Plasma membrane-specific interactome analysis reveals calpain 1 as a druggable modulator of rescued Phe508del-CFTR cell surface stability

Cystic fibrosis (CF) is a genetic disease caused by mutations in the gene encoding CF transmembrane conductance regulator (CFTR), a chloride channel normally expressed at the surface of epithelial cells. The most frequent mutation, resulting in Phe-508 deletion, causes CFTR misfolding and its premature degradation. Low temperature or pharmacological correctors can partly rescue the Phe508del-CFTR processing defect and enhance trafficking of this channel variant to the plasma membrane (PM). Nevertheless, the rescued channels have an increased endocytosis rate, being quickly removed from the PM by the peripheral protein quality-control pathway. We previously reported that rescued Phe508del-CFTR (rPhe508del) can be retained at the cell surface by stimulating signaling pathways that coax the adaptor molecule ezrin (EZR) to tether rPhe508del–Na+/H+-exchange regulatory factor-1 (NHERF1) complexes to the actin cytoskeleton, thereby averting the rapid internalization of this channel variant. However, the molecular basis for why rPhe508del fails to recruit active EZR to the PM remains elusive. Here, using a proteomics approach, we characterized and compared the core components of wt-CFTR– or rPhe508del–containing macromolecular complexes at the surface of human bronchial epithelial cells. We identified calpain 1 (CAPN1) as an exclusive rPhe508del interactor that prevents active EZR recruitment, impairs rPhe508del anchoring to actin, and reduces its stability in the PM. We show that either CAPN1 downregulation or its chemical inhibition dramatically improves the functional rescue of Phe508del-CFTR in airway cells. These observations suggest that CAPN1 constitutes an attractive target for pharmacological intervention, as part of CF combination therapies restoring Phe508del-CFTR function.This work was supported by a center grant UID/MULTI/04046/2019 to BioISI and project PTDC/BIA-CEL/28408/2017 and IF2012 to PM, both from FCT, Portugal. AMM was recipient of fellowship SFRH/BD/52490/2014 from BioSYS PhD programme PD65-2012, and PB of fellowship SFRH/BPD/94322/2013.N/

Response to treatment with interferon-alpha and ribavirin in patients with chronic Hepatitis C virus genotypes 2 and 3 depends on the degree of hepatic fibrosis

The combined therapy with interferon alfa plus ribavirin (INF+RBV) is considered the most appropriate treatment for patients with chronic hepatitis C virus genotypes 2 and 3 in Brazil. However, wide variations in the rates of sustained viral response (SVR) have been reported among such patients. We evaluated, retrospectively, factors associated with SVR in subjects with chronic hepatitis C virus genotypes 2 and 3 and that received medication from the Health Secretariat of the state of São Paulo. One-hundred-seventy-seven consecutive patients with chronic hepatitis C were treated for 24 or 48 weeks according to the viral genotype. Patients co-infected with associated hepatic diseases or who had problems with alcohol abuse were excluded. The genotype of the HCV-RNA was identified through restriction analysis, the viral load through quantitative PCR (Amplicor, Roche) and the degree of hepatic fibrosis according to the Metavir score. Demographic, virological and histological parameters were submitted to binary logistic regression analysis to identify the variables associated with SVR. The overall rate of SVR was 36.4% for the 177 patients, and genotype 2 or 3 was the main parameter independently associated with SVR. Among the 77 patients with these viral genotypes, only the stage of fibrosis had a significant effect on the SVR (odds ratio (OR) = 3.035; 95% CI (confidence interval) = 1.196-7.699; p=0.019). The rate of SVR among the subjects with fibrosis at an advanced stage (F3-F4) was 38%, compared to 75% for patients with fibrosis at an initial stage (F0-F2). Consequently, other therapeutic options should be considered for patients with genotypes 2 and 3 who have advanced fibrosis.Federal University of São PauloSírio-Libanês Hospital of São PauloUNIFESPSciEL

Oceanographic drivers of population differentiation in Indo-Pacific bottlenose (Tursiops aduncus) and humpback (Sousa spp.) dolphins of the northern Bay of Bengal

The Bay of Bengal is one of the most productive ecosystems in the northern Indian Ocean and it harbours a rich community of cetaceans, including Indo-Pacific bottlenose (Tursiops aduncus) and humpback (Sousa spp.) dolphins. The taxonomy of these genera has been controversial, but within the Indian Ocean both seem to be divided into phylogenetically discrete units that range from the east to the west. Within the Sousa genus, S. plumbea is distributed in the western Indian Ocean while S. chinensis is distributed in the eastern Indian and western Pacific Ocean. T. aduncus has a discontinuous distribution throughout the Indo-Pacific Ocean and two different phylogenetic units are known to exist, one along the eastern African coast and another one in the eastern Indian and west Pacific Ocean. In this study we investigate the phylogeography of Indo-Pacific humpback and bottlenose dolphins in the northern Bay of Bengal. We sequenced the mitochondrial DNA control region for 17 bottlenose and 15 humpback dolphins and compared the results with previously published sequences within each genus. In both cases, we found that Bangladesh dolphins are genetically different from neighbouring populations. While the Bangladesh T. aduncus seem to be more closely related to the African T. aduncus form than the Pacific form, Sousa spp. seem to be more closely related to individuals from Australia. The genetic uniqueness of these populations has important evolutionary implications, due to their isolation, coastal distribution in a geographic cul-de-sac characterized by an extreme infusion, redistribution and recycling of biological productivity, and conservation implications since their survival is threatened in particular by fatal interactions with fisheries. We suggest that the particular and extreme oceanographic conditions found in the Bay of Bengal may be driving speciation in these dolphins and other marine megafauna.info:eu-repo/semantics/publishedVersio

Monitoring biotechnological processes through quantitative image analysis: application to 2-phenylethanol production by Yarrowia lipolytica

Available online 20 March 2023Quantitative image analysis (QIA) is a simple and automated tool for process monitoring that, when combined with chemometric techniques, enables the association of changes in microbiota morphology to various operational parameters. To that effect, principal component analysis, multilinear regression, and ordinary least squares methods were applied to the obtained dataset of the biotransformation conditions for Y. lipolytica through the monitor of yeast morphology, substrates (glycerol, L-phenylalanine - L-Phe) consumption and metabolites (2-phenylethanol 2-PE) production was developed. Glycerol and L-Phe were successfully monitored by the proposed approach, though with a lower monitoring ability for 2-PE, and mostly related to yeast and cluster size and proportion, yeasts contents and cluster morphology. The chemometric approach also allowed to identify significant morphological modifications related with the change in the stirring speed in the experiments at 600rpm, 600/400rpm (600rpm for 24h, and 400rpm until the end of the experiment) and in pH from 5.5 to 7.5. This work demonstrated, for the first time, that QIA combined with chemometric analysis can be considered a valuable tool to monitor biotechnological processes, namely the 2-PE production by Y. lipolytica, by analyzing yeast and cluster morphology.info:eu-repo/semantics/publishedVersio

Influência das condições climáticas na fenologia do pessegueiro cv. 'Royal Time'

A fenologia estuda a morfologia dos diferentes estados do desenvolvimento das plantas e permite entender e datar o seu desenvolvimento, sendo particularmente importante nas espécies frutícolas, designadamente para o posicionamento das técnicas culturais a realizar ao longo do ciclo anual. Nas prunóideas de clima temperado, nas quais se inclui o pessegueiro, a cerejeira, a ameixeira e a amendoeira, o ciclo anual inicia-se com o desabrochamento dos gomos florais, que se vão desenvolvendo até atingir a plena floração. A chegada de pólen compatível ao estigma da flor pode conduzir à sua fertilização culminando no vingamento dos frutos. Este período de floração é condicionado pelo clima e é utilizado como indicador das alterações climáticas, sendo fundamental para entender como as plantas respondem a estas alterações, preocupação crescente por parte dos produtores devido ao aumento da frequência e intensidade de eventos climáticos extremos. Com o objetivo de estudar a influência das condições meteorológicas no período de floração em pessegueiros da Região da Beira Interior, foi observada a fenologia da cultivar Royal Time, num mesmo pomar, nos ciclos 2015, 2016 e 2018. Paralelamente, foi recolhida informação das variáveis climáticas temperatura e precipitação. Os resultados permitiram observar que, no ciclo 2015, o período de floração, desde o estado C ao estado G foi muito curto, de aproximadamente 17 dias, com a plena floração a ocorrer em 12 de março. Em 2016 o período de floração teve a duração de 36 dias, com a plena floração em 24 de março e em 2018 o período de floração estendeu-se por 35 dias, com a plena floração em 29 de março. Este estudo visa não só evidenciar a irregularidade climática que se observou nos últimos anos, como também contribuir para fornecer aos produtores informação quantificada sobre o efeito da irregularidade dos fatores climáticos na fenologiainfo:eu-repo/semantics/publishedVersio

Activation of the Maternal Immune System During Pregnancy Alters Behavioral Development of Rhesus Monkey Offspring

Background: Maternal infection during pregnancy is associated with an increased risk of schizophrenia and autism in the offspring. Supporting this correlation, experimentally activating the maternal immune system during pregnancy in rodents produces offspring with abnormal brain and behavioral development. We have developed a nonhuman primate model to bridge the gap between clinical populations and rodent models of maternal immune activation (MIA). Methods: A modified form of the viral mimic, synthetic double-stranded RNA (polyinosinic:polycytidylic acid stabilized with poly-L-lysine) was delivered to two separate groups of pregnant rhesus monkeys to induce MIA: 1) late first trimester MIA (n = 6), and 2) late second trimester MIA (n = 7). Control animals (n = 11) received saline injections at the same first or second trimester time points or were untreated. Sickness behavior, temperature, and cytokine profiles of the pregnant monkeys confirmed a strong inflammatory response to MIA. Results: Behavioral development of the offspring was studied for 24 months. Following weaning at 6 months of age, MIA offspring exhibited abnormal responses to separation from their mothers. As the animals matured, MIA offspring displayed increased repetitive behaviors and decreased affiliative vocalizations. When evaluated with unfamiliar conspecifics, first trimester MIA offspring deviated from species-typical macaque social behavior by inappropriately approaching and remaining in immediate proximity of an unfamiliar animal. Conclusions: In this rhesus monkey model, MIA yields offspring with abnormal repetitive behaviors, communication, and social interactions. These results extended the findings in rodent MIA models to more human-like behaviors resembling those in both autism and schizophrenia

A Modified e-Delphi Study

Therapeutic interventions for people with problematic use of psychoactive substances can help tackle specific needs related to substance addiction consequences. This modified e-Delphi study aimed to establish consensus on a training program for self-management of substance addiction consequences. The study was conducted between February and April 2022, with an experts’ sample of 28 participants in the first round and 24 in the second. A priori consensus criteria were defined for each round. The results revealed a very strong consensus was achieved on the structure of the program and on clinical areas, such as the problematic use of substances, general health knowledge, health-seeking behavior and adherence, self-knowledge and well-being, social role and personal dignity, and family process. Additionally, over 80% participant consensus was achieved on an extensive number of interventions categorized as psychoeducational, psychotherapeutic, socio therapeutic, brief interventions, social skills training, problem solving techniques, relaxation techniques, and counseling. These findings may be able to fulfill a gap concerning structured treatment approaches for people with problematic use of psychoactive substances. Supporting self-management of the consequences of substance addiction and its application can change nurses’ interventions.info:eu-repo/semantics/publishedVersio

Extraction and characterization of extracellular polymeric substances from aerobic granular sludge from a full-scale sequencing batch reactor in Portugal

Microbiotec'17 - Congress of Microbiology and Biotechnology 2017Background: Aerobic granular sludge (AGS) is a recently developed technology for wastewater treatment. This system is able to manage higher amounts of wastewater and requires less surface area than conventional systems. The granules consist of microorganisms embedded in a self-produced extracellular polymeric substances (EPS) matrix. EPS are high molecular weight polymers, which can be metabolic products of microorganisms (e.g. proteins, polysaccharides, humic substances, nucleic acids) or be due to cell lyses. Accumulation on the cells surface of such EPS forms a protective barrier for the cells from the external environment. This work focus on the extraction and quantification of EPS from AGS from a large scale bioreactor in Portugal, during approximately 4 months. Given the environmental and chemical differences that these granules are subjected to one of the goals was to assess variability in the EPS production and characterize the granules morphology and composition in a large scale environment.The authors thank the company SIMTEJO for supplying the granules, and the financial support of European Social Fund, under Programa Operacional under the project NORTE-08-5369-FSE-000007 and BioTecNorte operation (NORTE-010145-FEDER-000004). This work was also supported by Portuguese Funds from FCT - Fundação para a Ciência e a Tecnologia through the strategic funding of UID/Multi/50016/2013 and UID/BIO/04469/2013 units and COMPETE 2020 (POCI-01-0145-FEDER-006684). C.L. Amorim wishes to acknowledge the research grant from FCT (SFRH/BPD/96481/2013).info:eu-repo/semantics/publishedVersio

Pinus Susceptibility to Pitch Canker Triggers Specific Physiological Responses in Symptomatic Plants:An Integrated Approach

Fusarium circinatum, the causal agent of pine pitch canker (PPC), is an emergent and still understudied risk that threatens Pinus forests worldwide, with potential production and sustainability losses. In order to explore the response of pine species with distinct levels of susceptibility to PPC, we investigated changes in physiology, hormones, specific gene transcripts, and primary metabolism occurring in symptomatic Pinus pinea, Pinus pinaster, and Pinus radiata upon inoculation with F. circinatum. Pinus radiata and P. pinaster exhibiting high and intermediate susceptibility to PPC, respectively, suffered changes in plant water status and photosynthetic impairment. This was associated with sink metabolism induction, a general accumulation of amino acids and overexpression of pathogenesis-related genes. On the other hand, P. pinea exhibited the greatest resistance to PPC and stomatal opening, transpiration increase, and glycerol accumulation were observed in inoculated plants. A stronger induction of pyruvate decarboxylase transcripts and differential hormones regulation were also found for inoculated P. pinea in comparison with the susceptible Pinus species studied. The specific physiological changes reported herein are the first steps to understand the complex Pinus–Fusarium interaction and create tools for the selection of resistant genotypes thus contributing to disease mitigation