Colonization, diversity, and seasonality of fishes at pelagic fish aggregating devices.

The pelagic zone of the ocean can be a challenging environment in which to conduct research and as a result we lack the robust baseline abundance and diversity data, compared to what is available in more accessible coastal habitats, to be able to track changes or stressors to the biota in this environment. Many large-scale fisheries target pelagic fish, and much of the information available on these species is based on fisheries-dependent data that may be biased towards hotspots and commercially valuable fishes. Here, a long-term video and visual fish survey was conducted on two subsurface moored fish aggregating devices (FADs) in the pelagic waters of the central Bahamas to determine the feasibility of using moored pelagic FADs as tools for collecting fish abundance and diversity data. A wide range of species was documented, including large migratory fish that are the focus of commercial and recreational fisheries, and smaller often overlooked species on which little abundance or seasonality information exists. We found that FADs colonize quickly and reach a peak stable (albeit seasonally cyclical) abundance and diversity within the first several months after deployment. Species richness was higher in video surveys, but abundance was higher in visual surveys, except for sharks. Our results highlight the need to tailor survey methods to fit the context and study objective, and provide further evidence for the importance of fisheries-independent data in monitoring pelagic species.

The Proteome Profile of Olfactory Ecto-Mesenchymal Stem Cells-Derived from Patients with Familial Alzheimer's Disease Reveals New Insights for AD Study.

Alzheimer's disease (AD), the most common neurodegenerative disease and the first cause of dementia worldwide, has no effective treatment, and its pathological mechanisms are not yet fully understood. We conducted this study to explore the proteomic differences associated with Familial Alzheimer's Disease (FAD) in olfactory ecto-mesenchymal stem cells (MSCs) derived from PSEN1 (A431E) mutation carriers compared with healthy donors paired by age and gender through two label-free liquid chromatography-mass spectrometry approaches. The first analysis compared carrier 1 (patient with symptoms, P1) and its control (healthy donor, C1), and the second compared carrier 2 (patient with pre-symptoms, P2) with its respective control cells (C2) to evaluate whether the protein alterations presented in the symptomatic carrier were also present in the pre-symptom stages. Finally, we analyzed the differentially expressed proteins (DEPs) for biological and functional enrichment. These proteins showed impaired expression in a stage-dependent manner and are involved in energy metabolism, vesicle transport, actin cytoskeleton, cell proliferation, and proteostasis pathways, in line with previous AD reports. Our study is the first to conduct a proteomic analysis of MSCs from the Jalisco FAD patients in two stages of the disease (symptomatic and presymptomatic), showing these cells as a new and excellent in vitro model for future AD studies.

Patient-Derived Fibroblasts With Presenilin-1 Mutations, That Model Aspects of Alzheimer's Disease Pathology, Constitute a Potential Object for Early Diagnosis.

Alzheimer's disease (AD), a neurodegenerative disorder that can occur in middle or old age, is characterized by memory loss, a continuous decline in thinking, behavioral and social skills that affect the ability of an individual to function independently. It is divided into sporadic and familial subtypes. Early-onset familial AD (FAD) is linked to mutations in genes coding for the amyloid-ß protein precursor (AßPP), presenilin 1 (PS1), and presenilin 2 (PS2), which lead to alterations in AßPP processing, generation of the Amyloid-ß peptide and hyperphosphorylation of tau protein. Identification of early biomarkers for AD diagnosis represents a challenge, and it has been suggested that molecular changes in neurodegenerative pathways identified in the brain of AD patients can be detected in peripheral non-neural cells derived from familial or sporadic AD patients. In the present study, we determined the protein expression, the proteomic and in silico characterization of skin fibroblasts from FAD patients with PS1 mutations (M146L or A246E) or from healthy individuals. Our results shown that fibroblasts from AD patients had increased expression of the autophagy markers LC3II, LAMP2 and Cathepsin D, a significant increase in total GSK3, phosphorylated ERK1/2 (Thr202/Tyr204) and phosphorylated tau (Thr231, Ser396, and Ser404), but no difference in the phosphorylation of Akt (Ser473) or the α (Ser21) and ß (Ser9) GSK3 isoforms, highlighting the relevant role of abnormal protein post-translational modifications in age-related neurodegenerative diseases, such as AD. Both 2-DE gels and mass spectrometry showed significant differences in the expression of the signaling pathways associated with protein folding and the autophagic pathway mediated by chaperones with the expression of HSPA5, HSPE1, HSPD1, HSP90AA1, and HSPE1 and reticular stress in the FAD samples. Furthermore, expression of the heat shock proteins HSP90 and HSP70 was significantly higher in the cells from AD patients as confirmed by Western blot. Taken together our results indicate that fibroblasts from patients with FAD-PS1 present alterations in signaling pathways related to cellular stress, autophagy, lysosomes, and tau phosphorylation. Fibroblasts can therefore be useful in modeling pathways related to neurodegeneration, as well as for the identification of early AD biomarkers.

Identification and biochemical characterization of an ATP-dependent dihydroxyacetone kinase from Trypanosoma cruzi.

Dihydroxyacetone (DHA) can be used as an energy source by many cell types; however, it is toxic at high concentrations. The enzyme dihydroxyacetone kinase (DAK) has shown to be involved in DHA detoxification and osmoregulation. Among protozoa of the genus Trypanosoma, T. brucei, which causes sleeping sickness, is highly sensitive to DHA and does not have orthologous genes to DAK. Conversely, T. cruzi, the etiological agent of Chagas Disease, has two putative ATP-dependent DAK (TcDAKs) sequences in its genome. Here we show that T. cruzi epimastigote lysates present a DAK specific activity of 27.1 nmol/min/mg of protein and that this form of the parasite is able to grow in the presence of 2 mM DHA. TcDAK gene was cloned and the recombinant enzyme (recTcDAK) was expressed in Escherichia coli. An anti-recTcDAK serum reacted with a protein of the expected molecular mass of 61 kDa in epimastigotes. recTcDAK presented maximal activity using Mg+2, showing a Km of 6.5 µM for DHA and a K0.5 of 124.7 µM for ATP. As it was reported for other DAKs, recTcDAK activity was inhibited by FAD with an IC50 value of 0.33 mM. In conclusion, TcDAK is the first DAK described in trypanosomatids confirming another divergent metabolism between T. brucei and T. cruzi.

Quality of life in early-onset Alzheimer's disease due to a PSEN1-E280A mutation.

PURPOSE: The present study aims to explore the association between the quality of life (QoL) score and the clinical and sociodemographic variables in patients with the PSEN1-E280A mutation. We also seek to evaluate the differences between the QoL reported by the patients (P-QoL) and the scores reported by the caregivers (C-QoL). METHODS: An analysis of 75 patients with the PSEN1-E280A mutation with mild cognitive impairment and dementia was performed. We used the Quality of Life in Alzheimer Disease (QoL-AD) survey to evaluate QoL as an outcome and evaluated its association with sociodemographic, lifestyle, clinical, and past medical history variables. RESULTS: The largest difference in the median of the QoL-AD score was in those who needed help to eat, those with moderate or severe dementia, those classified as frail or pre-frail, those with moderate social risk, and those with depression. Also, C-QoL was lower than the P-QoL, and the QoL-AD of individuals with severe dementia was lower than for milder forms of the disease. Not needing help to eat, not having a stressful situation in the past 3 months, and the years of education were positively correlated with QoL-AD in the linear model. CONCLUSION: As studies in similar populations with AD, factors with more impact on QoL are those related to loss of functionality and independence. These factors are also associated with variables related to the current literature with the burden of the disease for the caregivers.

Are mental disorders related to disbelief in free will? A systematic review.

BACKGROUND: The nature and existence of free will have been debated for centuries. Since some psychiatric disorders are known to interfere with one's ability to control their actions and thoughts (e.g., schizophrenia), the investigation of the psychiatric facet of free will beliefs seems to be relevant. In this systematic review, we were interested in clarifying if and how having a mental disorder affects individuals' beliefs in free will by comparing psychiatric vs. non-psychiatric samples. METHODS: A systematic search of MEDLINE, Web of Science, EMBASE, and PsycINFO databases was performed between 04 and 09 November 2020. The search strategy included "free will" and related constructs and terms related to DSM-5 mental disorders characterized by psychotic, compulsive, avoidant, or impulsive symptoms. Eligible designs of studies included case-control and cohort studies. Study selection took place in committee meetings consisting of six researchers. Quality assessment of the selected studies was performed through the Joanna Briggs Institute Appraisal Checklist for Case Control Studies. RESULTS: After removing duplicates, a total of 12,218 titles/abstracts were screened. Inclusion and exclusion criteria were followed, and three articles were eventually selected. CONCLUSIONS: It is not possible to provide unequivocal confirmation that having a mental disorder can or cannot affect someone's belief in free will. Studies with different mental disorders should be conducted in this field. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42018109468.

Functional characterization of the putative FAD synthase from Mycoplasma hyopneumoniae.

In bacteria, the biosynthesis of the cofactor flavin adenine dinucleotide (FAD), important in many physiological responses, is catalyzed by the bifunctional enzyme FAD synthase (FADSyn) which converts riboflavin into FAD by both kinase and adenylylation activity. The in silico 3D structure of a putative FADSyn from Mycoplasma hyopneumoniae (MhpFADSyn), the etiological agent of enzootic pneumonia was already reported, nevertheless, the in vitro functional characterization was not yet demonstrated. Our phylogenetic analysis revealed that MhpFADSyn is close related to the bifunctional FADSyn from Corynebacterium ammoniagenes. However, only the domain related to adenylylation was assigned by InterPro database. The activity of MhpFADSyn was evaluated through in vitro enzymatic assays using cell extracts from IPTG-inducible heterologous expression of MhpFADSyn in Escherichia coli. The flavoproteins were analyzed by HPLC and results showed that IPTG-induced cell lysate resulted in the formation of twofold increased amounts of FAD if compared to non IPTG-induced cells. Consumption of riboflavin substrate was also threefold greater in IPTG-induced lysate compared to non IPTG-induced cell extract. Thus, the recombinant MhpFADSyn protein could be associated to FAD biosynthesis. These findings contribute to expand the range of potential drug targets in diseases control and unveil metabolic pathways that could be attribute to mycoplasmas.

Optical imaging of cleavage stage bovine embryos using hyperspectral and confocal approaches reveals metabolic differences between on-time and fast-developing embryos.

The assessment of embryo quality aims to enhance subsequent pregnancy and live birth outcomes. Metabolic analysis of embryos has immense potential in this regard. As a step towards this goal, here we assess the metabolism of bovine embryos using label-free optical imaging. We compared embryos defined as either on-time or fast-developing, as fast dividing embryos are more likely to develop to the blastocyst stage. Specifically, bovine embryos at 48 (Day 2) and 96 (Day 4) hours post fertilization were fixed and separated based on morphological assessment: on-time (Day 2: 2 cell; Day 4: 5-7 cell) or fast-developing (Day 2: 3-7 cell; Day 4: 8-16 cell). Embryos with different developmental rates on Day 2 and Day 4 were correlated with metabolic activity and DNA damage. Confocal microscopy was used to assess metabolic activity by quantification of cellular autofluorescence specific for the endogenous fluorophores NAD(P)H and FAD with a subsequent calculation of the optical redox ratio. Separately, hyperspectral microscopy was employed to assess a broader range of endogenous fluorophores. DNA damage was determined using γH2AX immunohistochemistry. Hyperspectral imaging showed significantly lower abundance of endogenous fluorophores in fast-developing compared to on-time embryos on Day 2, indicating a lower metabolic activity. On Day 4 of development there was no difference in the abundance of FAD between on-time and fast-developing embryos. There was, however, significantly higher levels of NAD(P)H in fast-developing embryos leading to a significantly lower optical redox ratio when compared to on-time embryos. Collectively, these results demonstrate that fast-developing embryos present a 'quiet' metabolic pattern on Day 2 and Day 4 of development, compared to on-time embryos. There was no difference in the level of DNA damage between on-time and fast-developing embryos on either day of development. To our knowledge, this is the first collective use of confocal and hyperspectral imaging in cleavage-stage bovine embryos in the absence of fluorescent tags.

Ciclos de atenção a dietas da moda e tendências de busca na internet pelo Google trends / Cycles of attention to fad diets and internet search trends by Google trends

Resumo Este trabalho objetivou descrever e analisar conteúdos de notícias associadas a ciclos de buscas ao Google ligados a 8 tipos de dieta: a cetogênica; da lua; da proteína; da sopa; detox; dos pontos; paleo; e Dukan. Estas foram selecionadas por terem sido apontadas pelo Google Trends® (GT) como as mais frequentemente associadas ao termo dieta. Os conteúdos dos sites vinculados aos maiores picos de buscas foram apontados pelo Google News®. O volume de buscas às dietas foram estimados pelo GT com filtros de região (Brasil) e temática (Saúde) considerando o período de 01/01/2012 a 01/01/2017. Os acessos retratam ciclos efêmeros com centenas de picos e quedas de interesse pelas 8 dietas da moda. A ênfase se concentra nas orientações para rápida perda de peso em prol de um ideal de saúde tido como proxy de um ideal estético. A dimensão dos riscos associados às dietas mais restritivas é citada frequentemente e validada por informações de especialistas. Destaca-se a exposição e a influência da opinião de celebridades relatando suas experiências dietéticas. Ao contrário do ideal de variedade e equilíbrio, as dietas da moda retratam a pressão pelo autocontrole alimentar como recurso à construção de um corpo idealizado e tipificado por imagens das celebridades.

Abstract This work aimed at identifying, describing and analyzing news content associated with Google search cycles linked to eight types of diet: ketogenic; moon; protein; soup; detox; points; paleo; and Dukan. They were selected because they were pointed out by Google Trends® (GT) as the most often associated with the term "diet". The content of sites linked to the highest search peaks was pointed out by Google News®. The volume of search attempts for diets was estimated by the GT with region filters (Brazil) and theme (Health) considering the period from 01/01/2012 to 01/01/2017. Results: hits portray ephemeral cycles, with hundreds of peaks and declining interest in the eight fad diets. The emphasis is on guidelines for quick weight loss in favor of optimal health as a proxy for an esthetic ideal. The scale of the risks associated with the more restrictive diets is often quoted and validated by expert information. The exposure and influence of the opinion of celebrities reporting their dietary experiences stand out. Contrary to the ideal of moderation, variety, and balance, fad diets portray the pressure for food self-control as a resource for building an idealized body typified by celebrity images.

Characterization and validation of olive FAD and SAD gene families: expression analysis in different tissues and during fruit development.

Stearoyl-ACP desaturases (SADs) and fatty acid desaturases (FADs) play a critical role in plant lipid metabolism and also affect oil fatty acid composition introducing double bonds into the hydrocarbon chains to produce unsaturated fatty acids. In the present study, the genomic sequences of three SAD and three FAD candidate genes were characterized in olive and their expression was evaluated in different plant tissues. OeSAD genes corresponded to olive SAD1 and SAD2 and to a newly identified OeSAD4, sharing the conserved protein structure with other plant species. On the other hand, the full-length genomic sequences of two microsomal OeFAD genes (FAD2-1 and FAD2-2) and the plastidial FAD6, were released. When the level of expression was tested on different tissues of cv. Leccino, OeSAD1 and OeSAD2 were mainly expressed in the fruits, while OeFAD genes showed the lowest expression in this tissue. The mRNA profiling of all genes was directly studied in fruits of Leccino and Coratina cultivars during fruit development. In both genotypes, the expression level of OeSAD1 and OeSAD2 had the highest value during and after the pit-hardening period, when oil accumulation in fruit mesocarp is intensively increasing. Furthermore, the expression level of both OeFAD2 genes, which were the main candidates for oleic acid desaturation, were almost negligible during fruit ripening. These results have made possible to define candidate genes of the machinery regulation of fatty acid composition in olive oil, providing information on their sequence, gene structure and chromosomal location.

Calculation of the Geometries and Infrared Spectra of the Stacked Cofactor Flavin Adenine Dinucleotide (FAD) as the Prerequisite for Studies of Light-Triggered Proton and Electron Transfer.

Flavin cofactors, like flavin adenine dinucleotide (FAD), are important electron shuttles in living systems. They catalyze a wide range of one- or two-electron redox reactions. Experimental investigations include UV-vis as well as infrared spectroscopy. FAD in aqueous solution exhibits a significantly shorter excited state lifetime than its analog, the flavin mononucleotide. This finding is explained by the presence of a "stacked" FAD conformation, in which isoalloxazine and adenine moieties form a π-complex. Stacking of the isoalloxazine and adenine rings should have an influence on the frequency of the vibrational modes. Density functional theory (DFT) studies of the closed form of FAD in microsolvation (explicit water) were used to reproduce the experimental infrared spectra, substantiating the prevalence of the stacked geometry of FAD in aqueous surroundings. It could be shown that the existence of the closed structure in FAD can be narrowed down to the presence of only a single water molecule between the third hydroxyl group (of the ribityl chain) and the N7 in the adenine ring of FAD.

Nutritional composition of fad diets published on websites and blogs / Composição nutricional de dietas da moda publicadas em sites e blogs

ABSTRACT Objective To analyze quantitatively the diets available on blogs and websites. Methods The following 15 diets were analyzed: low-carb diet, gluten-free diet and intermittent fasting diet using the Virtual Nutri software program. The nutrients analyzed were as follows: carbohydrates, fibers, proteins, lipids, iodine, sodium, calcium, magnesium, iron, zinc, manganese, phosphorus, selenium, vitamins A, C, B6, B12, D, niacin, and folate. The nutrient values were compared with the Dietary Reference Intake recommendation for adults between the ages of 19-50 years. The macronutrient adequacy percentage was based on a 2000-kcal diet for a healthy life, as recommended by the World Health Organization. Results It was observed that most of the composition of macro and micronutrients was lower than the one established by the World Health Organization and the Dietary Reference Intake recommendation for adults between the ages of 19-50 years. Conclusion Long-term dieting can be a risk factor for several deficiency diseases and may endanger the health of individuals.

RESUMO Objetivo Analisar quantitativamente dietas disponíveis em blogs e sites. Métodos Analisou-se 15 cardápios de dietas intituladas: Low Carb, Dieta Sem Glúten e Jejum Intermitente utilizando o software, Virtual Nutri. Os nutrientes analisados foram: carboidratos, fibras, proteínas, lipídios, iodo, sódio, cálcio, magnésio, ferro, zinco, manganês, potássio, fósforo, cobre, selênio, vitaminas A, C, B6, B12, D, niacina e folato. Os nutrientes tiveram seus valores comparados com recomendações diárias de ingestão das Dietary Reference Intake para adultos da faixa etária de 19 a 50 anos. O percentual de adequação foi feito baseado em uma dieta de 2000kcal para indivíduos saudáveis, de acordo com o preconizado pela Organização Mundial da Saúde. Resultados Observou-se que a maioria dos cardápios teve sua composição de macro e micronutrientes aquém do determinado pela Organização Mundial da Saúde e das recomendações das Dietary Reference Intake para adultos de 19 a 50 anos. Conclusão O seguimento dessas dietas em longo prazo pode ser um fator de risco a diversas doenças carenciais que pode trazer perigos à saúde dos indivíduos.

Expression analysis of genes involved in the synthesis of oleic and linoleic acids in Jatropha cinerea seeds from Northwestern Mexico / Análise de expressão de genes envolvidos na síntese de ácidos oleico e linoleico em sementes de Jatropha cinerea do noroeste do México

Plants belonging to genus Jatropha has arisen interest because of its high oil content that could be used to produce biodiesel. It is also widely reported that the main fatty acids in Jatropha oilseed are oleic and linoleic acids. However, there are scarce studies related to native species of Jatropha from Northwestern Mexico which are adapted to arid conditions, and the expression of genes involved in fatty acid synthesis for these species is still unknown. Therefore, the aim of this study was to analyze the expression of five genes, ACP1, KASII, D9SD, FAD2-1 and FAD2-2, which are involved in the oleic and linoleic acids synthesis in mature wild-seeds of Jatropha cinerea, a native species from Sonoran Desert, using semi-quantitative RT-PCR. The J. cinerea seeds were randomly collected in Bahía de Kino, Sonora (México) which is a region characterized by its harsh environments such as saline soils and extreme temperature changes and J. curcas mature seeds from a non-toxic variety from Veracruz, Mexico were used as a reference. The RT-PCR analysis of three biological replicates were considered to ensure data consistent. Our analysis showed a higher expression of KASII and FAD2-1 genes in J. cinerea seeds compared to J. curcas, meanwhile the expression of ACP1, D9SD and FAD2-2 were higher in J. curcas. Furthermore, Actin and FAD2-1 genes sequences here obtained are the first reported for J. cinerea, thus providing information to develop further studies.(AU)

Plantas pertencentes ao gênero Jatropha têm despertado interesse devido ao seu alto teor de óleo que poderia ser usado para produzir biodiesel. Também é amplamente relatado que os principais ácidos graxos das oleaginosas da Jatropha são os ácidos oléico e linoleico. No entanto, existem poucos estudos relacionados a espécies nativas de pinhão-manso do noroeste do México que estão adaptados a condições áridas, e a expressão de genes envolvidos na síntese de ácidos graxos para essas espécies ainda é desconhecida. Portanto, o objetivo deste estudo foi analisar a expressão de cinco genes, ACP1, KASII, D9SD, FAD2-1 e FAD2-2, que estão envolvidos na síntese de ácidos oleico e linoleico em sementes selvagens maduras de Jatropha cinerea, espécies nativas do Deserto de Sonora, usando RT-PCR semi-quantitativa. Sementes de J. cinerea foram coletadas aleatoriamente em Bahía de Kino, Sonora (México), região caracterizada por ambientes agressivos como solos salinos e mudanças extremas de temperatura, e sementes de J. curcas maduras de uma variedade não-tóxica de Veracruz, México, usado como referência. Análise RT-PCR de três repetições biológicas foram consideradas para garantir dados consistentes. Nossa análise mostrou uma maior expressão dos genes KASII e FAD2-1 em sementes de J. cinerea comparado a J. curcas, enquanto a expressão de ACP1, D9SD e FAD2-2 foi maior em J. curcas. Além disso, as sequências dos genes Actin e FAD2-1 obtidas são as primeiras relatadas para J. cinerea, fornecendo, assim, informações para o desenvolvimento de novos estudos.(AU)

Expression analysis of genes involved in the synthesis of oleic and linoleic acids in Jatropha cinerea seeds from Northwestern Mexico / Análise de expressão de genes envolvidos na síntese de ácidos oleico e linoleico em sementes de Jatropha cinerea do noroeste do México

ABSTRACT: Plants belonging to genus Jatropha has arisen interest because of its high oil content that could be used to produce biodiesel. It is also widely reported that the main fatty acids in Jatropha oilseed are oleic and linoleic acids. However, there are scarce studies related to native species of Jatropha from Northwestern Mexico which are adapted to arid conditions, and the expression of genes involved in fatty acid synthesis for these species is still unknown. Therefore, the aim of this study was to analyze the expression of five genes, ACP1, KASII, D9SD, FAD2-1 and FAD2-2, which are involved in the oleic and linoleic acids synthesis in mature wild-seeds of Jatropha cinerea, a native species from Sonoran Desert, using semi-quantitative RT-PCR. The J. cinerea seeds were randomly collected in Bahía de Kino, Sonora (México) which is a region characterized by its harsh environments such as saline soils and extreme temperature changes and J. curcas mature seeds from a non-toxic variety from Veracruz, Mexico were used as a reference. The RT-PCR analysis of three biological replicates were considered to ensure data consistent. Our analysis showed a higher expression of KASII and FAD2-1 genes in J. cinerea seeds compared to J. curcas, meanwhile the expression of ACP1, D9SD and FAD2-2 were higher in J. curcas. Furthermore, Actin and FAD2-1 genes sequences here obtained are the first reported for J. cinerea, thus providing information to develop further studies.

RESUMO: Plantas pertencentes ao gênero Jatropha têm despertado interesse devido ao seu alto teor de óleo que poderia ser usado para produzir biodiesel. Também é amplamente relatado que os principais ácidos graxos das oleaginosas da Jatropha são os ácidos oléico e linoleico. No entanto, existem poucos estudos relacionados a espécies nativas de pinhão-manso do noroeste do México que estão adaptados a condições áridas, e a expressão de genes envolvidos na síntese de ácidos graxos para essas espécies ainda é desconhecida. Portanto, o objetivo deste estudo foi analisar a expressão de cinco genes, ACP1, KASII, D9SD, FAD2-1 e FAD2-2, que estão envolvidos na síntese de ácidos oleico e linoleico em sementes selvagens maduras de Jatropha cinerea, espécies nativas do Deserto de Sonora, usando RT-PCR semi-quantitativa. Sementes de J. cinerea foram coletadas aleatoriamente em Bahía de Kino, Sonora (México), região caracterizada por ambientes agressivos como solos salinos e mudanças extremas de temperatura, e sementes de J. curcas maduras de uma variedade não-tóxica de Veracruz, México, usado como referência. Análise RT-PCR de três repetições biológicas foram consideradas para garantir dados consistentes. Nossa análise mostrou uma maior expressão dos genes KASII e FAD2-1 em sementes de J. cinerea comparado a J. curcas, enquanto a expressão de ACP1, D9SD e FAD2-2 foi maior em J. curcas. Além disso, as sequências dos genes Actin e FAD2-1 obtidas são as primeiras relatadas para J. cinerea, fornecendo, assim, informações para o desenvolvimento de novos estudos.

Overlapping riboflavin supply pathways in bacteria.

Riboflavin derivatives are essential cofactors for a myriad of flavoproteins. In bacteria, flavins importance extends beyond their role as intracellular protein cofactors, as secreted flavins are a key metabolite in a variety of physiological processes. Bacteria obtain riboflavin through the endogenous riboflavin biosynthetic pathway (RBP) or by the use of importer proteins. Bacteria frequently encode multiple paralogs of the RBP enzymes and as for other micronutrient supply pathways, biosynthesis and uptake functions largely coexist. It is proposed that bacteria shut down biosynthesis and would rather uptake riboflavin when the vitamin is environmentally available. Recently, the overlap of riboflavin provisioning elements has gained attention and the functions of duplicated paralogs of RBP enzymes started to be addressed. Results point towards the existence of a modular structure in the bacterial riboflavin supply pathways. Such structure uses subsets of RBP genes to supply riboflavin for specific functions. Given the importance of riboflavin in intra and extracellular bacterial physiology, this complex array of riboflavin provision pathways may have developed to contend with the various riboflavin requirements. In riboflavin-prototrophic bacteria, riboflavin transporters could represent a module for riboflavin provision for particular, yet unidentified processes, rather than substituting for the RBP as usually assumed.

The putative flavin carrier family FlcA-C is important for Aspergillus fumigatus virulence.

Aspergillus fumigatus is an opportunistic fungal pathogen and the most important species causing pulmonary fungal infections. The signaling by calcium is very important for A. fumigatus pathogenicity and it is regulated by the transcription factor CrzA. We have previously used used ChIP-seq (Chromatin Immunoprecipitation DNA sequencing) aiming to identify gene targets regulated by CrzA. We have identified among several genes regulated by calcium stress, the putative flavin transporter, flcA. This transporter belongs to a small protein family composed of FlcA, B, and C. The ΔflcA null mutant showed several phenotypes, such as morphological defects, increased sensitivity to calcium chelating-agent ethylene glycol tetraacetic acid (EGTA), cell wall or oxidative damaging agents and metals, repre-sentative of deficiencies in calcium signaling and iron homeostasis. Increasing calcium concentrations improved significantly the ΔflcA growth and conidiation, indicating that ΔflcA mutant has calcium insufficiency. Finally, ΔflcA-C mutants showed reduced flavin adenine dinucleotide (FAD) and were avirulent in a low dose murine infection model.

FAD binding properties of a cytosolic version of Escherichia coli NADH dehydrogenase-2.

Respiratory NADH dehydrogenase-2 (NDH-2) of Escherichia coli is a peripheral membrane-bound flavoprotein. By eliminating its C-terminal region, a water soluble truncated version was obtained in our laboratory. Overall conformation of the mutant version resembles the wild-type protein. Considering these data and the fact that the mutant was obtained as an apo-protein, the truncated version is an ideal model to study the interaction between the enzyme and its cofactor. Here, the FAD binding properties of this version were characterized using far-UV circular dichroism (CD), differential scanning calorimetry (DSC), limited proteolysis, and steady-state and dynamic fluorescence spectroscopy. CD spectra, thermal unfolding and DSC profiles did not reveal any major difference in secondary structure between apo- and holo-protein. In addition, digestion site accessibility and tertiary conformation were similar for both proteins, as seen by comparable chymotryptic cleavage patterns. FAD binding to the apo-protein produced a parallel increment of both FAD fluorescence quantum yield and steady-state emission anisotropy. On the other hand, addition of FAD quenched the intrinsic fluorescence emission of the truncated protein, indicating that the flavin cofactor should be closely located to the protein Trp residues. Analysis of the steady-state and dynamic fluorescence data confirms the formation of the holo-protein with a 1:1 binding stoichiometry and an association constant KA=7.0(±0.8)×10(4)M(-1). Taken together, the FAD-protein interaction is energetically favorable and the addition of FAD is not necessary to induce the enzyme folded state. For the first time, a detailed characterization of the flavin:protein interaction was performed among alternative NADH dehydrogenases.

The C-terminal extension of bacterial flavodoxin-reductases: involvement in the hydride transfer mechanism from the coenzyme.

To study the role of the mobile C-terminal extension present in bacterial class of plant type NADP(H):ferredoxin reductases during catalysis, we generated a series of mutants of the Rhodobacter capsulatus enzyme (RcFPR). Deletion of the six C-terminal amino acids beyond alanine 266 was combined with the replacement A266Y, emulating the structure present in plastidic versions of this flavoenzyme. Analysis of absorbance and fluorescence spectra suggests that deletion does not modify the general geometry of FAD itself, but increases exposure of the flavin to the solvent, prevents a productive geometry of FAD:NADP(H) complex and decreases the protein thermal stability. Although the replacement A266Y partially coats the isoalloxazine from solvent and slightly restores protein stability, this single change does not allow formation of active charge-transfer complexes commonly present in the wild-type FPR, probably due to restraints of C-terminus pliability. A proton exchange process is deduced from ITC measurements during coenzyme binding. All studied RcFPR variants display higher affinity for NADP(+) than wild-type, evidencing the contribution of the C-terminus in tempering a non-productive strong (rigid) interaction with the coenzyme. The decreased catalytic rate parameters confirm that the hydride transfer from NADPH to the flavin ring is considerably hampered in the mutants. Although the involvement of the C-terminal extension from bacterial FPRs in stabilizing overall folding and bent-FAD geometry has been stated, the most relevant contributions to catalysis are modulation of coenzyme entrance and affinity, promotion of the optimal geometry of an active complex and supply of a proton acceptor acting during coenzyme binding.

Alterations in brain structure and neurodevelopmental outcome in preterm infants hospitalized in different neonatal intensive care unit environments.

OBJECTIVE: To evaluate associations between neonatal intensive care unit (NICU) room type (open ward and private room) and medical outcomes; neurobehavior, electrophysiology, and brain structure at hospital discharge; and developmental outcomes at 2 years of age. STUDY DESIGN: In this prospective longitudinal cohort study, we enrolled 136 preterm infants born <30 weeks gestation from an urban, 75-bed level III NICU from 2007-2010. Upon admission, each participant was assigned to a bedspace in an open ward or private room within the same hospital, based on space and staffing availability, where they remained for the duration of hospitalization. The primary outcome was developmental performance at 2 years of age (n = 86 infants returned for testing, which was 83% of survivors) measured using the Bayley Scales of Infant and Toddler Development, 3rd Edition. Secondary outcomes were: (1) medical factors throughout the hospitalization; (2) neurobehavior; and (3) cerebral injury and maturation (determined by magnetic resonance imaging and electroencephalography). RESULTS: At term equivalent age, infants in private rooms were characterized by a diminution of normal hemispheric asymmetry and a trend toward having lower amplitude integrated electroencephalography cerebral maturation scores (P = .02; ß = -0.52 [CI -0.95, -0.10]). At age 2 years, infants from private rooms had lower language scores (P = .006; ß = -8.3 [CI -14.2, -2.4]) and a trend toward lower motor scores (P = .02; ß = -6.3 [CI -11.7, -0.99]), which persisted after adjustment for potential confounders. CONCLUSION: These findings raise concerns that highlight the need for further research into the potential adverse effects of different amounts of sensory exposure in the NICU environment.

Sodium azide mutagenesis resulted in a peanut plant with elevated oleate content

Screening of peanut seeds resulting from 0.39 percent sodium azide treatment with NIRS calibration equation for bulk seed samples identified a plant with more than 60 percent oleate. Oleate content in individual seeds of the plant, as predicted by NIRS calibration equation for intact single peanut seeds, ranged from 50.05 percent ~ 68.69 percent. Three seeds with >60 percent oleate thus identified were further confirmed by gas chromatography. Multiple sequence alignments of the FAD2B gene from Huayu 22 (wild type) and peanut seeds with elevated oleate (mutant type) revealed a C281T transition in the coding region causing an I94T substitution in the oleoyl-PC desaturase, which may be responsible for reduction in the enzyme activity.