Improving obesity research: Unveiling metabolic pathways through a 3D In vitro model of adipocytes using 3T3-L1 cells.

Obesity, a burgeoning global health crisis, has tripled in prevalence over the past 45 years, necessitating innovative research methodologies. Adipocytes, which are responsible for energy storage, play a central role in obesity. However, most studies in this field rely on animal models or adipocyte monolayer cell cultures, which are limited in their ability to fully mimic the complex physiology of a living organism, or pose challenges in terms of cost, time consumption, and ethical considerations. These limitations prompt a shift towards alternative methodologies. In response, here we show a 3D in vitro model utilizing the 3T3-L1 cell line, aimed at faithfully replicating the metabolic intricacies of adipocytes in vivo. Using a workable cell line (3T3-L1), we produced adipocyte spheroids and differentiated them in presence and absence of TNF-α. Through a meticulous proteomic analysis, we compared the molecular profile of our adipose spheroids with that of adipose tissue from lean and obese C57BL/6J mice. This comparison demonstrated the model's efficacy in studying metabolic conditions, with TNF-α treated spheroids displaying a notable resemblance to obese white adipose tissue. Our findings underscore the model's simplicity, reproducibility, and cost-effectiveness, positioning it as a robust tool for authentically mimicking in vitro metabolic features of real adipose tissue. Notably, our model encapsulates key aspects of obesity, including insulin resistance and an obesity profile. This innovative approach has the potential to significantly impact the discovery of novel therapeutic interventions for metabolic syndrome and obesity. By providing a nuanced understanding of metabolic conditions, our 3D model stands as a transformative contribution to in vitro research, offering a pathway for the development of small molecules and biologics targeting these pervasive health issues in humans.

Production and Chemical Characterization of Silage from Saccharine Sorghum Fertirrigated with Sugarcane Vinasse.

BACKGROUND: Distillery vinasse is one of the promising bio-fertilizers, as it contains significant amounts of essential chemical elements, allied with sorghum that is widely used in the diet of ruminant animals and has been considered as an alternative to the production of other cereals or forages. This study aimed to evaluate saccharin sorghum silage from fertilization with vinasse. METHODS: The research was conducted using the BRS-511, CR-1339, and CR-1342 geno-types. The silage was held for 170 days after sowing, with experimental design in blocks with triple factorial (genotypes x fertilization x inoculation) totaling 54 installments. At 95 days, the silos were opened for sample collection and analysis bromatological analysis. RESULTS: The results indicate the primary source of variation was genotype, characterizing them with different potentials in productivity and better results for BRS-511, CR-1339, and CR-1342. The bromatological results indicate good quality for CR-1339 and CR-1342 hybrids, however, better digestability for BRS-511. There was no observable difference between the factors of fertilization. The inoculation additive assists in the reduction of lignin appears to be high. PCA analysis showed differences between cultivars (BRS-511, CR-1339, and CR-1342) and fertili-zation. However, the PCAs showed the genotypes show similar results with conventional ferti-lization and sugarcane vinasse. CONCLUSION: The study reflected the possibility of producing sweet sorghum silage with soil sugarcane vinasse fertilization as fertilizer.

DNA methylation of the promoter region at the CREB1 binding site is a mechanism for the epigenetic regulation of brain-specific PKMζ.

Protein kinase M zeta, PKMζ, is a brain enriched kinase with a well characterized role in Long-Term Potentiation (LTP), the activity-dependent strengthening of synapses involved in long-term memory formation. However, little is known about the molecular mechanisms that maintain the tissue specificity of this kinase. Here, we characterized the epigenetic factors, mainly DNA methylation, regulating PKMζ expression in the human brain. The PRKCZ gene has an upstream promoter regulating Protein kinase C ζ (PKCζ), and an internal promoter driving PKMζ expression. A demethylated region, including a canonical CREB binding site, situated at the internal promoter was only observed in human CNS tissues. The induction of site-specific hypermethylation of this region resulted in decreased CREB1 binding and downregulation of PKMζ expression. Noteworthy, CREB binding sites were absent in the upstream promoter of PRKCZ locus, suggesting a specific mechanism for regulating PKMζ expression. These observations were validated using a system of human neuronal differentiation from induced pluripotent stem cells (iPSCs). CREB1 binding at the internal promoter was detected only in differentiated neurons, where PKMζ is expressed. The same epigenetic mechanism in the context of CREB binding site was identified in other genes involved in neuronal differentiation and LTP. Additionally, aberrant DNA hypermethylation at the internal promoter was observed in cases of Alzheimer's disease, correlating with decreased expression of PKMζ in patient brains. Altogether, we present a conserved epigenetic mechanism regulating PKMζ expression and other genes enhanced in the CNS with possible implications in neuronal differentiation and Alzheimer's disease.

Discovery and structural characterization of chicoric acid as a SARS-CoV-2 nucleocapsid protein ligand and RNA binding disruptor.

The nucleocapsid (N) protein plays critical roles in coronavirus genome transcription and packaging, representing a key target for the development of novel antivirals, and for which structural information on ligand binding is scarce. We used a novel fluorescence polarization assay to identify small molecules that disrupt the binding of the N protein to a target RNA derived from the SARS-CoV-2 genome packaging signal. Several phenolic compounds, including L-chicoric acid (CA), were identified as high-affinity N-protein ligands. The binding of CA to the N protein was confirmed by isothermal titration calorimetry, 1H-STD and 15N-HSQC NMR, and by the crystal structure of CA bound to the N protein C-terminal domain (CTD), further revealing a new modulatory site in the SARS-CoV-2 N protein. Moreover, CA reduced SARS-CoV-2 replication in cell cultures. These data thus open venues for the development of new antivirals targeting the N protein, an essential and yet underexplored coronavirus target.

Extracellular matrix stiffness regulates degradation of MST2 via SCF ßTrCP.

The Hippo pathway plays central roles in relaying mechanical signals during development and tumorigenesis, but how the proteostasis of the Hippo kinase MST2 is regulated remains unknown. Here, we found that chemical inhibition of proteasomal proteolysis resulted in increased levels of MST2 in human breast epithelial cells. MST2 binds SCFßTrCP E3 ubiquitin ligase and silencing ßTrCP resulted in MST2 accumulation. Site-directed mutagenesis combined with computational molecular dynamics studies revealed that ßTrCP binds MST2 via a non-canonical degradation motif. Additionally, stiffer extracellular matrix, as well as hyperactivation of integrins resulted in enhanced MST2 degradation mediated by integrin-linked kinase (ILK) and actomyosin stress fibers. Our study uncovers the underlying biochemical mechanisms controlling MST2 degradation and underscores how alterations in the microenvironment rigidity regulate the proteostasis of a central Hippo pathway component.

Structural dynamics of SARS-CoV-2 nucleocapsid protein induced by RNA binding.

The nucleocapsid (N) protein of the SARS-CoV-2 virus, the causal agent of COVID-19, is a multifunction phosphoprotein that plays critical roles in the virus life cycle, including transcription and packaging of the viral RNA. To play such diverse roles, the N protein has two globular RNA-binding modules, the N- (NTD) and C-terminal (CTD) domains, which are connected by an intrinsically disordered region. Despite the wealth of structural data available for the isolated NTD and CTD, how these domains are arranged in the full-length protein and how the oligomerization of N influences its RNA-binding activity remains largely unclear. Herein, using experimental data from electron microscopy and biochemical/biophysical techniques combined with molecular modeling and molecular dynamics simulations, we show that, in the absence of RNA, the N protein formed structurally dynamic dimers, with the NTD and CTD arranged in extended conformations. However, in the presence of RNA, the N protein assumed a more compact conformation where the NTD and CTD are packed together. We also provided an octameric model for the full-length N bound to RNA that is consistent with electron microscopy images of the N protein in the presence of RNA. Together, our results shed new light on the dynamics and higher-order oligomeric structure of this versatile protein.

pyKVFinder: an efficient and integrable Python package for biomolecular cavity detection and characterization in data science.

BACKGROUND: Biomolecular interactions that modulate biological processes occur mainly in cavities throughout the surface of biomolecular structures. In the data science era, structural biology has benefited from the increasing availability of biostructural data due to advances in structural determination and computational methods. In this scenario, data-intensive cavity analysis demands efficient scripting routines built on easily manipulated data structures. To fulfill this need, we developed pyKVFinder, a Python package to detect and characterize cavities in biomolecular structures for data science and automated pipelines. RESULTS: pyKVFinder efficiently detects cavities in biomolecular structures and computes their volume, area, depth and hydropathy, storing these cavity properties in NumPy arrays. Benefited from Python ecosystem interoperability and data structures, pyKVFinder can be integrated with third-party scientific packages and libraries for mathematical calculations, machine learning and 3D visualization in automated workflows. As proof of pyKVFinder's capabilities, we successfully identified and compared ADRP substrate-binding site of SARS-CoV-2 and a set of homologous proteins with pyKVFinder, showing its integrability with data science packages such as matplotlib, NGL Viewer, SciPy and Jupyter notebook. CONCLUSIONS: We introduce an efficient, highly versatile and easily integrable software for detecting and characterizing biomolecular cavities in data science applications and automated protocols. pyKVFinder facilitates biostructural data analysis with scripting routines in the Python ecosystem and can be building blocks for data science and drug design applications.

Vaccination coverage against human papillomavirus (HPV) and associated factors in female academics from a university in southwestern Goiás, Brazil.

OBJECTIVE: To check the coverage of the HPV vaccine in women enrolled in health courses at a university in southwest Goiás, Brazil, and the factors associated with vaccination. METHODS: This is a cross-sectional study, including female university students of health courses, aged 18 years or more. A standardized and self-applying questionnaire was used. Participants who received two or more doses of the vaccine were considered immunized. Multiple analysis was performed using multinomial logistic regression. RESULT: We observed that, of the 1510 participants, 473 (31.3%) had two or more doses of HPV vaccine, 167 (11.0%) one dose and 870 (57.6%) were unvaccinated. Participants under 21 years of age and in socioeconomic stratum A were 2 times more likely to have received two or more doses of the vaccine (Prevalence Ratio = 1.95; 95%CI 1.40-2.70 and Prevalence Ratio = 2.09; 95%CI 1.39-3.13, respectively). CONCLUSIONS: The research revealed extensive possibility for interventions with the aim of achieving greater vaccination coverage among female university students. Even women with more knowledge and high economic stratum showed low vaccination coverage, suggesting that results of higher vaccine coverage can be obtained with vaccination carried out in a school environment.

Molecular and cellular basis of hyperassembly and protein aggregation driven by a rare pathogenic mutation in DDX3X.

Current studies estimate that 1-3% of females with unexplained intellectual disability (ID) present de novo splice site, nonsense, frameshift, or missense mutations in the DDX3X protein (DEAD-Box Helicase 3 X-Linked). However, the cellular and molecular mechanisms by which DDX3X mutations impair brain development are not fully comprehended. Here, we show that the ID-linked missense mutation L556S renders DDX3X prone to aggregation. By using a combination of biophysical assays and imaging approaches, we demonstrate that this mutant assembles solid-like condensates and amyloid-like fibrils. Although we observed greatly reduced expression of the mutant allele in a patient who exhibits skewed X inactivation, this appears to be enough to sequestrate healthy proteins into solid-like ectopic granules, compromising cell function. Therefore, our data suggest ID-linked DDX3X L556S mutation as a disorder arising from protein misfolding and aggregation.

Cryo-EM structure of the mature and infective Mayaro virus at 4.4 Å resolution reveals features of arthritogenic alphaviruses.

Mayaro virus (MAYV) is an emerging arbovirus of the Americas that may cause a debilitating arthritogenic disease. The biology of MAYV is not fully understood and largely inferred from related arthritogenic alphaviruses. Here, we present the structure of MAYV at 4.4 Å resolution, obtained from a preparation of mature, infective virions. MAYV presents typical alphavirus features and organization. Interactions between viral proteins that lead to particle formation are described together with a hydrophobic pocket formed between E1 and E2 spike proteins and conformational epitopes specific of MAYV. We also describe MAYV glycosylation residues in E1 and E2 that may affect MXRA8 host receptor binding, and a molecular "handshake" between MAYV spikes formed by N262 glycosylation in adjacent E2 proteins. The structure of MAYV is suggestive of structural and functional complexity among alphaviruses, which may be targeted for specificity or antiviral activity.

Vaccination coverage against human papillomavirus (HPV) and associated factors in female academics from a university in southwestern Goiás, Brazil / Cobertura vacinal contra o papiloma vírus humano (HPV) e fatores associados em acadêmicas de uma universidade do sudoeste goiano

ABSTRACT OBJECTIVE To check the coverage of the HPV vaccine in women enrolled in health courses at a university in southwest Goiás, Brazil, and the factors associated with vaccination. METHODS This is a cross-sectional study, including female university students of health courses, aged 18 years or more. A standardized and self-applying questionnaire was used. Participants who received two or more doses of the vaccine were considered immunized. Multiple analysis was performed using multinomial logistic regression. RESULT We observed that, of the 1510 participants, 473 (31.3%) had two or more doses of HPV vaccine, 167 (11.0%) one dose and 870 (57.6%) were unvaccinated. Participants under 21 years of age and in socioeconomic stratum A were 2 times more likely to have received two or more doses of the vaccine (Prevalence Ratio = 1.95; 95%CI 1.40-2.70 and Prevalence Ratio = 2.09; 95%CI 1.39-3.13, respectively). CONCLUSIONS The research revealed extensive possibility for interventions with the aim of achieving greater vaccination coverage among female university students. Even women with more knowledge and high economic stratum showed low vaccination coverage, suggesting that results of higher vaccine coverage can be obtained with vaccination carried out in a school environment.

RESUMO OBJETIVO Verificar a cobertura da vacina contra o HPV em mulheres matriculadas em cursos da área de saúde de uma universidade do sudoeste do Estado de Goiás e os fatores associados à vacinação. MÉTODOS Trata-se de estudo transversal, incluindo universitárias dos cursos da área de saúde, com 18 anos ou mais. Foi utilizado questionário padronizado e autoaplicável. As participantes que receberam duas ou mais doses da vacina foram consideradas como imunizadas. A análise múltipla foi realizada por meio de regressão logística multinomial. RESULTADOS Observou-se que, das 1510 participantes, 473 (31,3%) com duas ou mais doses de vacina contra o HPV, 167 (11,0%) com uma dose e 870 (57,6%) não vacinadas. As participantes com menos de 21 anos e inseridas no estrato socioeconômico A tinham 2 vezes mais chance de terem recebido duas doses ou mais da vacina (Razão de Prevalência = 1,95; IC95% 1,40-2,70 e Razão de Prevalência = 2,09; IC95% 1.39-3,13, respectivamente). CONCLUSÕES A pesquisa revelou extensa possibilidade para intervenções com o objetivo de atingir maior cobertura vacinal entre as universitárias. Mesmo mulheres com mais conhecimento e de estrato econômico elevado apresentaram baixa cobertura vacinal, sugerindo que resultados de cobertura vacinal maior podem ser obtidos com a vacinação realizada em ambiente escolar.

Soil organic matter fractions and carbon management index under integrated crop-livestock system / Frações da matéria orgânica do solo e índice de manejo de carbono em sistema de integração lavoura-pecuária

The objective of this work was to evaluate the carbon content of the physical, chemical and oxidizable fractions of soil organic matter (SOM) and to calculate the carbon management index (CMI) in an area managed under an integrated crop-livestock system (ICLS) in the western region of Paraná - Brazil. The experiment was carried out at the experimental farm, belonging to the Universidade Estadual do Oeste do Paraná. Seventeen areas, which are managed in different ways, fifteen in ICLS and two areas of controls (Forest and Haymaking), using the design divided with two nested controls, with three replications were evaluated. Deformed and undisturbed soil samples were collected from all the areas to determine the total organic carbon (TOC), carbon stock, the physical, chemical and oxidizable fractions of SOM and the CMI in the layers of 0-0.05, 0.05-0.1 and 0.1-0.2 m. Little significant changes in the fractions were found for the management of the ICLS area in relation to the Forest and the area of Haymaking, although the Forest presented the best values for most of the studied fractions. It is recommended to adopt sustainable practices, such as ICLS, even though the average fractions tend to take time to match reference areas.

O objetivo desse trabalho foi avaliar os teores de carbono das frações física, química e oxidável da matéria orgânica do solo (MOS) e calcular o índice de manejo de carbono (IMC) em uma área manejada em sistema de integração lavoura pecuária (ILP) na região Oeste do Paraná - Brasil. O experimento foi realizado na fazenda experimental, pertencente à Universidade Estadual do Oeste do Paraná. Foram avaliadas dezessete áreas, que foram manejadas de diferentes formas, quinze em ILP e duas testemunhas (Mata e Fenação), sendo empregado o delineamento subsubsubdividido com duas testemunhas aninhadas, com três repetições. Em todas as áreas foram coletadas amostras deformadas e indeformadas de solo para serem determinados o carbono orgânico total, estoque de carbono e as frações físicas granulométricas, oxidáveis e químicas da MOS e o IMC nas camadas de 0-0,05, 0,05-0,1 e 0,1-0,2 m. Alterações pouco significativas das frações foram encontradas para os manejos da área em ILP em relação a mata e a área de fenação, entretanto a mata apresentou os melhores valores para a maioria das frações estudadas. Recomenda-se a adoção de práticas sustentáveis, como a ILP, mesmo que os teores médios das frações tendem a demorar tempo para igualar-se a áreas de referência.

Predictors of Post-Discharge 30-Day Hospital Readmission in Decompensated Heart Failure Patients

Abstract Background Heart failure (HF) is worldwide known as a public health issue with high morbimortality. One of the issues related to the evolution of HF is the high rate of hospital readmission caused by decompensation of the clinical condition, with high costs and worsening of ventricular function. Objective To quantify the readmission rate and identify the predictors of rehospitalization in patients with acute decompensated heart failure. Methods Hospital-based historic cohort of patients admitted with acute decompensated HF in a private hospital from Recife/PE, from January 2008 to February 2016, followed-up for at least 30 days after discharge. Demographic and clinical data of admission, hospitalization, and clinical and late outcomes were analyzed. Logistic regression was used as a strategy to identify the predictors of independent risks. Results 312 followed-up patients, average age 73 (± 14), 61% males, 51% NYHA Class III, and 58% ischemic etiology. Thirty-day readmission rate was 23%. Multivariate analysis identified the independent predictors ejection fraction < 40% (OR = 2.1; p = 0.009), hyponatremia (OR = 2.9; p = 0.022) and acute coronary syndrome (ACS) as the cause of decompensation (OR = 1.1; p = 0,026). The final model using those three variables presented reasonable discriminatory power (C-Statistics = 0.655 - HF 95%: 0.582 - 0.728) and good calibration (Hosmer-Lemeshow p = 0.925). Conclusions Among hospitalized patients with acute decompensated heart failure, the rate of readmission was high. Hyponatremia, reduced ejection fraction and ACS as causes of decompensation were robust markers for the prediction of hospital readmission within 30 days of discharge. (Int J Cardiovasc Sci. 2020; 33(2):175-184)

Decompensated Heart Failure with Mid-Range Ejection Fraction: Epidemiology and In-Hospital Mortality Risk Factors

Abstract Background: Recently, a new HF entity, with LVEF between 40-49%, was presented to comprehend and seek better therapy for HF with preserved LVEF (HFpEF) and borderline, in the means that HF with reduced LVEF (HFrEF) already has well-defined therapy in the literature. Objective: To compare the clinical-therapeutic profile of patients with HF with mid-range LVEF (HFmrEF) with HFpEF and HFrEF and to verify predictors of hospital mortality. Method: Historical cohort of patients admitted with decompensated HF at a supplementary hospital in Recife/PE between April/2007 - August/2017, stratified by LVEF (< 40%/40 - 49/≥ 50%), based on the guideline of the European Society of Cardiology (ESC) 2016. The groups were compared and Logistic Regression was used to identify predictors of independent risk for in-hospital death. Results: A sample of 493 patients, most with HFrEF (43%), HFpEF (30%) and HFmrEF (26%). Average age of 73 (± 14) years, 59% men. Hospital mortality 14%, readmission within 30 days 19%. In therapeutics, it presented statistical significance among the 3 groups, spironolactone, in HFrEF patients. Hospital death and readmission within 30 days did not make difference. In the HFmrEF group, factors independently associated with death were: valve disease (OR: 4.17, CI: 1.01-9.13), altered urea at admission (OR: 6.18, CI: 1.78-11.45) and beta-blocker hospitalization (OR: 0.29, CI: 0.08-0.97). In HFrEF, predictors were: prior renal disease (OR: 2.84, CI: 1.19-6.79), beta-blocker at admission (OR: 0.29, CI: 0.12-0.72) and ACEI/ ARB (OR: 0.21, CI: 0.09-0.49). In HFpEF, only valve disease (OR: 4.61, CI: 1.33-15.96) and kidney disease (OR: 5.18, CI: 1.68-11.98) were relevant. Conclusion: In general, HFmrEF presented intermediate characteristics between HFrEF and HFpEF. Independent predictors of mortality may support risk stratification and management of this group.

Genes Controlled by DNA Methylation Are Involved in Wilms Tumor Progression.

To identify underlying mechanisms involved with metastasis formation in Wilms tumors (WTs), we performed comprehensive DNA methylation and gene expression analyses of matched normal kidney (NK), WT blastemal component, and metastatic tissues (MT) from patients treated under SIOP 2001 protocol. A linear Bayesian framework model identified 497 differentially methylated positions (DMPs) between groups that discriminated NK from WT, but MT samples were divided in two groups. Accordingly, methylation variance grouped NK and three MT samples tightly together and all WT with four MT samples that showed high variability. WT were hypomethylated compared to NK, and MT had a hypermethylated pattern compared to both groups. The methylation patterns were in agreement with methylases and demethylases expression. Methylation data pointed to the existence of two groups of metastases. While hierarchical clustering analysis based on the expression of all 2569 differentially expressed genes (DEGs) discriminated WT and MT from all NK samples, the hierarchical clustering based on the expression of 44 genes with a differentially methylated region (DMR) located in their promoter region revealed two groups: one containing all NKs and three MTs and one containing all WT and four MTs. Methylation changes might be controlling expression of genes associated with WT progression. The 44 genes are candidates to be further explored as a signature for metastasis formation in WT.

Physical Activity Level and Muscular Power of Physical Education Students / Nível de Atividade Física e Potência Muscular de Estudantes de Educação Física

Being physically active is an indispensable condition for health and quality of life. In addition, literature has presented promising studies on muscle power associations with other parameters of health-related physical fitness. However, there is still little evidence to assess the level of physical activity and muscle power. Thus, the objective of this study was to evaluate the level of physical activity and muscular power of physical education students of a course in Rio de Janeiro. In order to reach the proposed goal, 37 physical education students of both sexes, with a mean age of 26.13 years, participated in responding the International Physical Activity Questionnaire (IPAQ) and performing the horizontal impulse jump test for muscular power. As a result, it was observed that 86% of men were classified as very active and active. For women, 63,63% were identified as very active and active. For power, mean values of 2.02 ± 0.16 for men and 1.48 ± 0.27 for women were obtained. Therefore, it was concluded that both males and females are physically active, although males represent a greater quantitative muscle power, which is largely inferior to previously published studies. (AU).

Ser ativo fisicamente constitui condição indispensável para a saúde a qualidade de vida. Além disso, a literatura tem apresentado estudos promissores sobre as associações da potência muscular com outros parâmetros da aptidão física relacionada à saúde. No entanto, ainda são poucas as evidências que buscaram avaliar nível de atividade física e potência muscular. Dessa forma, o objetivo desse trabalho foi avaliar o nível de atividade física e a potência muscular de estudantes de educação física de um curso no Rio de Janeiro. Para atingir ao objetivo proposto 37 estudantes de educação física, de ambos os sexos, com média de idade de 26,13 anos participaram ao responder ao Questionário Internacional de Atividade Física (IPAQ) e executar o teste de salto de impulsão horizontal para potência muscular. Como resultado, foi observado que 86% dos homens foram classificados como muito ativos e ativos. Já para mulheres 63,63% foram identificadas como muito ativas e ativas. Para potência obteve-se os valores de média de 2,02 ± 0,16 para homens e 1,48 ± 0,27 para mulheres. Portanto, concluiu-se que tanto o sexo masculino quanto o feminino são ativos fisicamente, embora os homens representem um maior quantitativo a potência muscular mostrou-se inferior, em grande parte, aos estudos anteriormente publicados. (AU).

Mechanistic insights revealed by a UBE2A mutation linked to intellectual disability.

Ubiquitin-conjugating enzymes (E2) enable protein ubiquitination by conjugating ubiquitin to their catalytic cysteine for subsequent transfer to a target lysine side chain. Deprotonation of the incoming lysine enables its nucleophilicity, but determinants of lysine activation remain poorly understood. We report a novel pathogenic mutation in the E2 UBE2A, identified in two brothers with mild intellectual disability. The pathogenic Q93E mutation yields UBE2A with impaired aminolysis activity but no loss of the ability to be conjugated with ubiquitin. Importantly, the low intrinsic reactivity of UBE2A Q93E was not overcome by a cognate ubiquitin E3 ligase, RAD18, with the UBE2A target PCNA. However, UBE2A Q93E was reactive at high pH or with a low-pKa amine as the nucleophile, thus providing the first evidence of reversion of a defective UBE2A mutation. We propose that Q93E substitution perturbs the UBE2A catalytic microenvironment essential for lysine deprotonation during ubiquitin transfer, thus generating an enzyme that is disabled but not dead.

Remaining straw and release of nutrients from oat managed in integrated crop-livestock / Palhada remanescente e liberação de nutrientes da aveia preta manejada em sistema de integração lavoura-pecuária

This study aimed to evaluate the decomposition and nutrient release from residues of the culture of oats and fallow in crop-livestock integration system. The experimental design was a randomized complete block design with two replications, as parcels being formed by four managements (fallow, oats without grazing, oats grazed once and twice), and the subplots for evaluation periods along the soybean crop in succession (0, 10, 20, 30, 50, 100 and 120 days after sowing). The residual amounts of dry matter and the contents of Carbon (C), Nitrogen (N), Phosphorus (P) and Potassium (K) were determined. Oats without grazing and fallow with natural reappearance of turnip + ryegrass were the treatments that presented the highest amount of dry matter remaining, reaching 5,219 and 6,781 kg ha-1, respectively. The amount of nutrients, N, P and K released from the residues, were similar independent from the management adopted, with exponential reduction proportional to the reduction of the remaining dry matter. K was the nutrient released faster from the residues and should be considered at the time of fertilization calculation of the culture to be implanted. The integrated crop-livestock system in which takes place one and two grazing oats, even reducing soil cover, enables high nutrient cycling.

O objetivo do trabalho foi avaliar a decomposição e liberação de nutrientes dos resíduos da cultura da aveia preta e do pousio conduzidos em sistema de integração lavoura-pecuária. O delineamento experimental utilizado foi o de blocos casualizados, em esquema de parcelas subdivididas, com duas repetições, sendo as parcelas constituídas por quatro manejos (pousio, aveia sem pastejo, aveia pastejada uma e duas vezes) e as subparcelas, pelas épocas de avaliação ao longo do cultivo da soja em sucessão (0, 10, 20, 30, 50, 100 e 120 dias após a semeadura). Foram determinadas as quantidades residuais de matéria seca e os teores de Carbono (C), Nitrogênio (N), Fósforo (P) e Potássio (K). A aveia sem pastejo e o pousio com ressemeadura natural de aveia + azevém, foram os manejos que apresentaram as maiores quantidades de matéria seca remanescentes, chegando a 5.219 e 6.781 kg ha-1, respectivamente. A quantidade dos nutrientes, N, P e K liberados dos resíduos, foram semelhantes independente do manejo adotado, com redução exponencial e proporcional à redução da matéria seca remanescente. O K foi o nutriente liberado mais rapidamente dos resíduos e deve ser considerado no momento do cálculo de adubação da cultura a ser implantada. O sistema de integração lavoura-pecuária no qual se realiza um e dois pastejos da aveia, mesmo reduzindo a cobertura do solo, possibilita elevada ciclagem de nutrientes.

Glutaminase Affects the Transcriptional Activity of Peroxisome Proliferator-Activated Receptor γ (PPARγ) via Direct Interaction.

Phosphate-activated glutaminases catalyze the deamidation of glutamine to glutamate and play key roles in several physiological and pathological processes. In humans, GLS encodes two multidomain splicing isoforms: KGA and GAC. In both isoforms, the canonical glutaminase domain is flanked by an N-terminal region that is folded into an EF-hand-like four-helix bundle. However, the splicing event replaces a well-structured three-repeat ankyrin domain in KGA with a shorter, unordered C-terminal stretch in GAC. The multidomain architecture, which contains putative protein-protein binding motifs, has led to speculation that glutaminases are involved in cellular processes other than glutamine metabolism; in fact, some proteins have been identified as binding partners of KGA and the isoforms of its paralogue gene, GLS2. Here, a yeast two-hybrid assay identified nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) as a new binding partner of the glutaminase. We show that KGA and GAC directly bind PPARγ with a low-micromolar dissociation constant; the interaction involves the N-terminal and catalytic domains of glutaminases as well as the ligand-binding domain of the nuclear receptor. The interaction occurs within the nucleus, and by sequestering PPARγ from its responsive element DR1, the glutaminases decreased nuclear receptor activity as assessed by a luciferase reporter assay. Altogether, our findings reveal an unexpected glutaminase-binding partner and, for the first time, directly link mitochondrial glutaminases to an unanticipated role in gene regulation.