Work schedule and seasonal influences on sleep and fatigue in helicopter and fixed-wing aircraft operations in extreme environments.

Sleep and fatigue were investigated in aviation search and rescue, firefighting, emergency medical services and offshore transfer operations in 210 participants, for 21 days each, across 17 datasets in seven countries. Sleep data were collected using wrist-worn actigraphs and sleep diaries. Sustained attention was assessed using a 5-min Psychomotor Vigilance Task (PVT). Duty information was provided from corporate IT systems. Despite the number of 24 h operations, most work occurred during daytime hours, and most sleep occurred at night. There were seasonal changes in work and sleep patterns, with naps used to augment total sleep time. The proportion of sleep occurring during duty varied from zero to 30%. Differences in PVT response times were trivial to small. Legislation that defines flight, duty time and minimum rest limits assume that sleep is not obtained during duty periods, apart from some napping under Fatigue Risk Management Systems (FRMS). However, especially in cases where the aviation service requires waiting for tasks (e.g. search and rescue, emergency medical response), this assumption may not always hold. FRMS should accommodate different modes of working that safely facilitate sleep during duty time where appropriate.

Is patient activation a mediator of the effect of a health promoting intervention in adults at high risk of type 2 diabetes? A longitudinal path model analysis within a randomised trial.

BACKGROUND: Exploring mediators of preventive intervention effects has important implications for the planning of complex interventions. Our aim was to assess the extent to which knowledge, skills and confidence to manage health, here measured as "patient activation", was a mediator of the effect of the intervention "Live your life without diabetes" on weight, waist circumference and systolic blood pressure at 12 months follow-up in adults at high risk of type 2 diabetes. METHODS: Autoregressive path models with three time points of measurement, and contemporaneous and constant b paths were used in a randomised controlled trial (RCT). The RCT took place in a Danish municipal healthcare center and included 127 individuals aged 28 to 70 years with fasting plasma glucose: 6.1-6.9 mmol/l and/or glycated haemoglobin (HbA1c): 42.0-47.9 mmol/mol. Participants were randomised to routine care (n = 64), or intervention (n = 63). The intervention group received an empirical and theory-based intervention delivered over four two-h group sessions during five weeks, and two further sessions after one and six months. The outcomes were weight, waist circumference and systolic blood pressure, and the mediator was patient activation, measured by the self-reported Patient Activation Measure (PAM). Data for the present study was derived from questionnaires and clinical measures from baseline, three- and 12-months follow-up. RESULTS: Mediated effects via PAM on: weight: - 0.09 kg (95% CI - 0.38 to 0.20) out of the total effect - 1.09 kg (95% CI - 3.05 to 0.87); waist circumference: - 0.04 cm (95% CI - 0.36 to 0.28) out of the total effect - 1.86 cm (95% CI - 4.10 to 0.39); and systolic blood pressure: - 0.31 mmHg (- 1.10 to 0.49) out of the total effect - 2.73 mmHg (95% CI - 6.34 to 0.87). CONCLUSION: We found no mediating effects of patient activation as a single variable of the intervention "Live your life without diabetes" on weight, waist circumference and systolic blood pressure at 12 months follow-up in adults at high risk of type 2 diabetes. Our study demonstrates an analytic approach for estimating mediating effects in complex interventions that comply with the criteria on temporal ordered data. Future studies should include possible interacting variables.

Deubiquitinating enzyme amino acid profiling reveals a class of ubiquitin esterases.

The reversibility of ubiquitination by the action of deubiquitinating enzymes (DUBs) serves as an important regulatory layer within the ubiquitin system. Approximately 100 DUBs are encoded by the human genome, and many have been implicated with pathologies, including neurodegeneration and cancer. Non-lysine ubiquitination is chemically distinct, and its physiological importance is emerging. Here, we couple chemically and chemoenzymatically synthesized ubiquitinated lysine and threonine model substrates to a mass spectrometry-based DUB assay. Using this platform, we profile two-thirds of known catalytically active DUBs for threonine esterase and lysine isopeptidase activity and find that most DUBs demonstrate dual selectivity. However, with two anomalous exceptions, the ovarian tumor domain DUB class demonstrates specific (iso)peptidase activity. Strikingly, we find the Machado-Joseph disease (MJD) class to be unappreciated non-lysine DUBs with highly specific ubiquitin esterase activity rivaling the efficiency of the most active isopeptidases. Esterase activity is dependent on the canonical catalytic triad, but proximal hydrophobic residues appear to be general determinants of non-lysine activity. Our findings also suggest that ubiquitin esters have appreciable cellular stability and that non-lysine ubiquitination is an integral component of the ubiquitin system. Its regulatory sophistication is likely to rival that of canonical ubiquitination.

Structural basis for RING-Cys-Relay E3 ligase activity and its role in axon integrity.

MYCBP2 is a ubiquitin (Ub) E3 ligase (E3) that is essential for neurodevelopment and regulates axon maintenance. MYCBP2 transfers Ub to nonlysine substrates via a newly discovered RING-Cys-Relay (RCR) mechanism, where Ub is relayed from an upstream cysteine to a downstream substrate esterification site. The molecular bases for E2-E3 Ub transfer and Ub relay are unknown. Whether these activities are linked to the neural phenotypes is also unclear. We describe the crystal structure of a covalently trapped E2~Ub:MYCBP2 transfer intermediate revealing key structural rearrangements upon E2-E3 Ub transfer and Ub relay. Our data suggest that transfer to the dynamic upstream cysteine, whilst mitigating lysine activity, requires a closed-like E2~Ub conjugate with tempered reactivity, and Ub relay is facilitated by a helix-coil transition. Furthermore, neurodevelopmental defects and delayed injury-induced degeneration in RCR-defective knock-in mice suggest its requirement, and that of substrate esterification activity, for normal neural development and programmed axon degeneration.

Testing the "Zero-Sum Game" Hypothesis: An Examination of School Health Policies and Practices and Inequalities in Educational Outcomes.

BACKGROUND: Health and education are intrinsically linked, while both are significantly patterned by socioeconomic status throughout the life course. Nevertheless, the impact of promoting health via schools on education is seen by some as a "zero-sum game"; ie, focusing resources on health improvement activity distracts schools from their core business of educating pupils, potentially compromising educational attainment. There is emerging evidence that school health improvement interventions may beneficially influence both health and attainment. However, few studies have examined the relationship between school health improvement activity and socioeconomic inequalities in educational attainment. METHODS: Wales-wide, school-level survey data on school health policies and practices was linked with routinely collected data on academic attainment. Primary outcomes included attendance and academic attainment at age 14 (Key Stage 3) and 16 (Key Stage 4). Linear regression models were constructed separately for high and low Free School Meal (FSM) schools, adjusting for confounders. Interaction terms were fitted to test whether there was an interaction between FSM, health improvement activity, and outcomes. RESULTS: There were positive associations between almost all school health variables and KS3 attainment among high, but not low FSM schools. Similarly, for attendance, there were positive associations of several health variables among high but not low FSM schools. There were no associations for KS4 attainment. CONCLUSIONS: Our findings did not support the "zero-sum game" hypothesis; in fact, among more deprived schools there was a tendency for better attendance and attainment at age 14 in schools with more embedded health improvement action.

Photocrosslinking Activity-Based Probes for Ubiquitin RING E3 Ligases.

Activity-based protein profiling is an invaluable technique for studying enzyme biology and facilitating the development of therapeutics. Ubiquitin E3 ligases (E3s) are one of the largest enzyme families and regulate a host of (patho)physiological processes. The largest subtype are the RING E3s of which there are >600 members. RING E3s have adaptor-like activity that can be subject to diverse regulatory mechanisms and have become attractive drug targets. Activity-based probes (ABPs) for measuring RING E3 activity do not exist. Here we re-engineer ubiquitin-charged E2 conjugating enzymes to produce photocrosslinking ABPs. We demonstrate activity-dependent profiling of two divergent cancer-associated RING E3s, RNF4 and c-Cbl, in response to their native activation signals. We also demonstrate profiling of endogenous RING E3 ligase activation in response to epidermal growth factor (EGF) stimulation. These photocrosslinking ABPs should advance E3 ligase research and the development of selective modulators against this important class of enzymes.

Cellular IP6 Levels Limit HIV Production while Viruses that Cannot Efficiently Package IP6 Are Attenuated for Infection and Replication.

HIV-1 hijacks host proteins to promote infection. Here we show that HIV is also dependent upon the host metabolite inositol hexakisphosphate (IP6) for viral production and primary cell replication. HIV-1 recruits IP6 into virions using two lysine rings in its immature hexamers. Mutation of either ring inhibits IP6 packaging and reduces viral production. Loss of IP6 also results in virions with highly unstable capsids, leading to a profound loss of reverse transcription and cell infection. Replacement of one ring with a hydrophobic isoleucine core restores viral production, but IP6 incorporation and infection remain impaired, consistent with an independent role for IP6 in stable capsid assembly. Genetic knockout of biosynthetic kinases IPMK and IPPK reveals that cellular IP6 availability limits the production of diverse lentiviruses, but in the absence of IP6, HIV-1 packages IP5 without loss of infectivity. Together, these data suggest that IP6 is a critical cofactor for HIV-1 replication.

Use of the School Setting During the Summer Holidays: Mixed-Methods Evaluation of Food and Fun Clubs in Wales.

OBJECTIVE: School summer holiday clubs in deprived areas of Wales were evaluated to examine opportunities for healthy eating and physical activity and explore delivery processes. METHODS: Ten Food and Fun clubs participated in 2016. Quantitative data (child and parent surveys; N = 196, N = 84) assessed the opportunity to provide children with breakfast and lunch. A sub-sample of children wore an accelerometer (N = 41) to evaluate the opportunity for achieving 1-hour of moderate to vigorous activity (MVPA) at club. Features of successful club delivery were identified through; focus groups (child and parent; N = 74, N = 69) and interviews (staff/volunteer; N = 32). RESULTS: Opportunities for healthy eating were delivered with high fidelity: 86% of children reported breakfast consumption and 75% eating a healthy lunch. On club days, children reported consuming fewer sugary snacks (66%), fewer sugary drinks (81%), and more fruits and vegetables (67%). About 71% of children achieved the recommended MVPA at club, with children engaging in more MVPA (+17 minutes/day, p < .01) on average compared to non-club days. Successful delivery processes were: use of school facilities and staff; flexible partnership-working; and whole family involvement. CONCLUSIONS: Schools appear to provide a suitable setting for the delivery of healthy eating and physical activity opportunities during school summer holidays.

Role theory of schools and adolescent health.

Evidence that health and education are interlinked is transforming investment in adolescent health. However, no comprehensive theory of how schools influence mental and physical health, which could guide, and be tested through, empirical studies, exists. Using neuroscience, sociology, and other disciplines, we theorise that schools catering for students age 11-18 years can influence health by affecting the behavioural roles that are available for students to perform, the resources available to influence student behaviour, and how peers and teachers (known as the audience) respond. Some schools offer opportunities for students to adopt diverse pro-school roles and to maintain these roles via constructive feedback. Other schools focus narrowly on high academic attainment. Where pro-school roles are unavailable, are beyond students' resources, or elicit negative responses from teachers and peers, students might experience anxiety and choose to adopt anti-school roles, particularly in later adolescence. Behaviours that harm health, such as violence and drug use, are central to anti-school roles because they can facilitate belonging and status within anti-school peer groups and symbolise alternative transitions to adulthood.

Interventions integrating health and academic interventions to prevent substance use and violence: a systematic review and synthesis of process evaluations.

BACKGROUND: Within increasingly constrained school timetables, interventions that integrate academic and health education to reduce substance use and violence may hold promise as a category of intervention that can positively affect both academic and health outcomes. There are no current systematic reviews exploring the effectiveness of such interventions or factors that affect their implementation. METHODS: A total of 19 bibliographic databases and 32 websites were searched. References were also extracted from the reference lists of included studies, and experts and authors were contacted to identify relevant studies. We included reports with no restrictions on language or date. References were screened on title/abstract and those not thus excluded were screened on full report. Data extraction and appraisal followed the Critical Appraisal Skills Programme, Evidence for Policy and Practice Information and Co-ordinating Centre and Cochrane tools. Extracted process data were qualitatively meta-synthesised for common themes. RESULTS: Seventy-eight thousand four hundred fifty-one unique references were identified, and 62 reports were included. A total of 16 reports (reporting on 15 studies of 12 interventions) evaluated process. Key facilitators of integrated academic and health curricula were supportive senior management and alignment of the intervention with school ethos; a positive teaching environment, including positive perceptions around the ability to be flexible in the adaptation and delivery of integrated academic and health curricula; positive pre-existing student and teacher attitudes towards intervention content; and parental support of interventions, largely through reinforcement of messaging at home. Important barriers were over-burdened teachers, with little time to learn and implement integrated curricula. CONCLUSION: Several useful facilitating and inhibiting factors linked to the implementation of interventions that integrate academic and health education for reduced substance use and/or violence were identified, providing tentative but insightful evidence of context-specific issues that may impact intervention success. However, overall, there is still a considerable gap in our understanding of how to achieve the successful implementation of these interventions.

Trivalent RING Assembly on Retroviral Capsids Activates TRIM5 Ubiquitination and Innate Immune Signaling.

TRIM5 is a RING domain E3 ubiquitin ligase with potent antiretroviral function. TRIM5 assembles into a hexagonal lattice on retroviral capsids, causing envelopment of the infectious core. Concomitantly, TRIM5 initiates innate immune signaling and orchestrates disassembly of the viral particle, yet how these antiviral responses are regulated by capsid recognition is unclear. We show that hexagonal assembly triggers N-terminal polyubiquitination of TRIM5 that collectively drives antiviral responses. In uninfected cells, N-terminal monoubiquitination triggers non-productive TRIM5 turnover. Upon TRIM5 assembly on virus, a trivalent RING arrangement allows elongation of N-terminally anchored K63-linked ubiquitin chains (N-K63-Ub). N-K63-Ub drives TRIM5 innate immune stimulation and proteasomal degradation. Inducing ubiquitination before TRIM5 assembly triggers premature degradation and ablates antiviral restriction. Conversely, driving N-K63 ubiquitination after TRIM5 assembly enhances innate immune signaling. Thus, the hexagonal geometry of TRIM5's antiviral lattice converts a capsid-binding protein into a multifunctional antiviral platform.

A systematic review and synthesis of theories of change of school-based interventions integrating health and academic education as a novel means of preventing violence and substance use among students.

BACKGROUND: Schools can play an important role in promoting health. However, many education policies and institutions are increasingly emphasising academic attainment targets, which appear to be diminishing the time available for health education lessons. Interventions that integrate both health and academic learning may present an ideal solution, simultaneously addressing health education and academic development. The theories of change underlying these interventions are therefore of interest, but are poorly studied. METHODS: A systematic review of evaluations of interventions that integrate academic and health education for reduced substance use and/or violence was carried out. As part of this, reports describing theory were assessed for quality and data extracted. Theoretical data were synthesised within and across individual interventions using reciprocal translation and meta-ethnographic line of argument synthesis to produce an overall theory of change for interventions that integrate health and academic education to prevent substance use and violence. RESULTS: Forty-eight reports provided theoretical descriptions of 18 interventions. An overarching theory that emerged was that eroding 'boundaries' at multiple and mutually reinforcing levels-by integrating academic and health education, by transforming relationships between teachers and students, by generalising learning from classrooms to the wider school environment and by ensuring consistent messages from schools and families-is intended to lead to the development of a community of engaged students oriented towards pro-social behaviour and away from substance use, violence and other risk behaviours. CONCLUSIONS: Eroding 'boundaries' between health and academic education, teachers and students, classrooms and the wider school and schools and families were seen to be the most critical to establishing new frameworks of family, classroom or school organisation that are conducive to promoting both academic and social-emotional outcomes. Whether such interventions are feasible to implement and effective in reducing risk behaviours will be examined in other reports arising from the review.

Effects of the Learning Together intervention on bullying and aggression in English secondary schools (INCLUSIVE): a cluster randomised controlled trial.

BACKGROUND: Bullying, aggression, and violence among children and young people are some of the most consequential public mental health problems. We tested the Learning Together intervention, which involved students in efforts to modify their school environment using restorative practice and by developing social and emotional skills. METHODS: We did a cluster randomised trial, with economic and process evaluations, of the Learning Together intervention compared with standard practice (controls) over 3 years in secondary schools in south-east England. Learning Together consisted of staff training in restorative practice; convening and facilitating a school action group; and a student social and emotional skills curriculum. Primary outcomes were self-reported experience of bullying victimisation (Gatehouse Bullying Scale; GBS) and perpetration of aggression (Edinburgh Study of Youth Transitions and Crime (ESYTC) school misbehaviour subscale) measured at 36 months. We analysed data using intention-to-treat longitudinal mixed-effects models. This trial was registered with the ISRCTN registry (10751359). FINDINGS: We included 40 schools (20 in each group); no schools withdrew. 6667 (93·6%) of 7121 students participated at baseline and 5960 (83·3%) of 7154 at 36 months. Mean GBS bullying score at 36 months was 0·34 (SE 0·02) in the control group versus 0·29 (SE 0·02) in the intervention group, with a significant adjusted mean difference (-0·03, 95% CI -0·06 to -0·001; adjusted effect size -0·08). Mean ESYTC score at 36 months was 4·33 (SE 0·20) in the control group versus 4·04 (0·21) in the intervention group, with no evidence of a difference between groups (adjusted difference -0·13, 95% CI -0·43 to 0·18; adjusted effect size -0·03). Costs were an additional £58 per pupil in intervention schools than in control schools. INTERPRETATION: Learning Together had small but significant effects on bullying, which could be important for public health, but no effect on aggression. Interventions to promote student health by modifying the whole-school environment are likely to be one of the most feasible and efficient ways of addressing closely related risk and health outcomes in children and young people. FUNDING: National Institute for Health Research, Educational Endowment Foundation.

Integration of academic and health education for the prevention of physical aggression and violence in young people: systematic review, narrative synthesis and intervention components analysis.

OBJECTIVES: To systematically review evidence on the effectiveness of interventions including integration of academic and health education for reducing physical aggression and violence, and describe the content of these interventions. DATA SOURCES: Between November and December 2015, we searched 19 databases and 32 websites and consulted key experts in the field. We updated our search in February 2018. ELIGIBILITY CRITERIA: We included randomised trials of school-based interventions integrating academic and health education in students aged 4-18 and not targeted at health-related subpopulations (eg, learning or developmental difficulties). We included evaluations reporting a measure of interpersonal violence or aggression. DATA EXTRACTION AND ANALYSIS: Data were extracted independently in duplicate, interventions were analysed to understand similarities and differences and outcomes were narratively synthesised by key stage (KS). RESULTS: We included 13 evaluations of 10 interventions reported in 20 papers. Interventions included either full or partial integration, incorporated a variety of domains beyond the classroom, and used literature, local development or linking of study skills and health promoting skills. Evidence was concentrated in KS2, with few evaluations in KS3 or KS4, and evaluations had few consistent effects; evaluations in KS3 and KS4 did not suggest effectiveness. DISCUSSION: Integration of academic and health education may be a promising approach, but more evidence is needed. Future research should consider the 'lifecourse' aspects of these interventions; that is, do they have a longitudinal effect? Evaluations did not shed light on the value of different approaches to integration.

JACK trial protocol: a phase III multicentre cluster randomised controlled trial of a school-based relationship and sexuality education intervention focusing on young male perspectives.

INTRODUCTION: Teenage pregnancy remains a worldwide health concern which is an outcome of, and contributor to, health inequalities. The need for gender-aware interventions with a focus on males in addressing teenage pregnancy has been highlighted as a global health need by WHO and identified in systematic reviews of (relationship and sexuality education (RSE)). This study aims to test the effectiveness of an interactive film-based RSE intervention, which draws explicit attention to the role of males in preventing an unintended pregnancy by reducing unprotected heterosexual teenage sex among males and females under age 16 years. METHODS AND ANALYSIS: A phase III cluster randomised trial with embedded process and economic evaluations. If I Were Jack encompasses a culturally sensitive interactive film, classroom materials, a teacher-trainer session and parent animations and will be delivered to replace some of the usual RSE for the target age group in schools in the intervention group. Schools in the control group will not receive the intervention and will continue with usual RSE. Participants will not be blinded to allocation. Schools are the unit of randomisation stratified per country and socioeconomic status. We aim to recruit 66 UK schools (24 in Northern Ireland; 14 in each of England, Scotland and Wales), including approximately 7900 pupils. A questionnaire will be administered at baseline and at 12-14 months postintervention. The primary outcome is reported unprotected sex, a surrogate measure associated with unintended teenage pregnancy. Secondary outcomes include knowledge, attitudes, skills and intentions relating to avoiding teenage pregnancy in addition to frequency of engagement in sexual intercourse, contraception use and diagnosis of sexually transmitted infections. ETHICS AND DISSEMINATION: Ethical approval was obtained from Queen's University Belfast. Results will be published in peer-reviewed journals and disseminated to stakeholders. Funding is from the National Institute for Health Research. TRIAL REGISTRATION NUMBER: ISRCTN99459996.

Truncation- and motif-based pan-cancer analysis reveals tumor-suppressing kinases.

A major challenge in cancer genomics is identifying "driver" mutations from the many neutral "passenger" mutations within a given tumor. To identify driver mutations that would otherwise be lost within mutational noise, we filtered genomic data by motifs that are critical for kinase activity. In the first step of our screen, we used data from the Cancer Cell Line Encyclopedia and The Cancer Genome Atlas to identify kinases with truncation mutations occurring within or before the kinase domain. The top 30 tumor-suppressing kinases were aligned, and hotspots for loss-of-function (LOF) mutations were identified on the basis of amino acid conservation and mutational frequency. The functional consequences of new LOF mutations were biochemically validated, and the top 15 hotspot LOF residues were used in a pan-cancer analysis to define the tumor-suppressing kinome. A ranked list revealed MAP2K7, an essential mediator of the c-Jun N-terminal kinase (JNK) pathway, as a candidate tumor suppressor in gastric cancer, despite its mutational frequency falling within the mutational noise for this cancer type. The majority of mutations in MAP2K7 abolished its catalytic activity, and reactivation of the JNK pathway in gastric cancer cells harboring LOF mutations in MAP2K7 or the downstream kinase JNK suppressed clonogenicity and growth in soft agar, demonstrating the functional relevance of inactivating the JNK pathway in gastric cancer. Together, our data highlight a broadly applicable strategy to identify functional cancer driver mutations and define the JNK pathway as tumor-suppressive in gastric cancer.

How Do Contextual Factors Influence Implementation and Receipt of Positive Youth Development Programs Addressing Substance Use and Violence? A Qualitative Meta-Synthesis of Process Evaluations.

OBJECTIVE: Positive youth development (PYD) often aims to prevent tobacco, alcohol, and drugs use and violence. We systematically reviewed PYD interventions, synthesizing process, and outcomes evidence. Synthesis of outcomes, published elsewhere, found no overall evidence of reducing substance use or violence but notable variability of fidelity. Our synthesis of process evaluations examined how implementation varied and was influenced by context. DATA SOURCE: Process evaluations of PYD aiming to reduce substance use and violence. Study Inclusion Criteria: Overall review published since 1985; written in English; focused on youth aged 11 to 18 years; focused on interventions addressing multiple positive assets; reported on theory, process, or outcomes; and concerned with reducing substance use or violence. Synthesis of process evaluations examined how implementation varies with or is influenced by context. DATA EXTRACTION: Two reviewers in parallel. DATA SYNTHESIS: Thematic synthesis. RESULTS: We identified 12 reports. Community engagement enhanced program appeal. Collaboration with other agencies could broaden the activities offered. Calm but authoritative staff increased acceptability. Staff continuity underpinned diverse activities and durable relationships. Empowering participants were sometimes in tension with requiring them to engage in diverse activities. CONCLUSION: Our systematic review identified factors that might help improve the fidelity and acceptability of PYD interventions. Addressing these might enable PYD to fulfill its potential as a means of promoting health.

Intracellular antibody signalling is regulated by phosphorylation of the Fc receptor TRIM21.

Cell surface Fc receptors activate inflammation and are tightly controlled to prevent autoimmunity. Antibodies also simulate potent immune signalling from inside the cell via the cytosolic antibody receptor TRIM21, but how this is regulated is unknown. Here we show that TRIM21 signalling is constitutively repressed by its B-Box domain and activated by phosphorylation. The B-Box occupies an E2 binding site on the catalytic RING domain by mimicking E2-E3 interactions, inhibiting TRIM21 ubiquitination and preventing immune activation. TRIM21 is derepressed by IKKß and TBK1 phosphorylation of an LxxIS motif in the RING domain, at the interface with the B-Box. Incorporation of phosphoserine or a phosphomimetic within this motif relieves B-Box inhibition, promoting E2 binding, RING catalysis, NF-κB activation and cytokine transcription upon infection with DNA or RNA viruses. These data explain how intracellular antibody signalling is regulated and reveal that the B-Box is a critical regulator of RING E3 ligase activity.