Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling.

BACKGROUND: Arginyltransferase (Ate1) orchestrates posttranslational protein arginylation, a pivotal regulator of cellular proteolytic processes. In eukaryotic cells, two interconnected systems-the ubiquitin proteasome system (UPS) and macroautophagy-mediate proteolysis and cooperate to maintain quality protein control and cellular homeostasis. Previous studies have shown that N-terminal arginylation facilitates protein degradation through the UPS. Dysregulation of this machinery triggers p62-mediated autophagy to ensure proper substrate processing. Nevertheless, how Ate1 operates through this intricate mechanism remains elusive. METHODS: We investigated Ate1 subcellular distribution through confocal microscopy and biochemical assays using cells transiently or stably expressing either endogenous Ate1 or a GFP-tagged Ate1 isoform transfected in CHO-K1 or MEFs, respectively. To assess Ate1 and p62-cargo clustering, we analyzed their colocalization and multimerization status by immunofluorescence and nonreducing immunoblotting, respectively. Additionally, we employed Ate1 KO cells to examine the role of Ate1 in autophagy. Ate1 KO MEFs cells stably expressing GFP-tagged Ate1-1 isoform were used as a model for phenotype rescue. Autophagy dynamics were evaluated by analyzing LC3B turnover and p62/SQSTM1 levels under both steady-state and serum-starvation conditions, through immunoblotting and immunofluorescence. We determined mTORC1/AMPk activation by assessing mTOR and AMPk phosphorylation through immunoblotting, while mTORC1 lysosomal localization was monitored by confocal microscopy. RESULTS: Here, we report a multifaceted role for Ate1 in the autophagic process, wherein it clusters with p62, facilitates autophagic clearance, and modulates its signaling. Mechanistically, we found that cell-specific inactivation of Ate1 elicits overactivation of the mTORC1/AMPk signaling hub that underlies a failure in autophagic flux and subsequent substrate accumulation, which is partially rescued by ectopic expression of Ate1. Statistical significance was assessed using a two-sided unpaired t test with a significance threshold set at P<0.05. CONCLUSIONS: Our findings uncover a critical housekeeping role of Ate1 in mTORC1/AMPk-regulated autophagy, as a potential therapeutic target related to this pathway, that is dysregulated in many neurodegenerative and cancer diseases.

Assessment of a new home-based care pathway for children newly diagnosed with type 1 diabetes.

AIM: To compare the outcomes of home-based and conventional hospital-based care for children newly diagnosed with type 1 diabetes mellitus. METHODS: A descriptive study was conducted of all children newly diagnosed with diabetes mellitus at the Timone Hospital in Marseille, France, between November 2017 and July 2019. The patients received either home-based or in-patient hospital care. The primary outcome was the length of initial hospital stay. The secondary outcome measures were glycemic control in the first year of treatment, families' diabetes knowledge, the effect of diabetes on quality of life, and overall quality of care. RESULTS: A total of 85 patients were included, 37 in the home-based care group and 48 in the in-patient care group. The initial length of hospital stay was 6 days in the home-based care group versus 9 days in the in-patient care group. Levels of glycemic control, diabetes knowledge and quality of care were comparable in the two groups despite a higher rate of socioeconomic deprivation in the home-based care group. CONCLUSION: Home-based care for children with diabetes is safe and effective. This new healthcare pathway provides good overall social care, especially for socioeconomically deprived families.

The clustering of physical health conditions and associations with co-occurring mental health problems and problematic alcohol use: a cross-sectional study.

BACKGROUND: There is strong evidence for the co-occurrence of mental health conditions and alcohol problems, yet physical health outcomes among this group are not well characterised. This study aimed to identify clusters of physical health conditions and their associations with mental health and problematic alcohol use in England's general population. METHODS: Cross-sectional analysis of the 2014 Adult Psychiatric Morbidity Survey (N = 7546) was conducted. The survey used standardised measures of problematic alcohol use and mental health conditions, including the Alcohol Use Disorders Identification Test (AUDIT) and the Clinical Interview Schedule-Revised. Participants self-reported any lifetime physical health conditions. Latent class analysis considered 12 common physical illnesses to identify clusters of multimorbidity. Multinomial logistic regression (adjusting for age, gender, ethnicity, education, and occupational grade) was used to explore associations between mental health, hazardous drinking (AUDIT 8 +), and co-occurring physical illnesses. RESULTS: Five clusters were identified with statistically distinct and clinically meaningful disease patterns: 'Physically Healthy' (76.62%), 'Emerging Multimorbidity' (3.12%), 'Hypertension & Arthritis' (14.28%), 'Digestive & Bowel Problems'' (3.17%), and 'Complex Multimorbidity' (2.8%). Having a mental health problem was associated with increased odds of 'Digestive & Bowel Problems' (adjusted multinomial odds ratio (AMOR) = 1.58; 95% CI [1.15-2.17]) and 'Complex Multimorbidity' (AMOR = 2.02; 95% CI [1.49-2.74]). Individuals with co-occurring mental health conditions and problematic alcohol use also had higher odds of 'Digestive & Bowel Problems' (AMOR = 2.64; 95% CI [1.68-4.15]) and 'Complex Multimorbidity' (AMOR = 2.62; 95% CI [1.61-4.23]). CONCLUSIONS: Individuals with a mental health condition concurrent with problematic alcohol use experience a greater burden of physical illnesses, highlighting the need for timely treatment which is likely to include better integration of alcohol and mental health services.

The 19S proteasome subunit Rpt5 reversibly associates with cold-stable microtubules in glial cells at low temperatures.

The ubiquitin-proteasome system (UPS) degrades intracellular proteins through the 26S proteasome. We analysed how cold stress affects the UPS in glial cells. Together with a reduction in the 20S proteolytic activity and increased levels of polyubiquitinated proteins, exposure of glial cell cultures to cold induces a partial disassembly of the 26S proteasome. In particular, we found that Rpt5, a subunit of the 19S proteasome, relocates to cold-stable microtubules, although no apparent cytoskeletal redistribution was detected for other analysed subunits of the 19S or 20S complexes. Furthermore, we demonstrate that both the expression of the microtubule-associated protein MAP6 and the post-translational acetylation of α-tubulin modulate the association of Rpt5 with microtubules. This reversible association could be related to functional preservation of the proteolytic complex during cold stress.

Spontaneous rupture of an aneurysm of the superior thyroid artery: A rare cause of acute respiratory failure.

Ruptured aneurysm of superior thyroid artery is a very rare pathological event. Underlying causes such as trauma, malignancy or iatrogenic are not systematically found. Resulting cervical hematoma is life threatening and can lead to acute respiratory failure, dysphagia, vocal cord paralysis and hemomediastinum. Endovascular treatment combined with surgical drainage has been described as an effective treatment. In this case report, a 63-year-old man presented a spontaneous rupture of an aneurysm of superior thyroid artery resulting in cervical hematoma and acute respiratory failure.

Ablation of arginyl-tRNA-protein transferase in oligodendrocytes impairs central nervous system myelination.

Addition of arginine (Arg) from tRNA can cause major alterations of structure and function of protein substrates. This post-translational modification, termed protein arginylation, is mediated by the enzyme arginyl-tRNA-protein transferase 1 (Ate1). Arginylation plays essential roles in a variety of cellular processes, including cell migration, apoptosis, and cytoskeletal organization. Ate1 is associated with neuronal functions such as neurogenesis and neurite growth. However, the role of Ate1 in glial development, including oligodendrocyte (OL) differentiation and myelination processes in the central nervous system, is poorly understood. The present study revealed a peak in Ate1 protein expression during myelination process in primary cultured OLs. Post-transcriptional downregulation of Ate1 reduced the number of OL processes, and branching complexity, in vitro. We conditionally ablated Ate1 from OLs in mice using 2',3'-cyclic nucleotide 3'-phosphodiesterase-Cre promoter ("Ate1-KO" mice), to assess the role of Ate1 in OL function and axonal myelination in vivo. Immunostaining for OL differentiation markers revealed a notable reduction of mature OLs in corpus callosum of 14-day-old Ate1-KO, but no changes in spinal cord, in comparison with wild-type controls. Local proliferation of OL precursor cells was elevated in corpus callosum of 21-day-old Ate1-KO, but was unchanged in spinal cord. Five-month-old Ate1-KO displayed reductions of mature OL number and myelin thickness, with alterations of motor behaviors. Our findings, taken together, demonstrate that Ate1 helps maintain proper OL differentiation and myelination in corpus callosum in vivo, and that protein arginylation plays an essential role in developmental myelination.

Copeptin assays in children for the differential diagnosis of polyuria-polydipsia syndrome and reference levels in hospitalized children.

OBJECTIVES: Polyuria-polydipsia syndrome (PPS) is a common presentation in children but the differential diagnosis rests on burdensome water deprivation tests. The aims of this study were to determine a copeptin threshold to distinguish patients with central diabetes insipidus from those with primary polydipsia and to estimate the normal range of copeptin concentrations in children. DESIGN: Single-centre retrospective descriptive study. PATIENTS: Two hundred and seventy-eight children aged 2 months to 18 years who consulted for PPS (N = 40) or other reasons (control group, N = 238) at La Timone University Hospital in Marseille, France, between April 2015 and September 2019 and had a copeptin assay. MEASUREMENTS: Ultrasensitive copeptin assays on blood samples. RESULTS: Among the children with PPS, the mean copeptin concentrations were 1.72, 55.2 and 15.7 pmol/l in those with central diabetes insipidus (N = 21), nephrogenic diabetes insipidus (N = 3), and primary polydipsia (N = 16), respectively. Copeptin levels lower than 3.53 pmol/l were diagnostic of central diabetes insipidus with 100% sensitivity and 87.4% specificity (p < .001). The 5th-95th copeptin percentile range in the control group was 2.53-21.03 pmol/L. Copeptin levels were significantly higher in boys than in girls but there was no association with age, pubertal stage, body mass index, or the reason for consulting. CONCLUSIONS: Our results indicate copeptin assays may be valuable in the differential diagnosis of PPS in children. Larger prospective studies are required to establish their accuracy in everyday clinical practice.

Analysis of sex differences in the clinical presentation, management and prognosis of infective endocarditis in Spain.

INTRODUCTION: Sex-dependent differences of infective endocarditis (IE) have been reported. Women suffer from IE less frequently than men and tend to present more severe manifestations. Our objective was to analyse the sex-based differences of IE in the clinical presentation, treatment, and prognosis. MATERIAL AND METHODS: We analysed the sex differences in the clinical presentation, modality of treatment and prognosis of IE in a national-level multicentric cohort between 2008 and 2018. All data were prospectively recorded by the GAMES cohort (Spanish Collaboration on Endocarditis). RESULTS: A total of 3451 patients were included, of whom 1105 were women (32.0%). Women were older than men (mean age, 68.4 vs 64.5). The most frequently affected valves were the aortic valve in men (50.6%) and mitral valve in women (48.7%). Staphylococcus aureus aetiology was more frequent in women (30.1% vs 23.1%; p<0.001).Surgery was performed in 38.3% of women and 50% of men. After propensity score (PS) matching for age and estimated surgical risk (European System for Cardiac Operative Risk Evaluation II (EuroSCORE II)), the analysis of the matched cohorts revealed that women were less likely to undergo surgery (OR 0.74; 95% CI 0.59 to 0.91; p=0.05).The observed overall in-hospital mortality was 32.8% in women and 25.7% in men (OR for the mortality of female sex 1.41; 95% CI 1.21 to 1.65; p<0.001). This statistical difference was not modified after adjusting for all possible confounders. CONCLUSIONS: Female sex was an independent factor related to mortality after adjusting for confounders. In addition, women were less frequently referred for surgical treatment.

Development and validation of a multivariable risk prediction model for head and neck cancer using the UK Biobank.

Head and neck cancer (HNC) is the eighth most common cancer in the UK, with over 12,000 new cases every year. The incidence of HNC is predicted to increase by 33% by 2035. Risk modelling produces personalised risk estimates for specific diseases, which can be used to inform education, screening programmes and recruitment to clinical trials. The present study describes the development and validation of the first risk prediction model for absolute risk of HNC, using a nested case­control study within the UK Biobank dataset. The UK Biobank recruited 502,647 individuals aged 40­69 years from around the UK. In total, 859 cases of HNC were identified, with 253 incident cases (individuals who developed HNC in the 7 years following recruitment to the UK Biobank study). Logistic regression was used to develop the model, then the model performance was validated using a cohort from the North West of England. Overall, increasing age, male sex, positive history of smoking and alcohol consumption and higher levels of material deprivation were significantly associated with a higher risk of HNC. Consuming at least five portions of fruit and vegetables per day, exercising at least once per week and higher BMI offered a protective effect against HNC. The C­statistic was 0.69 [95% confidence interval (CI), 0.66­0.71] and the model displayed good calibration. Upon external validation, the C­statistic was 0.64 (95% CI, 0.60­0.68) with reasonable calibration. The model developed and validated in the present study allows calculation of a personalised risk estimate for HNC. This could be used to guide clinicians when counselling individuals on risk behaviour, and there is potential for such models to inform recruitment to screening trials.

Arginylated Calreticulin Increases Apoptotic Response Induced by Bortezomib in Glioma Cells.

After retrotranslocation from the endoplasmic reticulum to the cytoplasm, calreticulin is modified by the enzyme arginyltransferase-1 (ATE1). Cellular levels of arginylated calreticulin (R-CRT) are regulated in part by the proteasomal system. Under various stress conditions, R-CRT becomes associated with stress granules (SGs) or reaches the plasma membrane (PM), where it participates in pro-apoptotic signaling. The mechanisms underlying the resistance of tumor cells to apoptosis induced by specific drugs remain unclear. We evaluated the regulatory role of R-CRT in apoptosis of human glioma cell lines treated with the proteasome inhibitor bortezomib (BT). Two cell lines (HOG, MO59K) displaying distinctive susceptibility to apoptosis induction were studied further. BT efficiency was found to be correlated with a subcellular distribution of R-CRT. In MO59K (apoptosis-resistant), R-CRT was confined to SGs formed following BT treatment. In contrast, HOG (apoptosis-susceptible) treated with BT showed lower SG formation and higher levels of cytosolic and PM R-CRT. Increased R-CRT level was associated with enhanced mobilization of intracellular Ca2+ and with sustained apoptosis activation via upregulation of cell death receptor DR5. R-CRT overexpression in the cytoplasm of MO59K rendered the cells susceptible to BT-induced, DR5-mediated cell death. Our findings suggest that R-CRT plays an essential role in the effect of BT treatment on tumor cells and that ATE1 is a strong candidate target for future studies of cancer diagnosis and therapy.

Relevance of protein-protein interactions on the biological identity of nanoparticles.

Considering that the use of nanoparticles (NPs) as carriers of therapeutic or theranostic agents has increased in the last years, it is mandatory to understand the interaction between NPs and living systems. In contact with biological fluids, the NPs (synthetic identity) are covered with biomolecules that form a protein corona, which defines the biological identity. It is well known that the protein corona formation is mediated by non-specific physical interactions, but protein-protein interactions (PPI), involving specific recognition sites of the polypeptides, are also involved. This work explores the relationship between the synthetic and biological identities of layered double hydroxides nanoparticles (LDH-NPs) and the effect of the protein corona on the cellular response. With such a purpose, the synthetic identity was modified by coating LDH-NPs with either a single protein or a complex mixture of them, followed by the characterization of the protein corona formed in a commonly used cell culture medium. A proteomic approach was used to identify the protein corona molecules and the PPI network was constructed with a novel bioinformatic tool. The coating on LDH-NPs defines the biological identity in such a way that the composition of the protein corona as well as PPI are changed. Electrostatic interactions appear not to be the only driving force regulating the interactions between NPs, proteins and cells since the specific recognition also play a fundamental role. However, the biological identity of LDH-NPs does not affect the interactions with cells that shows negligible cytotoxicity and high internalization levels.