate these issues and may include detail project programming, space management, effective constructability review and efficient co-ordination of personnel, plant and materials among others. The value of this research is to aid management and operation of brownfield sites by identifying issues impacting on health and safety management in project process.

Exercise rehabilitation following intensive care unit discharge for recovery from critical illness

Background: Skeletal muscle wasting and weakness are significant complications of critical illness, associated with the degree of illness severity and periods of reduced mobility during mechanical ventilation. They contribute to the profound physical and functional deficits observed in survivors. These impairments may persist for many years following discharge from the intensive care unit (ICU) and may markedly influence health-related quality of life. Rehabilitation is a key strategy in the recovery of patients following critical illness. Exercise based interventions are aimed at targeting this muscle wasting and weakness. Physical rehabilitation delivered during ICU admission has been systematically evaluated and shown to be beneficial. However its effectiveness when initiated after ICU discharge has yet to be established. Objectives: To assess the effectiveness of exercise rehabilitation programmes, initiated after ICU discharge, on functional exercise capacity and health-related quality of life in adult ICU survivors who have been mechanically ventilated for more than 24 hours. Search methods:We searched the following databases: the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library), OvidSP MEDLINE, Ovid SP EMBASE, and CINAHL via EBSCO host to 15th May 2014. We used a specific search strategy for each database. This included synonyms for ICU and critical illness, exercise training and rehabilitation. We searched the reference lists of included studies and contacted primary authors to obtain further information regarding potentially eligible studies. We also searched major clinical trials registries (Clinical Trials and Current Controlled Trials) and the personal libraries of the review authors. We applied no language or publication restriction. We reran the search in February 2015. We will deal with any studies of interest when we update the review.  Selection criteria:We included randomized controlled trials (RCTs), quasi-RCTs, and controlled clinical trials (CCTs) that compared an exercise interventioninitiated after ICU discharge to any other intervention or a control or ‘usual care’ programme in adult (≥18years) survivors ofcritical illness. Data collection and analysis:We used standard methodological procedures expected by The Cochrane Collaboration. Main results:We included six trials (483 adult ICU participants). Exercise-based interventions were delivered on the ward in two studies; both onthe ward and in the community in one study; and in the community in three studies. The duration of the intervention varied according to the length of stay in hospital following ICU discharge (up to a fixed duration of 12 weeks).Risk of bias was variable for all domains across all trials. High risk of bias was evident in all studies for performance bias, although blinding of participants and personnel in therapeutic rehabilitation trials can be pragmatically challenging. Low risk of bias was at least 50% for all other domains across all trials, although high risk of bias was present in one study for random sequence generation (selection bias), incomplete outcome data (attrition bias) and other sources. Risk of bias was unclear for remaining studies across the domains.All six studies measured effect on the primary outcome of functional exercise capacity, although there was wide variability in natureof intervention, outcome measures and associated metrics, and data reporting. Overall quality of the evidence was very low. Only two studies using the same outcome measure for functional exercise capacity, had the potential for pooling of data and assessment of heterogeneity. On statistical advice, this was considered inappropriate to perform this analysis and study findings were therefore qualitatively described. Individually, three studies reported positive results in favour of the intervention. A small benefit (versus. control)was evident in anaerobic threshold in one study (mean difference, MD (95% confidence interval, CI), 1.8 mlO2/kg/min (0.4 to 3.2),P value = 0.02), although this effect was short-term, and in a second study, both incremental (MD 4.7 (95% CI 1.69 to 7.75) Watts, P value = 0.003) and endurance (MD 4.12 (95% CI 0.68 to 7.56) minutes, P value = 0.021) exercise testing demonstrated improvement.Finally self-reported physical function increased significantly following a rehabilitation manual (P value = 0.006). Remaining studies found no effect of the intervention.Similar variability in with regard findings for the primary outcome of health-related quality of life were also evident. Only two studies evaluated this outcome. Following statistical advice, these data again were considered inappropriate for pooling to determine overall effect and assessment of heterogeneity. Qualitative description of findings was therefore undertaken. Individually, neither study reported differences between intervention and control groups for health-related quality of life as a result of the intervention. Overall quality of the evidence was very low.Mortality was reported by all studies, ranging from 0% to 18.8%. Only one non-mortality adverse event was reported across all patients in all studies (a minor musculoskeletal injury). Withdrawals, reported in four studies, ranged from 0% to 26.5% in control groups,and 8.2% to 27.6% in intervention groups. Loss to follow-up, reported in all studies, ranged from 0% to 14% in control groups, and 0% to 12.5% in intervention groups. Authors’ conclusions:We are unable, at this time, to determine an overall effect on functional exercise capacity, or health-related quality of life, of an exercise based intervention initiated after ICU discharge in survivors of critical illness. Meta-analysis of findings was not appropriate. This was due to insufficient study number and data. Individual study findings were inconsistent. Some studies reported a beneficial effect of the intervention on functional exercise capacity, and others not. No effect was reported on health-related quality of life. Methodological rigour was lacking across a number of domains influencing quality of the evidence. There was also wide variability in the characteristics of interventions, outcome measures and associated metrics, and data reporting.If further trials are identified, we may be able to determine the effect of exercise-based interventions following ICU discharge, on functional exercise capacity and health-related quality of life in survivors of critical illness.

HES6 drives a critical AR transcriptional programme to induce castration-resistant prostate cancer through activation of an E2F1-mediated cell cycle network

Castrate-resistant prostate cancer (CRPC) is poorly characterized and heterogeneous and while the androgen receptor (AR) is of singular importance, other factors such as c-Myc and the E2F family also play a role in later stage disease. HES6 is a transcription co-factor associated with stem cell characteristics in neural tissue. Here we show that HES6 is up-regulated in aggressive human prostate cancer and drives castration-resistant tumour growth in the absence of ligand binding by enhancing the transcriptional activity of the AR, which is preferentially directed to a regulatory network enriched for transcription factors such as E2F1. In the clinical setting, we have uncovered a HES6-associated signature that predicts poor outcome in prostate cancer, which can be pharmacologically targeted by inhibition of PLK1 with restoration of sensitivity to castration. We have therefore shown for the first time the critical role of HES6 in the development of CRPC and identified its potential in patient-specific therapeutic strategies.