Ubiquitination and proteasomal degradation of ATG12 regulates its proapoptotic activity

During macroautophagy, conjugation of ATG12 to ATG5 is essential for LC3 lipidation and autophagosome formation. Additionally, ATG12 has ATG5-independent functions in diverse processes including mitochondrial fusion and mitochondrial-dependent apoptosis. In this study, we investigated the regulation of free ATG12. In stark contrast to the stable ATG12–ATG5 conjugate, we find that free ATG12 is highly unstable and rapidly degraded in a proteasome-dependent manner. Surprisingly, ATG12, itself a ubiquitin-like protein, is directly ubiquitinated and this promotes its proteasomal degradation. As a functional consequence of its turnover, accumulation of free ATG12 contributes to proteasome inhibitor-mediated apoptosis, a finding that may be clinically important given the use of proteasome inhibitors as anticancer agents. Collectively, our results reveal a novel interconnection between autophagy, proteasome activity, and cell death mediated by the ubiquitin-like properties of ATG12

Divergent drivers of carbon dioxide and methane dynamics in an agricultural coastal floodplain: post-flood hydrological and biological drivers

Many coastal floodplains have been artificially drained for agriculture, altering hydrological connectivity and the delivery of groundwater-derived solutes including carbon dioxide (CO2) and methane (CH4) to surface waters. Here, we investigated the drivers of CO2 and CH4 within the artificial drains.of a coastal floodplain under sugarcane plantation and quantify the contribution of groundwater discharge to CO2 and CH4 dynamics over a flood event (290 mm of rainfall). High temporal resolution, in situ observations of dissolved CO2 and CH4, carbon stable isotopes of CH4 (delta C-13-CH4), and the natural groundwater tracer radon (Rn-222) allowed us to quantify. CO2, CH4 and groundwater dynamics during the rapid recession of a flood over a five day period. Extreme super-saturation of free CO2 ([CO2*]) up to 2,951 mu M (25,480% of atmospheric equilibrium) was driven by large groundwater input into the drains (maximum 87 cm day-(1)), caused by a steep hydraulic head in the adjacent water table. Groundwater input sustained between 95 and 124% of the surface [CO2*] flux during the flood recession by delivering high carbonate alkalinity groundwater (DIC = 10,533 mu M, similar to pH = 7.05) to acidic surface water (pH <4), consequently transforming all groundwater-derived DIC to [CO2*]. In contrast, groundwater was not a major direct driver of CH4 contributing only 14% of total CH4 fluxes. A progressive increase in CH4 concentrations of up to similar to 2400 nM day-(1) occurred as a combination of increased substrate availability delivered by post-flood drainage water and longer residence times, which allowed for a biogenic CH4 signal to develop. The progressive enrichment in delta C-13-CH4 values (- 70%. to-48%.) and increase in CH4 concentrations (46-2460 nM) support coupled production-oxidation, with concentrations and delta C-13 values remaining higher (2,798 nM and-47%.) than pre-flood conditions (534 nM and-55 parts per thousand) three weeks after the flood. Our findings demonstrate how separate processes can drive the aquatic CO2 and CH4 response to a flood event in a drained coastal floodplain, and the key role groundwater had in post-flood [CO2*] evasion to the atmosphere, but not CH4. (C) 2016 Elsevier B.V. All rights reserved

RIPK3 activation leads to cytokine synthesis that continues after loss of cell membrane integrity

Necroptosis is a form of programmed cell death that is defined by activation of the kinase RIPK3 and subsequent cell membrane permeabilization by the effector MLKL. RIPK3 activation can also promote immune responses via production of cytokines and chemokines. How active cytokine production is coordinated with the terminal process of necroptosis is unclear. Here, we report that cytokine production continues within necroptotic cells even after they have lost cell membrane integrity and irreversibly committed to death. This continued cytokine production is dependent on mRNA translation and requires maintenance of endoplasmic reticulum integrity that remains after plasma membrane integrity is lost. The continued translation of cytokines by cellular corpses contributes to necroptotic cell uptake by innate immune cells and priming of adaptive immune responses to antigens associated with necroptotic corpses. These findings imply that cell death and production of inflammatory mediators are coordinated to optimize the immunogenicity of necroptotic cells

Mitochondria are required for pro-ageing features of the senescent phenotype

Cell senescence is an important tumour suppressor mechanism and driver of ageing. Both functions are dependent on the development of the senescent phenotype, which involves an overproduction of pro‐inflammatory and pro‐oxidant signals. However, the exact mechanisms regulating these phenotypes remain poorly understood. Here, we show the critical role of mitochondria in cellular senescence. In multiple models of senescence, absence of mitochondria reduced a spectrum of senescence effectors and phenotypes while preserving ATP production via enhanced glycolysis. Global transcriptomic analysis by RNA sequencing revealed that a vast number of senescent‐associated changes are dependent on mitochondria, particularly the pro‐inflammatory phenotype. Mechanistically, we show that the ATM, Akt and mTORC1 phosphorylation cascade integrates signals from the DNA damage response (DDR) towards PGC‐1β‐dependent mitochondrial biogenesis, contributing to a ROS‐mediated activation of the DDR and cell cycle arrest. Finally, we demonstrate that the reduction in mitochondrial content in vivo, by either mTORC1 inhibition or PGC‐1β deletion, prevents senescence in the ageing mouse liver. Our results suggest that mitochondria are a candidate target for interventions to reduce the deleterious impact of senescence in ageing tissues

First narrow-band search for continuous gravitational waves from known pulsars in advanced detector data

Spinning neutron stars asymmetric with respect to their rotation axis are potential sources of continuous gravitational waves for ground-based interferometric detectors. In the case of known pulsars a fully coherent search, based on matched filtering, which uses the position and rotational parameters obtained from electromagnetic observations, can be carried out. Matched filtering maximizes the signalto- noise (SNR) ratio, but a large sensitivity loss is expected in case of even a very small mismatch between the assumed and the true signal parameters. For this reason, narrow-band analysis methods have been developed, allowing a fully coherent search for gravitational waves from known pulsars over a fraction of a hertz and several spin-down values. In this paper we describe a narrow-band search of 11 pulsars using data from Advanced LIGO’s first observing run. Although we have found several initial outliers, further studies show no significant evidence for the presence of a gravitational wave signal. Finally, we have placed upper limits on the signal strain amplitude lower than the spin-down limit for 5 of the 11 targets over the bands searched; in the case of J1813-1749 the spin-down limit has been beaten for the first time. For an additional 3 targets, the median upper limit across the search bands is below the spin-down limit. This is the most sensitive narrow-band search for continuous gravitational waves carried out so far

Pulpotomy for the Management of Irreversible Pulpitis in Mature Teeth (PIP) : a feasibility study

Fundings: This study is funded by the National Institute for Health Research (NIHR) Health Technology Assessment Program (project reference NIHR129230). The views expressed are those of the authors and not necessarily those of the NIHR or the Department of Health and Social Care. The funding body has had no role in the design of the study and will have no role in the collection, analysis, and interpretation of the data and in the writing of any future manuscript. Acknowledgements The authors would like to thank all the patients, dentists and dental team members who are participating in the PIP Trial. We would also like to thank the members of the TSC and DMEC. We would like to acknowledge the funding for the project from the National Institute for Health Research Health Technology Assessment Programme (Project Number NIHR129230). The views and opinions expressed herein are those of the authors and do not necessarily reflect those of the HTA programme, NIHR, NHS or the Department of Health. Sponsor: University of Dundee Funder: National Institute for Health Research (NIHR), Health Technology Assessment (HTA) Programme, Project number: NIHR129230 The PIP study group consists of the co-chief investigators, grant holders, project management group and the Trial Management Committee as outlined as follows: Co-chief investigators: Jan E Clarkson (JC) and Craig R Ramsay (CR) Grant holders: Sondos Albradri (SA), Avijit Banerjee (AB), Katie Banister (KB), Dwayne Boyers (DB), David Conway (DC), Chris Deery (CD), Beatriz Goulao (BG), Ekta Gupta (EG), Fadi Jarad (FJ), Thomas Lamont (TL), Graeme MacLennan (GMacL), Francesco Mannocci (FM) Zoe Marshmann (ZM), Tina McGuff (TMcG), David Ricketts (DR), Douglas Robertson (DR) Marjon van der Pol (MvdP) and Linda Young (LY). Trial Management Committee: Sondos Albradri (SA), Avijit Banerjee (AB), Katie Banister (KB), Chris Deery (CD), Rosanne Bell (RB), David Conway (DC), Dwayne Boyers (DB), Lori Brown (LB), Pina Donaldson (PD), Anne Duncan (AD), Katharine Dunn (KD), Patrick Fee (PF), Mark Forrest (MF), Jill Gouick (JG), Beatriz Goulao (BG), Ekta Gupta (EG), Alice Hamilton (AH), Fadi Jarad (FJ), Jennifer Kettle (JK), Thomas Lamont (TL), Graeme MacLennan (GMacL), Lorna Macpherson (LM), Francesco Mannocci (FM), Zoe Marshmann (ZM), Fiona Mitchell (FM), Tina McGuff (TMcG), David Ricketts (DR), Douglas Robertson (DR), Marjon van der Pol (MvdP), Gabriella Wojewodka (GW) and Linda Young (LY)Peer reviewedPublisher PD

Clinical Utility of Random Anti–Tumor Necrosis Factor Drug–Level Testing and Measurement of Antidrug Antibodies on the Long-Term Treatment Response in Rheumatoid Arthritis

Objective: To investigate whether antidrug antibodies and/or drug non-trough levels predict the long-term treatment response in a large cohort of patients with rheumatoid arthritis (RA) treated with adalimumab or etanercept and to identify factors influencing antidrug antibody and drug levels to optimize future treatment decisions.  Methods: A total of 331 patients from an observational prospective cohort were selected (160 patients treated with adalimumab and 171 treated with etanercept). Antidrug antibody levels were measured by radioimmunoassay, and drug levels were measured by enzyme-linked immunosorbent assay in 835 serial serum samples obtained 3, 6, and 12 months after initiation of therapy. The association between antidrug antibodies and drug non-trough levels and the treatment response (change in the Disease Activity Score in 28 joints) was evaluated.  Results: Among patients who completed 12 months of followup, antidrug antibodies were detected in 24.8% of those receiving adalimumab (31 of 125) and in none of those receiving etanercept. At 3 months, antidrug antibody formation and low adalimumab levels were significant predictors of no response according to the European League Against Rheumatism (EULAR) criteria at 12 months (area under the receiver operating characteristic curve 0.71 [95% confidence interval (95% CI) 0.57, 0.85]). Antidrug antibody–positive patients received lower median dosages of methotrexate compared with antidrug antibody–negative patients (15 mg/week versus 20 mg/week; P = 0.01) and had a longer disease duration (14.0 versus 7.7 years; P = 0.03). The adalimumab level was the best predictor of change in the DAS28 at 12 months, after adjustment for confounders (regression coefficient 0.060 [95% CI 0.015, 0.10], P = 0.009). Etanercept levels were associated with the EULAR response at 12 months (regression coefficient 0.088 [95% CI 0.019, 0.16], P = 0.012); however, this difference was not significant after adjustment. A body mass index of ≥30 kg/m2 and poor adherence were associated with lower drug levels.  Conclusion: Pharmacologic testing in anti–tumor necrosis factor–treated patients is clinically useful even in the absence of trough levels. At 3 months, antidrug antibodies and low adalimumab levels are significant predictors of no response according to the EULAR criteria at 12 months