Changes in Proteasome Structure and Function Caused by HAMLET in Tumor Cells

BACKGROUND: Proteasomes control the level of endogenous unfolded proteins by degrading them in the proteolytic core. Insufficient degradation due to altered protein structure or proteasome inhibition may trigger cell death. This study examined the proteasome response to HAMLET, a partially unfolded protein-lipid complex, which is internalized by tumor cells and triggers cell death. METHODOLOGY/PRINCIPAL FINDINGS: HAMLET bound directly to isolated 20S proteasomes in vitro and in tumor cells significant co-localization of HAMLET and 20S proteasomes was detected by confocal microscopy. This interaction was confirmed by co-immunoprecipitation from extracts of HAMLET-treated tumor cells. HAMLET resisted in vitro degradation by proteasomal enzymes and degradation by intact 20S proteasomes was slow compared to fatty acid-free, partially unfolded alpha-lactalbumin. After a brief activation, HAMLET inhibited proteasome activity in vitro and in parallel a change in proteasome structure occurred, with modifications of catalytic (beta1 and beta5) and structural subunits (alpha2, alpha3, alpha6 and beta3). Proteasome inhibition was confirmed in extracts from HAMLET-treated cells and there were indications of proteasome fragmentation in HAMLET-treated cells. CONCLUSIONS/SIGNIFICANCE: The results suggest that internalized HAMLET is targeted to 20S proteasomes, that the complex resists degradation, inhibits proteasome activity and perturbs proteasome structure. We speculate that perturbations of proteasome structure might contribute to the cytotoxic effects of unfolded protein complexes that invade host cells

First data with the ATLAS Level-1 Calorimeter Trigger

The ATLAS Level-1 Calorimeter Trigger is one of the main elements of the first stage of event selection for the ATLAS experiment at the LHC. The input stage consists of a mixed analogue/digital component taking trigger sums from the ATLAS calorimeters. The trigger logic is performed in a digital, pipelined system with several stages of processing, largely based on FPGAs, which perform programmable algorithms in parallel with a fixed latency to process about 300 Gbyte/s of input data. The real-time output consists of counts of different types of physics objects, and energy sums. The final system consists of over 300 custom-built VME modules, of several different types. The installation at ATLAS of these modules, and the necessary infrastructure, was completed at the end of 2007. The system has since undergone intensive testing, both in standalone mode, and in conjunction with the whole of the ATLAS detector in combined running. The final steps of commissioning, and experience with running the full-scale system are presented. Results of integration tests performed with the upstream calorimeters, and downstream trigger and data-flow systems, are shown, along with an analysis of the performance of the calorimeter trigger in full ATLAS data-taking. This includes trigger operation during the cosmic muon runs from before LHC start-up, and a first look at LHC proton beam data

Improving patient experience and safety at transitions of care through the Your Care Needs You (YCNY) intervention: a study protocol for a cluster randomised controlled feasibility trial

Background Patients, particularly older people, often experience safety issues when transitioning from hospital to home. Although the evidence is currently equivocal as to how we can improve this transition of care, interventions that support patient involvement may be more effective. The ‘Your Care Needs You’ (YCNY) intervention supports patients to ‘know more’ and ‘do more’ whilst in hospital in order that they better understand their health condition and medications, maintain their daily activities, and can seek help at home if required. The intervention aims to reduce emergency hospital readmissions and improve safety and experience during the transition to home. Methods As part of the Partners At Care Transitions (PACT) programme of research, a multi-centred cluster randomised controlled trial (cRCT) will be conducted to explore the feasibility of the YCNY intervention and trial methodology. Data will be used to refine the intervention and develop a protocol for a definitive cRCT. Ten acute hospital wards (the clusters) from varying medical specialties including older peoples’ medicine, trauma and orthopaedics, cardiology, intermediate care, and stroke will be randomised to deliver YCNY or usual care on a 3:2 basis. Up to 200 patients aged 75 years and over and discharged to their own homes will be recruited to the study. Patients will complete follow-up questionnaires at 5-, 30-, and 90-days post-discharge and readmission data up to 90-days post-discharge will be extracted from their medical records. Study outcomes will include measures of feasibility (e.g. screening, recruitment, and retention data) and processes required to collect routine data at a patient and ward level. In addition, interviews and observations involving up to 24 patients/carers and 28 staff will be conducted to qualitatively assess the acceptability, usefulness, and feasibility of the intervention and implementation package to patients and staff. A separate sub-study will be conducted to explore how accurately primary outcome data (30-day emergency hospital readmissions) can be gathered for the definitive cRCT. Discussion This study will establish the feasibility of the YCNY intervention which aims to improve safety and experience during transitions of care. It will identify key methodological and implementation issues that need to be addressed prior to assessing the effectiveness of the YCNY intervention in a definitive cluster randomised controlled trial. Trial registration UK Clinical Research Network Portfolio: 42191; ISTCRN: ISRCTN51154948. Registered 16/07/2019