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2.
Biochem Soc Trans ; 51(3): 959-969, 2023 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-37318088

RESUMO

Protein synthesis is essential to support homeostasis, and thus, must be highly regulated during cellular response to harmful environments. All stages of translation are susceptible to regulation under stress, however, the mechanisms involved in translation regulation beyond initiation have only begun to be elucidated. Methodological advances enabled critical discoveries on the control of translation elongation, highlighting its important role in translation repression and the synthesis of stress-response proteins. In this article, we discuss recent findings on mechanisms of elongation control mediated by ribosome pausing and collisions and the availability of tRNAs and elongation factors. We also discuss how elongation intersects with distinct modes of translation control, further supporting cellular viability and gene expression reprogramming. Finally, we highlight how several of these pathways are reversibly regulated, emphasizing the dynamics of translation control during stress-response progression. A comprehensive understanding of translation regulation under stress will produce fundamental knowledge of protein dynamics while opening new avenues and strategies to overcome dysregulated protein production and cellular sensitivity to stress.


Assuntos
Elongação Traducional da Cadeia Peptídica , Ribossomos , RNA Mensageiro/metabolismo , Ribossomos/metabolismo , Proteínas/metabolismo , RNA de Transferência/metabolismo
3.
Proc Natl Acad Sci U S A ; 117(36): 22157-22166, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32855298

RESUMO

Subpopulations of ribosomes are responsible for fine tuning the control of protein synthesis in dynamic environments. K63 ubiquitination of ribosomes has emerged as a new posttranslational modification that regulates protein synthesis during cellular response to oxidative stress. K63 ubiquitin, a type of ubiquitin chain that functions independently of the proteasome, modifies several sites at the surface of the ribosome, however, we lack a molecular understanding on how this modification affects ribosome structure and function. Using cryoelectron microscopy (cryo-EM), we resolved the first three-dimensional (3D) structures of K63 ubiquitinated ribosomes from oxidatively stressed yeast cells at 3.5-3.2 Å resolution. We found that K63 ubiquitinated ribosomes are also present in a polysome arrangement, similar to that observed in yeast polysomes, which we determined using cryoelectron tomography (cryo-ET). We further showed that K63 ubiquitinated ribosomes are captured uniquely at the rotated pretranslocation stage of translation elongation. In contrast, cryo-EM structures of ribosomes from mutant cells lacking K63 ubiquitin resolved at 4.4-2.7 Å showed 80S ribosomes represented in multiple states of translation, suggesting that K63 ubiquitin regulates protein synthesis at a selective stage of elongation. Among the observed structural changes, ubiquitin mediates the destabilization of proteins in the 60S P-stalk and in the 40S beak, two binding regions of the eukaryotic elongation factor eEF2. These changes would impact eEF2 function, thus, inhibiting translocation. Our findings help uncover the molecular effects of K63 ubiquitination on ribosomes, providing a model of translation control during oxidative stress, which supports elongation halt at pretranslocation.


Assuntos
Estresse Oxidativo , Ribossomos/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/metabolismo , Microscopia Crioeletrônica , Regulação Fúngica da Expressão Gênica , Modelos Moleculares , Mutação
4.
J Biol Chem ; 297(3): 101077, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34391779

RESUMO

Ubiquitin signaling is a conserved, widespread, and dynamic process in which protein substrates are rapidly modified by ubiquitin to impact protein activity, localization, or stability. To regulate this process, deubiquitinating enzymes (DUBs) counter the signal induced by ubiquitin conjugases and ligases by removing ubiquitin from these substrates. Many DUBs selectively regulate physiological pathways employing conserved mechanisms of ubiquitin bond cleavage. DUB activity is highly regulated in dynamic environments through protein-protein interaction, posttranslational modification, and relocalization. The largest family of DUBs, cysteine proteases, are also sensitive to regulation by oxidative stress, as reactive oxygen species (ROS) directly modify the catalytic cysteine required for their enzymatic activity. Current research has implicated DUB activity in human diseases, including various cancers and neurodegenerative disorders. Due to their selectivity and functional roles, DUBs have become important targets for therapeutic development to treat these conditions. This review will discuss the main classes of DUBs and their regulatory mechanisms with a particular focus on DUB redox regulation and its physiological impact during oxidative stress.


Assuntos
Enzimas Desubiquitinantes/genética , Enzimas Desubiquitinantes/metabolismo , Enzimas Desubiquitinantes/fisiologia , Animais , Endopeptidases/metabolismo , Homeostase , Humanos , Oxirredução , Estresse Oxidativo , Processamento de Proteína Pós-Traducional , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Ubiquitina/metabolismo , Ubiquitinação
5.
Int J Mol Sci ; 21(3)2020 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-32050486

RESUMO

The eukaryotic proteome has to be precisely regulated at multiple levels of gene expression, from transcription, translation, and degradation of RNA and protein to adjust to several cellular conditions. Particularly at the translational level, regulation is controlled by a variety of RNA binding proteins, translation and associated factors, numerous enzymes, and by post-translational modifications (PTM). Ubiquitination, a prominent PTM discovered as the signal for protein degradation, has newly emerged as a modulator of protein synthesis by controlling several processes in translation. Advances in proteomics and cryo-electron microscopy have identified ubiquitin modifications of several ribosomal proteins and provided numerous insights on how this modification affects ribosome structure and function. The variety of pathways and functions of translation controlled by ubiquitin are determined by the various enzymes involved in ubiquitin conjugation and removal, by the ubiquitin chain type used, by the target sites of ubiquitination, and by the physiologic signals triggering its accumulation. Current research is now elucidating multiple ubiquitin-mediated mechanisms of translational control, including ribosome biogenesis, ribosome degradation, ribosome-associated protein quality control (RQC), and redox control of translation by ubiquitin (RTU). This review discusses the central role of ubiquitin in modulating the dynamism of the cellular proteome and explores the molecular aspects responsible for the expanding puzzle of ubiquitin signals and functions in translation.


Assuntos
Biossíntese de Proteínas , Ubiquitina/metabolismo , Ubiquitinação , Animais , Humanos , Modelos Moleculares , Estresse Oxidativo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise , Estabilidade de RNA , Proteínas Ribossômicas/metabolismo , Ribossomos/metabolismo
6.
J Proteome Res ; 18(1): 309-318, 2019 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-30489083

RESUMO

During oxidative stress, K63-linked polyubiquitin chains modify a variety of proteins including ribosomes. Knowledge of the precise sites of K63 ubiquitin is key to understand its function during the response to stress. To identify the sites of K63 ubiquitin, we developed a new mass spectrometry based method that quantified >1100 K63 ubiquitination sites in yeast that responded to oxidative stress induced by H2O2. We determined that under stress, K63 ubiquitin-modified proteins were involved in several cellular functions including ion transport, protein trafficking, and translation. The most abundant ubiquitin sites localized to the head of the 40S subunit of the ribosome, modified assembled polysomes, and affected the binding of translation factors. The results suggested a new pathway of post-initiation control of translation during oxidative stress and illustrated the importance of high-resolution mapping of noncanonical ubiquitination events.


Assuntos
Proteínas Fúngicas/metabolismo , Estresse Oxidativo , Proteômica/métodos , Ubiquitinação , Sítios de Ligação , Peróxido de Hidrogênio/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Biossíntese de Proteínas , Processamento de Proteína Pós-Traducional , Ribossomos/metabolismo , Ubiquitina/metabolismo , Leveduras
7.
J Dairy Res ; 86(1): 25-33, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30757981

RESUMO

This study aimed to calculate economic values (EVs) and economic selection indices for milk production systems in small rural properties. The traits 305-d milk yield in kg (MY), fat (FP) and protein (PP) percentage, daily fat (FY) and protein (PY) yield, cow live weight in kg (LW), calving interval (CI), and logarithm of daily somatic cell count (SCC) in milk were considered the goals and selection criteria. The production systems were identified from 29 commercial properties based on the inventory of revenues and costs and of zootechnical field data. Later, bioeconomic models were developed to calculate the productive performance, revenues, and costs concerning milk production to estimate EVs, which were calculated as the difference in annual profit with dairy production resulting from a change in one unit of the trait while keeping the others constant and dividing the value by the number of cows. After the EVs were known, ten economic selection indices were estimated for each system so they could be compared by modifying the selection criteria and calculating the relative importance of each selection criteria, the accuracy of the economic selection index, and response expected to the selection in USD, among other parameters. One of the systems detected was called less intensive (LS) and was characterized by having ten cows in lactation that produced 13·5 l/d and consumed 1·8 kg of concentrate/d. The second system detected was called more intensive (IS) and had 22 cows in lactation that produced 17·5 l/d and consumed 3·4 kg of concentrate/d. Monthly profits per cows in lactation of USD 2·60 and USD 68·77 were recorded for LS and IS, respectively. The EVs of the traits MY, FP, and PP were all positive, while for the other traits they were all negative in all situations. The best economic selection indices were those featuring selection criteria MY, LW, and CI, while the trait LW had the greatest importance in both systems. These results indicate that animal frame must be controlled in order to maximize the system's profit.


Assuntos
Cruzamento/economia , Cruzamento/métodos , Bovinos/genética , Indústria de Laticínios/economia , Lactação/genética , Seleção Genética , Animais , Brasil , Bovinos/fisiologia , Contagem de Células , Custos e Análise de Custo , Fazendas , Feminino , Renda , Masculino , Leite/classificação , Leite/citologia , População Rural , Clima Tropical
8.
J Bacteriol ; 200(9)2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29463605

RESUMO

Microbes in biofilms face the challenge of substrate limitation. In particular, oxygen often becomes limited for cells in Pseudomonas aeruginosa biofilms growing in the laboratory or during host colonization. Previously we found that phenazines, antibiotics produced by P. aeruginosa, balance the intracellular redox state of cells in biofilms. Here, we show that genes involved in denitrification are induced in phenazine-null (Δphz) mutant biofilms grown under an aerobic atmosphere, even in the absence of nitrate. This finding suggests that resident cells employ a bet-hedging strategy to anticipate the potential availability of nitrate and counterbalance their highly reduced redox state. Consistent with our previous characterization of aerobically grown colonies supplemented with nitrate, we found that the pathway that is induced in Δphz mutant colonies combines the nitrate reductase activity of the periplasmic enzyme Nap with the downstream reduction of nitrite to nitrogen gas catalyzed by the enzymes Nir, Nor, and Nos. This regulatory relationship differs from the denitrification pathway that functions under anaerobic growth, with nitrate as the terminal electron acceptor, which depends on the membrane-associated nitrate reductase Nar. We identified the sequences in the promoter regions of the nap and nir operons that are required for the effects of phenazines on expression. We also show that specific phenazines have differential effects on nap gene expression. Finally, we provide evidence that individual steps of the denitrification pathway are catalyzed at different depths within aerobically grown biofilms, suggesting metabolic cross-feeding between community subpopulations.IMPORTANCE An understanding of the unique physiology of cells in biofilms is critical to our ability to treat fungal and bacterial infections. Colony biofilms of the opportunistic pathogen Pseudomonas aeruginosa grown under an aerobic atmosphere but without nitrate express a denitrification pathway that differs from that used for anaerobic growth. We report that the components of this pathway are induced by electron acceptor limitation and that they are differentially expressed over the biofilm depth. These observations suggest that (i) P. aeruginosa exhibits "bet hedging," in that it expends energy and resources to prepare for nitrate availability when other electron acceptors are absent, and (ii) cells in distinct biofilm microniches may be able to exchange substrates to catalyze full denitrification.


Assuntos
Proteínas de Bactérias/metabolismo , Biofilmes/efeitos dos fármacos , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Fenazinas/farmacologia , Pseudomonas aeruginosa/efeitos dos fármacos , Proteínas de Bactérias/genética , Biofilmes/crescimento & desenvolvimento , Desnitrificação , Regiões Promotoras Genéticas , Pseudomonas aeruginosa/metabolismo
9.
Phytopathology ; 106(12): 1486-1494, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27532426

RESUMO

Bacterial fruit blotch (BFB), caused by the seedborne bacterium Acidovorax citrulli, is an economically important threat to cucurbitaceous crops worldwide. Since the first report of BFB in Brazil in 1990, outbreaks have occurred sporadically on watermelon and, more frequently, on melon, resulting in significant yield losses. At present, the genetic diversity and the population structure of A. citrulli strains in Brazil remain unclear. A collection of 74 A. citrulli strains isolated from naturally infected tissues of different cucurbit hosts in Brazil between 2000 and 2014 and 18 A. citrulli reference strains from other countries were compared by pulsed-field gel electrophoresis (PFGE), multilocus sequence analysis (MLSA) of housekeeping and virulence-associated genes, and pathogenicity tests on seedlings of different cucurbit species. The Brazilian population comprised predominantly group I strains (98%), regardless of the year of isolation, geographical region, or host. Whole-genome restriction digestion and PFGE analysis revealed that three unique and previously unreported A. citrulli haplotypes (assigned as haplotypes B22, B23, and B24) occurred in Brazil. The greatest diversity of A. citrulli (four haplotypes) was found among strains collected from the northeastern region of Brazil, which accounts for more than 90% of the country's melon production. MLSA clearly distinguished A. citrulli strains into two well-supported clades, in agreement with observations based on PFGE analysis. Five Brazilian A. citrulli strains, representing different group I haplotypes, were moderately aggressive on watermelon seedlings compared with four group II strains that were highly aggressive. In contrast, no significant differences in BFB severity were observed between group I and II A. citrulli strains on melon and squash seedlings. Finally, we observed a differential effect of temperature on in vitro growth of representative group I and II A. citrulli haplotypes. Specifically, of 18 group II strains tested, all grew at 40 and 41°C, whereas only 3 of 15 group I strains (haplotypes B8[P], B3[K], and B15) grew at 40°C. Three strains representing haplotype B8(P) were the only group I strains that grew at 41°C. These results contribute to a better understanding of the genetic diversity of A. citrulli associated with BFB outbreaks in Brazil, and reinforce the efficiency of MLSA and PFGE analysis for assessing population structure. This study also provides the first evidence to suggest that temperature might be a driver in the ecological adaptation of A. citrulli populations.


Assuntos
Citrullus/microbiologia , Comamonadaceae/isolamento & purificação , Cucurbitaceae/microbiologia , Doenças das Plantas/microbiologia , Brasil , Comamonadaceae/classificação , Comamonadaceae/genética , Comamonadaceae/patogenicidade , Produtos Agrícolas , Eletroforese em Gel de Campo Pulsado , Frutas/microbiologia , Genética Populacional , Haplótipos , Tipagem de Sequências Multilocus , Plântula/microbiologia , Temperatura , Virulência
10.
bioRxiv ; 2024 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-38948861

RESUMO

Under stress conditions, cells reprogram their molecular machineries to mitigate damage and promote survival. Ubiquitin signaling is globally increased during oxidative stress, controlling protein fate and supporting stress defenses at several subcellular compartments. However, the rules driving subcellular ubiquitin localization to promote these concerted response mechanisms remain understudied. Here, we show that K63-linked ubiquitin chains, known to promote proteasome-independent pathways, accumulate primarily in non-cytosolic compartments during oxidative stress induced by sodium arsenite in mammalian cells. Our subcellular ubiquitin proteomic analyses of non-cytosolic compartments expanded 10-fold the pool of proteins known to be ubiquitinated during arsenite stress (2,046) and revealed their involvement in pathways related to immune signaling and translation control. Moreover, subcellular proteome analyses revealed proteins that are recruited to non-cytosolic compartments under stress, including a significant enrichment of helper ubiquitin-binding adaptors of the ATPase VCP that processes ubiquitinated substrates for downstream signaling. We further show that VCP recruitment to non-cytosolic compartments under arsenite stress occurs in a ubiquitin-dependent manner mediated by its adaptor NPLOC4. Additionally, we show that VCP and NPLOC4 activities are critical to sustain low levels of non-cytosolic K63-linked ubiquitin chains, supporting a cyclical model of ubiquitin conjugation and removal that is disrupted by cellular exposure to reactive oxygen species. This work deepens our understanding of the role of localized ubiquitin and VCP signaling in the basic mechanisms of stress response and highlights new pathways and molecular players that are essential to reshape the composition and function of the human subcellular proteome under dynamic environments.

11.
Health Syst (Basingstoke) ; 13(1): 48-61, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38370321

RESUMO

Lack of timeliness and capacity are seen as fundamental problems that jeopardise healthcare delivery systems everywhere. Many believe the shortage of medical providers is causing this timeliness problem. This action research presents how one doctor implemented the theory of constraints (TOC) to improve the throughput (quantity of patients treated) of his ophthalmology imaging practice by 64% in a few weeks with little to no expense. The five focusing steps (5FS) guided the TOC implementation - which included the drum-buffer-rope scheduling and buffer management - and occurred in a matter of days. The implementation provided significant bottom-line results almost immediately. This article explains each step of the 5FS in general terms followed by specific applications to healthcare services, as well as the detailed use in this action research. Although TOC successfully addressed the practice problems, this implementation was not sustained after the TOC champion left the organisation. However, this drawback provided valuable knowledge. The article provides insightful knowledge to help readers implement TOC in their environments to provide immediate and significant results at little to no expense.

12.
Cell Rep ; 42(11): 113359, 2023 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-37917585

RESUMO

Oxidative stress causes K63-linked ubiquitination of ribosomes by the E2 ubiquitin conjugase Rad6. How Rad6-mediated ubiquitination of ribosomes affects translation, however, is unclear. We therefore perform Ribo-seq and Disome-seq in Saccharomyces cerevisiae and show that oxidative stress causes ribosome pausing at specific amino acid motifs, which also leads to ribosome collisions. However, these redox-pausing signatures are lost in the absence of Rad6 and do not depend on the ribosome-associated quality control (RQC) pathway. We also show that Rad6 is needed to inhibit overall translation in response to oxidative stress and that its deletion leads to increased expression of antioxidant genes. Finally, we observe that the lack of Rad6 leads to changes during translation that affect activation of the integrated stress response (ISR) pathway. Our results provide a high-resolution picture of the gene expression changes during oxidative stress and unravel an additional stress response pathway affecting translation elongation.


Assuntos
Proteínas de Saccharomyces cerevisiae , Ubiquitina , Ubiquitina/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , gama-Glutamil Hidrolase/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Ribossomos/metabolismo , Estresse Oxidativo
13.
Health Syst (Basingstoke) ; 11(1): 1-16, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35127055

RESUMO

Despite ever-increasing resources devoted to healthcare, lack of capacity and timeliness are still chronic problems worldwide. This systematic review aims to present an overview of the Theory of Constraints (TOC) implementations in healthcare services and their outcomes. We analysed 42 TOC implementations (15 full-text articles, 12 video proceedings, and 2 theses/disserations) from major scientific electronic databases and TOC International Certification Organization Conferences. All implementations reported positive outcomes, both tangible and intangible. The two main improvements reported by authors were in productivity (98%; n = 41) - more patients treated - and in the timeliness of care (83%; n = 35). Furthermore, the selected studies reported dramatic improvements: 50% mean reductions in patient waiting time; 38% reduction in patient length of stay; 43% mean increase in operating room productivity and 34% mean increase in throughput. TOC implementations attained positive results in all levels of the health and social care chain. Most TOC recommendations and changes showed almost immediate results and required little or no additional cost to implement. Evidence supports TOC as a promising solution for the chronic healthcare problem, improving quality and timeliness, both necessary conditions for providing effective healthcare.

14.
Cell Rep ; 39(8): 110860, 2022 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-35613580

RESUMO

Protein ubiquitination is an essential process that rapidly regulates protein synthesis, function, and fate in dynamic environments. Within its non-proteolytic functions, we showed that K63-linked polyubiquitinated conjugates heavily accumulate in yeast cells exposed to oxidative stress, stalling ribosomes at elongation. K63-ubiquitinated conjugates accumulate mostly because of redox inhibition of the deubiquitinating enzyme Ubp2; however, the role and regulation of ubiquitin-conjugating enzymes (E2) in this pathway remained unclear. Here, we show that the E2 Rad6 associates and modifies ribosomes during stress. We further demonstrate that Rad6 and its human homolog UBE2A are redox regulated by forming a reversible disulfide with the E1 ubiquitin-activating enzyme (Uba1). This redox regulation is part of a negative feedback regulation, which controls the levels of K63 ubiquitination under stress. Finally, we show that Rad6 activity is necessary to regulate translation, antioxidant defense, and adaptation to stress, thus providing an additional physiological role for this multifunctional enzyme.


Assuntos
Ribossomos , Enzimas de Conjugação de Ubiquitina , Humanos , Oxirredução , Estresse Oxidativo , Ribossomos/metabolismo , Enzimas Ativadoras de Ubiquitina/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitinação
15.
Artigo em Inglês | MEDLINE | ID: mdl-34639391

RESUMO

The emergency department (ED) crowding is a critical healthcare issue worldwide that leads to long waits and poorer healthcare outcomes. Goldratt's theory of constraints (TOC) has been used effectively to improve such problematic environments for more than three decades. While most TOC solutions are simple, with many viewing them as purely common sense, they represent paradigm shifts in how to manage complex, uncertain, and silo environments. Goldratt used a simple dice game with a straight flow (I-shape) to illustrate the impact of dependent resources and statistical fluctuations in managing resources. Additionally, games help to overcome resistance to change and gain ownership by having participants develop their solutions. This new cooperative game illustrates an ED environment where patients may follow different care pathways according to their clinical needs, timeliness of care is measured in minutes, the demand is highly uncertain, and treatment must frequently start almost immediately. A Monte Carlo simulation validated the TOC solution to this ED game, achieving results similar to the real TOC's implementations. Moreover, this article provides a thorough process to Socratically introduce TOC to healthcare professionals and others to recognize that the EDs' (like other healthcare systems') core problem is the traditional approach to managing them.


Assuntos
Aglomeração , Serviço Hospitalar de Emergência , Atenção à Saúde , Humanos
16.
Artigo em Inglês | MEDLINE | ID: mdl-34574417

RESUMO

This study identifies physical activity (PA) and sedentary behaviour (SB) clusters in preschoolers compliant (C) or non-compliant (NC) with sleep recommendations; and associates these clusters with obesity markers. PA and SB were objectively assessed (Actigraph WGT3-X) in 272 preschoolers (4.4 ± 0.7 years old). Sleep duration was parent-reported, and preschoolers were classified as C (3-4 years old: 600-780 min/day; 5 years old: 540-660 min/day) or NC with sleep recommendations. Body mass index (BMI) and waist circumference (WC) were assessed according to international protocols. Moderate to vigorous physical activity (MVPA) and light physical activity (LPA) were categorized as low/high (<60 min/>60 min/day or <180 min/180 min/day, respectively). SB was defined according to mean values between clusters. Latent profile analysis was performed. Associations between the observed clusters and obesity markers were determined using linear regression (RStudio; 1.3.1073). Four cluster solutions for C and NC preschoolers were identified. A negative association between C/Low MVPA cluster and BMI, and a positive association between NC/Low MVPA and BMI (ß = -0.8, 95%CI = -1.6;-0.1, and ß = 0.9, 95%CI = 0.1;1.7, respectively) were observed. No association was seen for SB clusters. Adequate sleep duration may have a protective role for preschoolers' BMI, even if the children do not comply with MVPA recommendations.


Assuntos
Obesidade , Comportamento Sedentário , Índice de Massa Corporal , Criança , Pré-Escolar , Humanos , Obesidade/epidemiologia , Sono , Circunferência da Cintura
17.
Cells ; 9(6)2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32580388

RESUMO

Ubiquitination is a post-translational modification that regulates cellular processes by altering the interactions of proteins to which ubiquitin, a small protein adduct, is conjugated. Ubiquitination yields various products, including mono- and poly-ubiquitinated substrates, as well as unanchored poly-ubiquitin chains whose accumulation is considered toxic. We previously showed that transgenic, unanchored poly-ubiquitin is not problematic in Drosophila melanogaster. In the fruit fly, free chains exist in various lengths and topologies and are degraded by the proteasome; they are also conjugated onto other proteins as one unit, eliminating them from the free ubiquitin chain pool. Here, to further explore the notion of unanchored chain toxicity, we examined when free poly-ubiquitin might become problematic. We found that unanchored chains can be highly toxic if they resemble linear poly-ubiquitin that cannot be modified into other topologies. These species upregulate NF-κB signaling, and modulation of the levels of NF-κB components reduces toxicity. In additional studies, we show that toxicity from untethered, linear chains is regulated by isoleucine 44, which anchors a key interaction site for ubiquitin. We conclude that free ubiquitin chains can be toxic, but only in uncommon circumstances, such as when the ability of cells to modify and regulate them is markedly restricted.


Assuntos
Imunidade Inata/genética , Isoleucina/metabolismo , NF-kappa B/metabolismo , Processamento de Proteína Pós-Traducional/genética , Ubiquitina/metabolismo , Ubiquitinação/genética , Animais , Drosophila melanogaster , Transdução de Sinais
18.
Antioxid Redox Signal ; 31(15): 1133-1149, 2019 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-31482721

RESUMO

Aims: Ubiquitin is a highly conserved protein modifier that heavily accumulates during the oxidative stress response. Here, we investigated the role of the ubiquitination system, particularly at the linkage level, in the degradation of oxidized proteins. The function of ubiquitin in the removal of oxidized proteins remains elusive because of the wide range of potential targets and different roles that polyubiquitin chains play. Therefore, we describe in detail the dynamics of the K48 ubiquitin response as the canonical signal for protein degradation. We identified ubiquitin targets and defined the relationship between protein ubiquitination and oxidation during the stress response. Results: Combining oxidized protein isolation, linkage-specific ubiquitination screens, and quantitative proteomics, we found that K48 ubiquitin accumulated at both the early and late phases of the stress response. We further showed that a fraction of oxidized proteins are conjugated with K48 ubiquitin. We identified ∼750 ubiquitinated proteins and ∼400 oxidized proteins that were modified during oxidative stress, and around half of which contain both modifications. These proteins were highly abundant and function in translation and energy metabolism. Innovation and Conclusion: Our work showed for the first time that K48 ubiquitin modifies a large fraction of oxidized proteins, demonstrating that oxidized proteins can be targeted by the ubiquitin/proteasome system. We suggest that oxidized proteins that rapidly accumulate during stress are subsequently ubiquitinated and degraded during the late phase of the response. This delay between oxidation and ubiquitination may be necessary for reprogramming protein dynamics, restoring proteostasis, and resuming cell growth.


Assuntos
Lisina/metabolismo , Poliubiquitina/metabolismo , Metabolismo Energético/fisiologia , Humanos , Lisina/química , Oxirredução , Poliubiquitina/química , Proteômica/métodos , Ubiquitina/metabolismo
19.
JMIR Med Inform ; 7(1): e9845, 2019 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-30907730

RESUMO

BACKGROUND: Concerns about privacy and personal data protection resulted in reforms of the existing legislation in the European Union (EU). The General Data Protection Regulation (GDPR) aims to reform the existing directive on the topic of personal data protection of EU citizens with a strong emphasis on more control of the citizens over their data and in the establishment of rules for the processing of personal data. OpenEHR is a standard that embodies many principles of interoperable and secure software for electronic health records (EHRs) and has been advocated as the best approach for the development of hospital information systems. OBJECTIVE: This study aimed to understand to what extent the openEHR standard can help in the compliance of EHR systems to the GDPR requirements. METHODS: A list of requirements for an EHR to support GDPR compliance and also a list of the openEHR design principles were made. The requirements were categorized and compared with the principles by experts on openEHR and GDPR. RESULTS: A total of 50 GDPR requirements and 8 openEHR design principles were identified. The openEHR principles conformed to 30% (15/50) of GDPR requirements. All the openEHR principles were aligned with GDPR requirements. CONCLUSIONS: This study showed that the openEHR principles conform well to GDPR, underlining the common wisdom that truly realizing security and privacy requires it to be built in from the start. By using an openEHR-based EHR, the institutions are closer to becoming compliant with GDPR while safeguarding the medical data.

20.
Stud Health Technol Inform ; 264: 773-777, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31438029

RESUMO

ObsCare is an obstetric-specific Electronic Health Record in use in nine Portuguese obstetric departments. Like other EHRs, it faces major challenges related to semantic interoperability and data quality. openEHR is proposed to address those needs. This study aimed to describe a summary representation of Obscare workflow and to validate whether archetypes in the openEHR Clinical Knowledge Manager repository can represent ObsCare clinical concepts. The study included the phases: a) ObsCare form selection; b) Description of the workflow care process; c) Detailed data extraction; and d) CKM models analysis. 379 variables were analyzed: 219 were fully represented in CKM repository; 99 were partially represented and needed archetype modification; and 61 were not represented and need new archetypes. To conclude, our study showed that the openEHR CKM repository requires further enhancements to be able to fully answer to the needs of an obstetric-specific EHR, the ObsCare software.


Assuntos
Registros Eletrônicos de Saúde , Software , Confiabilidade dos Dados , Atenção à Saúde , Feminino , Humanos , Trabalho de Parto , Gravidez
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