Critical importance of DNA binding for CSL protein functions in fission yeast.

CSL proteins [named after the homologs CBF1 (RBP-Jκ in mice), Suppressor of Hairless and LAG-1] are conserved transcription factors found in animals and fungi. In the fission yeast Schizosaccharomyces pombe, they regulate various cellular processes, including cell cycle progression, lipid metabolism and cell adhesion. CSL proteins bind to DNA through their N-terminal Rel-like domain and central ß-trefoil domain. Here, we investigated the importance of DNA binding for CSL protein functions in fission yeast. We created CSL protein mutants with disrupted DNA binding and found that the vast majority of CSL protein functions depend on intact DNA binding. Specifically, DNA binding is crucial for the regulation of cell adhesion, lipid metabolism, cell cycle progression, long non-coding RNA expression and genome integrity maintenance. Interestingly, perturbed lipid metabolism leads to chromatin structure changes, potentially linking lipid metabolism to the diverse phenotypes associated with CSL protein functions. Our study highlights the critical role of DNA binding for CSL protein functions in fission yeast.

Genetic effects on molecular network states explain complex traits.

The complexity of many cellular and organismal traits results from the integration of genetic and environmental factors via molecular networks. Network structure and effect propagation are best understood at the level of functional modules, but so far, no concept has been established to include the global network state. Here, we show when and how genetic perturbations lead to molecular changes that are confined to small parts of a network versus when they lead to modulation of network states. Integrating multi-omics profiling of genetically heterogeneous budding and fission yeast strains with an array of cellular traits identified a central state transition of the yeast molecular network that is related to PKA and TOR (PT) signaling. Genetic variants affecting this PT state globally shifted the molecular network along a single-dimensional axis, thereby modulating processes including energy and amino acid metabolism, transcription, translation, cell cycle control, and cellular stress response. We propose that genetic effects can propagate through large parts of molecular networks because of the functional requirement to centrally coordinate the activity of fundamental cellular processes.

R-loops and regulatory changes in chronologically ageing fission yeast cells drive non-random patterns of genome rearrangements.

Aberrant repair of DNA double-strand breaks can recombine distant chromosomal breakpoints. Chromosomal rearrangements compromise genome function and are a hallmark of ageing. Rearrangements are challenging to detect in non-dividing cell populations, because they reflect individually rare, heterogeneous events. The genomic distribution of de novo rearrangements in non-dividing cells, and their dynamics during ageing, remain therefore poorly characterized. Studies of genomic instability during ageing have focussed on mitochondrial DNA, small genetic variants, or proliferating cells. To characterize genome rearrangements during cellular ageing in non-dividing cells, we interrogated a single diagnostic measure, DNA breakpoint junctions, using Schizosaccharomyces pombe as a model system. Aberrant DNA junctions that accumulated with age were associated with microhomology sequences and R-loops. Global hotspots for age-associated breakpoint formation were evident near telomeric genes and linked to remote breakpoints elsewhere in the genome, including the mitochondrial chromosome. Formation of breakpoint junctions at global hotspots was inhibited by the Sir2 histone deacetylase and might be triggered by an age-dependent de-repression of chromatin silencing. An unexpected mechanism of genomic instability may cause more local hotspots: age-associated reduction in an RNA-binding protein triggering R-loops at target loci. This result suggests that biological processes other than transcription or replication can drive genome rearrangements. Notably, we detected similar signatures of genome rearrangements that accumulated in old brain cells of humans. These findings provide insights into the unique patterns and possible mechanisms of genome rearrangements in non-dividing cells, which can be promoted by ageing-related changes in gene-regulatory proteins.

A Comprehensive Strategy for Stepwise Design of a Lab PROTOTYPE for the Removal of Emerging Contaminants in Water Using Cyclodextrin Polymers as Adsorbent Material.

The significant environmental issue of water pollution caused by emerging contaminants underscores the imperative for developing novel cleanup methods that are efficient, economically viable, and that are intended to operate at high capacity and under continuous flows at the industrial scale. This study shows the results of the operational design to build a prototype for the retention at lab scale of pollutant residues in water by using as adsorbent material, insoluble polymers prepared by ß-cyclodextrin and epichlorohydrin as a cross-linking agent. Laboratory in-batch tests were run to find out the adsorbent performances against furosemide and hydrochlorothiazide as pollutant models. The initial evaluation concerning the dosage of adsorbent, pH levels, agitation, and concentration of pharmaceutical pollutants enabled us to identify the optimal conditions for conducting the subsequent experiments. The adsorption kinetic and the mechanisms involved were evaluated revealing that the experimental data perfectly fit the pseudo second-order model, with the adsorption process being mainly governed by chemisorption. With KF constant values of 0.044 (L/g) and 0.029 (L/g) for furosemide and hydrochlorothiazide, respectively, and the determination coefficient (R2) being higher than 0.9 for both compounds, Freundlich yielded the most favorable outcomes, suggesting that the adsorption process occurs on heterogeneous surfaces involving both chemisorption and physisorption processes. The maximum monolayer adsorption capacity (qmax) obtained by the Langmuir isotherm revealed a saturation of the ß-CDs-EPI polymer surface 1.45 times higher for furosemide (qmax = 1.282 mg/g) than hydrochlorothiazide (qmax = 0.844 mg/g). Based on these results, the sizing design and building of a lab-scale model were carried out, which in turn will be used later to evaluate its performance working in continuous flow in a real scenario.

Longevity is determined by ETS transcription factors in multiple tissues and diverse species.

Ageing populations pose one of the main public health crises of our time. Reprogramming gene expression by altering the activities of sequence-specific transcription factors (TFs) can ameliorate deleterious effects of age. Here we explore how a circuit of TFs coordinates pro-longevity transcriptional outcomes, which reveals a multi-tissue and multi-species role for an entire protein family: the E-twenty-six (ETS) TFs. In Drosophila, reduced insulin/IGF signalling (IIS) extends lifespan by coordinating activation of Aop, an ETS transcriptional repressor, and Foxo, a Forkhead transcriptional activator. Aop and Foxo bind the same genomic loci, and we show that, individually, they effect similar transcriptional programmes in vivo. In combination, Aop can both moderate or synergise with Foxo, dependent on promoter context. Moreover, Foxo and Aop oppose the gene-regulatory activity of Pnt, an ETS transcriptional activator. Directly knocking down Pnt recapitulates aspects of the Aop/Foxo transcriptional programme and is sufficient to extend lifespan. The lifespan-limiting role of Pnt appears to be balanced by a requirement for metabolic regulation in young flies, in which the Aop-Pnt-Foxo circuit determines expression of metabolic genes, and Pnt regulates lipolysis and responses to nutrient stress. Molecular functions are often conserved amongst ETS TFs, prompting us to examine whether other Drosophila ETS-coding genes may also affect ageing. We show that five out of eight Drosophila ETS TFs play a role in fly ageing, acting from a range of organs and cells including the intestine, adipose and neurons. We expand the repertoire of lifespan-limiting ETS TFs in C. elegans, confirming their conserved function in ageing and revealing that the roles of ETS TFs in physiology and lifespan are conserved throughout the family, both within and between species.

Removal of an Azo Dye from Wastewater through the Use of Two Technologies: Magnetic Cyclodextrin Polymers and Pulsed Light.

Water pollution by dyes is a huge environmental problem; there is a necessity to produce new decolorization methods that are effective, cost-attractive, and acceptable in industrial use. Magnetic cyclodextrin polymers offer the advantage of easy separation from the dye solution. In this work, the ß-CD-EPI-magnetic (ß-cyclodextrin-epichlorohydrin) polymer was synthesized, characterized, and tested for removal of the azo dye Direct Red 83:1 from water, and the fraction of non-adsorbed dye was degraded by an advanced oxidation process. The polymer was characterized in terms of the particle size distribution and surface morphology (FE-SEM), elemental analysis (EA), differential scanning calorimetry (DSC), thermal gravimetric analysis (TGA), infrared spectrophotometry (IR), and X-ray powder diffraction (XRD). The reported results hint that 0.5 g and pH 5.0 were the best conditions to carry out both kinetic and isotherm models. A 30 min contact time was needed to reach equilibrium with a qmax of 32.0 mg/g. The results indicated that the pseudo-second-order and intraparticle diffusion models were involved in the assembly of Direct Red 83:1 onto the magnetic adsorbent. Regarding the isotherms discussed, the Freundlich model correctly reproduced the experimental data so that adsorption was confirmed to take place onto heterogeneous surfaces. The calculation of the thermodynamic parameters further demonstrates the spontaneous character of the adsorption phenomena (ΔG° = −27,556.9 J/mol) and endothermic phenomena (ΔH° = 8757.1 J/mol) at 25 °C. Furthermore, a good reusability of the polymer was evidenced after six cycles of regeneration, with a negligible decline in the adsorption extent (10%) regarding its initial capacity. Finally, the residual dye in solution after treatment with magnetic adsorbents was degraded by using an advanced oxidation process (AOP) with pulsed light and hydrogen peroxide (343 mg/L); >90% of the dye was degraded after receiving a fluence of 118 J/cm2; the discoloration followed a pseudo first-order kinetics where the degradation rate was 0.0196 cm2/J. The newly synthesized ß-CD-EPI-magnetic polymer exhibited good adsorption properties and separability from water which, when complemented with a pulsed light-AOP, may offer a good alternative to remove dyes such as Direct Red 83:1 from water. It allows for the reuse of both the polymer and the dye in the dyeing process.

Pyruvate kinase variant of fission yeast tunes carbon metabolism, cell regulation, growth and stress resistance.

Cells balance glycolysis with respiration to support their metabolic needs in different environmental or physiological contexts. With abundant glucose, many cells prefer to grow by aerobic glycolysis or fermentation. Using 161 natural isolates of fission yeast, we investigated the genetic basis and phenotypic effects of the fermentation-respiration balance. The laboratory and a few other strains depended more on respiration. This trait was associated with a single nucleotide polymorphism in a conserved region of Pyk1, the sole pyruvate kinase in fission yeast. This variant reduced Pyk1 activity and glycolytic flux. Replacing the "low-activity" pyk1 allele in the laboratory strain with the "high-activity" allele was sufficient to increase fermentation and decrease respiration. This metabolic rebalancing triggered systems-level adjustments in the transcriptome and proteome and in cellular traits, including increased growth and chronological lifespan but decreased resistance to oxidative stress. Thus, low Pyk1 activity does not lead to a growth advantage but to stress tolerance. The genetic tuning of glycolytic flux may reflect an adaptive trade-off in a species lacking pyruvate kinase isoforms.

General amino acid control in fission yeast is regulated by a nonconserved transcription factor, with functions analogous to Gcn4/Atf4.

Eukaryotes respond to amino acid starvation by enhancing the translation of mRNAs encoding b-ZIP family transcription factors (GCN4 in Saccharomyces cerevisiae and ATF4 in mammals), which launch transcriptional programs to counter this stress. This pathway involves phosphorylation of the eIF2 translation factor by Gcn2-protein kinases and is regulated by upstream ORFs (uORFs) in the GCN4/ATF4 5' leaders. Here, we present evidence that the transcription factors that mediate this response are not evolutionarily conserved. Although cells of the fission yeast Schizosaccharomyces pombe respond transcriptionally to amino acid starvation, they lack clear Gcn4 and Atf4 orthologs. We used ribosome profiling to identify mediators of this response in S. pombe, looking for transcription factors that behave like GCN4 We discovered a transcription factor (Fil1) translationally induced by amino acid starvation in a 5' leader and Gcn2-dependent manner. Like Gcn4, Fil1 is required for the transcriptional response to amino acid starvation, and Gcn4 and Fil1 regulate similar genes. Despite their similarities in regulation, function, and targets, Fil1 and Gcn4 belong to different transcription factor families (GATA and b-ZIP, respectively). Thus, the same functions are performed by nonorthologous proteins under similar regulation. These results highlight the plasticity of transcriptional networks, which maintain conserved principles with nonconserved regulators.

Fitness Landscape of the Fission Yeast Genome.

The relationship between DNA sequence, biochemical function, and molecular evolution is relatively well-described for protein-coding regions of genomes, but far less clear in noncoding regions, particularly, in eukaryote genomes. In part, this is because we lack a complete description of the essential noncoding elements in a eukaryote genome. To contribute to this challenge, we used saturating transposon mutagenesis to interrogate the Schizosaccharomyces pombe genome. We generated 31 million transposon insertions, a theoretical coverage of 2.4 insertions per genomic site. We applied a five-state hidden Markov model (HMM) to distinguish insertion-depleted regions from insertion biases. Both raw insertion-density and HMM-defined fitness estimates showed significant quantitative relationships to gene knockout fitness, genetic diversity, divergence, and expected functional regions based on transcription and gene annotations. Through several analyses, we conclude that transposon insertions produced fitness effects in 66-90% of the genome, including substantial portions of the noncoding regions. Based on the HMM, we estimate that 10% of the insertion depleted sites in the genome showed no signal of conservation between species and were weakly transcribed, demonstrating limitations of comparative genomics and transcriptomics to detect functional units. In this species, 3'- and 5'-untranslated regions were the most prominent insertion-depleted regions that were not represented in measures of constraint from comparative genomics. We conclude that the combination of transposon mutagenesis, evolutionary, and biochemical data can provide new insights into the relationship between genome function and molecular evolution.

Long noncoding RNA repertoire and targeting by nuclear exosome, cytoplasmic exonuclease, and RNAi in fission yeast.

Long noncoding RNAs (lncRNAs), which are longer than 200 nucleotides but often unstable, contribute a substantial and diverse portion to pervasive noncoding transcriptomes. Most lncRNAs are poorly annotated and understood, although several play important roles in gene regulation and diseases. Here we systematically uncover and analyze lncRNAs in Schizosaccharomyces pombe. Based on RNA-seq data from twelve RNA-processing mutants and nine physiological conditions, we identify 5775 novel lncRNAs, nearly 4× the previously annotated lncRNAs. The expression of most lncRNAs becomes strongly induced under the genetic and physiological perturbations, most notably during late meiosis. Most lncRNAs are cryptic and suppressed by three RNA-processing pathways: the nuclear exosome, cytoplasmic exonuclease, and RNAi. Double-mutant analyses reveal substantial coordination and redundancy among these pathways. We classify lncRNAs by their dominant pathway into cryptic unstable transcripts (CUTs), Xrn1-sensitive unstable transcripts (XUTs), and Dicer-sensitive unstable transcripts (DUTs). XUTs and DUTs are enriched for antisense lncRNAs, while CUTs are often bidirectional and actively translated. The cytoplasmic exonuclease, along with RNAi, dampens the expression of thousands of lncRNAs and mRNAs that become induced during meiosis. Antisense lncRNA expression mostly negatively correlates with sense mRNA expression in the physiological, but not the genetic conditions. Intergenic and bidirectional lncRNAs emerge from nucleosome-depleted regions, upstream of positioned nucleosomes. Our results highlight both similarities and differences to lncRNA regulation in budding yeast. This broad survey of the lncRNA repertoire and characteristics in S. pombe, and the interwoven regulatory pathways that target lncRNAs, provides a rich framework for their further functional analyses.

Thymol Encapsulated into HP-ß-Cyclodextrin as an Alternative to Synthetic Fungicides to Induce Lemon Resistance against Sour Rot Decay.

Consumers demand the use of eco-friendly fungicides to treat fruit and vegetables and governmental authorities have unauthorized the application of chemical antifungals for the efficient control of sour rot. In the present research, the microwave irradiation (MW) method was used to encapsulate thymol into 2-hydroxylpropyl-beta-cyclodextrin (HP-ß-CD) and the effect of these HP-ß-CD on controlling sour rot in citrus fruit, caused by Geotrichum citri-aurantii, was evaluated. Amounts of 25 and 50 mM of HP-ß-CD-thymol were used, and compared with propiconazole, to control the decay of inoculated lemon fruit. The treatments were performed in curative and preventive experiments. The incidence and severity of Geotrichum citri-aurantii in 25 and 50 mM HP-ß-CD-thymol-treated fruit were reduced in both experiments. The preventive 50 mM HP-ß-CD-thymol treatment showed the best effect, reducing the sour rot, respiration rate and fruit weight loss during storage at 20 °C. HP-ß-CD-thymol increased polyphenol concentration and the activity of antioxidant enzymes, such as catalase (CAT), ascorbate peroxidase (APX) and peroxidase (POD) in lemon peel, and the highest effects were found with the 50-mM dose. In conclusion, the results show that the use of thymol encapsulated by MW into HP-ß-CD could be an effective and sustainable tool, a substitute to the synthetic fungicides, for G. citri-auriantii control in citrus fruit.

Comprehensive Characterization of Linalool-HP-ß-Cyclodextrin Inclusion Complexes.

The objective of the present study is to obtain linalool- cyclodextrin (CDs) solid complexes for possible applications in the food industry. For this purpose, a detailed study of linalool complexation was carried out at different pH values, to optimize the type of CDs and reaction medium that support the highest quantity of encapsulated linalool. Once demonstrated the ability of hydroxypropyl-ß-cyclodextrin (HP-ß-CDs), to form inclusion complexes with linalool (KC = 921 ± 21 L mol-1) and given their greater complexation efficacy (6.788) at neutral pH, HP-ß-CDs were selected to produce solid inclusion complexes by using two different energy sources, ultrasounds and microwave irradiation, subsequently spraying the solutions obtained in the Spray Dryer. To provide scientific solidity to the experimental results, the complexes obtained were characterized by using different instrumental techniques in order to confirm the inclusion of linalool in the HP-ß-CDs hydrophobic cavity. The linalool solid complexes obtained were characterized by using 1H nuclear magnetic resonance (1H-NMR) and 2D nuclear magnetic resonance (ROSEY), differential scanning calorimetry, thermogravimetry and Fourier transform infrared spectrometry. Moreover, the structure of the complex obtained were also characterized by molecular modeling.

Modelling the Inactivation and Possible Regrowth of Salmonella enterica Treated with Chlorophyllin-Chitosan Complex and Visible Light.

The study focuses on predictive modelling of inactivation of Salmonella enterica after treatment with chlorophyllin-chitosan complex and visible light. Salmonella cells were incubated with chlorophyllin-chitosan complex (0.001% chlorophyllin and 0.1% chitosan) for different times (5-60 min) and then illuminated with visible light (λ=405 nm, H e=38 J/cm2). Inactivation curves and post-treatment regrowth curves were built based on microbiological viability tests and data were fitted to ten inactivation and two regrowth models. The photoactivated complex reduced Salmonella population, which were unable to regrow. Weibull and Baranyi models were the best to describe the inactivation and regrowth kinetics respectively. In conclusion, data from the kinetic analysis and predictive modelling confirmed that photoactivated chlorophyllin-chitosan complex is a promising non-thermal approach for inactivation of Gram-negative pathogens, since no bacterial regrowth after treatment has been predicted.

Thorough characterization and stability of HP-ß-cyclodextrin thymol inclusion complexes prepared by microwave technology: A required approach to a successful application in food industry.

BACKGROUND: The aims of the present study were to obtain a stable dry powder formulation of cyclodextrins (CDs) encapsulating thymol, for successful use as an ingredient on an industrial scale, and to characterize the thymol-CDs complexes using different techniques. RESULTS: Thymol was successfully solubilized in aqueous solutions and the Kc value increased with the pH of the media until the pH was neutral, giving the highest values (2583 ± 176 L mol-1 ) for HP-ß-cyclodextrins (HP-ß-CDs). The best encapsulation efficiency of thymol in solid complexes was obtained using the microwave (MWI) encapsulation method. The different characterization techniques have demonstrated the affinity of HP-ß-CDs for thymol molecules, forming stable complexes. CONCLUSIONS: The results support the use of the MWI method in the preparation of solid HP-ß-CD-thymol complexes, due to greater encapsulation efficiency and technological and economic advantages for industrial applications. © 2018 Society of Chemical Industry.

Natural genetic variation impacts expression levels of coding, non-coding, and antisense transcripts in fission yeast.

Our current understanding of how natural genetic variation affects gene expression beyond well-annotated coding genes is still limited. The use of deep sequencing technologies for the study of expression quantitative trait loci (eQTLs) has the potential to close this gap. Here, we generated the first recombinant strain library for fission yeast and conducted an RNA-seq-based QTL study of the coding, non-coding, and antisense transcriptomes. We show that the frequency of distal effects (trans-eQTLs) greatly exceeds the number of local effects (cis-eQTLs) and that non-coding RNAs are as likely to be affected by eQTLs as protein-coding RNAs. We identified a genetic variation of swc5 that modifies the levels of 871 RNAs, with effects on both sense and antisense transcription, and show that this effect most likely goes through a compromised deposition of the histone variant H2A.Z. The strains, methods, and datasets generated here provide a rich resource for future studies.

The fission yeast rDNA-binding protein Reb1 regulates G1 phase under nutritional stress.

Yeast Reb1 and its mammalian ortholog TTF1 are conserved Myb-type DNA-binding proteins that bind to specific sites near the 3'-end of rRNA genes (rDNA). Here, they participate in the termination of transcription driven by RNA polymerase I and block DNA replication forks approaching in the opposite direction. We found that Schizosaccharomyces pombe Reb1 also upregulates transcription of the ste9(+) gene that is required for nitrogen-starvation-induced growth arrest with a G1 DNA content and sexual differentiation. Ste9 activates the anaphase-promoting complex or cyclosome ('APC/C') in G1, targeting B-cyclin for proteasomal degradation in response to nutritional stress. Reb1 binds in vivo and in vitro to a specific DNA sequence at the promoter of ste9(+), similar to the sequence recognized in the rDNA, and this binding is required for ste9(+) transcriptional activation and G1 arrest. This suggests that Reb1 acts as a link between rDNA metabolism and cell cycle control in response to nutritional stress. In agreement with this new role for Reb1 in the regulation of the G1-S transition, reb1Δ and wee1(ts) mutations are synthetically lethal owing to the inability of these cells to lengthen G1 before entering S phase. Similarly, reb1Δ cdc10(ts) cells are unable to arrest in G1 and die at the semi-permissive temperature.

Evaluation of the In Vitro Antimicrobial Efficacy against Staphylococcus aureus and epidermidis of a Novel 3D-Printed Degradable Drug Delivery System Based on Polycaprolactone/Chitosan/Vancomycin-Preclinical Study.

Acute and chronic bone infections, especially those caused by methicillin-resistant Staphylococcus aureus (MRSA), remains a major complication and therapeutic challenge. It is documented that local administration of vancomycin offers better results than the usual routes of administration (e.g., intravenous) when ischemic areas are present. In this work, we evaluate the antimicrobial efficacy against S. aureus and S. epidermidis of a novel hybrid 3D-printed scaffold based on polycaprolactone (PCL) and a chitosan (CS) hydrogel loaded with different vancomycin (Van) concentrations (1, 5, 10, 20%). Two cold plasma treatments were used to improve the adhesion of CS hydrogels to the PCL scaffolds by decreasing PCL hydrophobicity. Vancomycin release was measured by means of HPLC, and the biological response of ah-BM-MSCs growing in the presence of the scaffolds was evaluated in terms of cytotoxicity, proliferation, and osteogenic differentiation. The PCL/CS/Van scaffolds tested were found to be biocompatible, bioactive, and bactericide, as demonstrated by no cytotoxicity (LDH activity) or functional alteration (ALP activity, alizarin red staining) of the cultured cells and by bacterial inhibition. Our results suggest that the scaffolds developed would be excellent candidates for use in a wide range of biomedical fields such as drug delivery systems or tissue engineering applications.

Broad functional profiling of fission yeast proteins using phenomics and machine learning.

Many proteins remain poorly characterized even in well-studied organisms, presenting a bottleneck for research. We applied phenomics and machine-learning approaches with Schizosaccharomyces pombe for broad cues on protein functions. We assayed colony-growth phenotypes to measure the fitness of deletion mutants for 3509 non-essential genes in 131 conditions with different nutrients, drugs, and stresses. These analyses exposed phenotypes for 3492 mutants, including 124 mutants of 'priority unstudied' proteins conserved in humans, providing varied functional clues. For example, over 900 proteins were newly implicated in the resistance to oxidative stress. Phenotype-correlation networks suggested roles for poorly characterized proteins through 'guilt by association' with known proteins. For complementary functional insights, we predicted Gene Ontology (GO) terms using machine learning methods exploiting protein-network and protein-homology data (NET-FF). We obtained 56,594 high-scoring GO predictions, of which 22,060 also featured high information content. Our phenotype-correlation data and NET-FF predictions showed a strong concordance with existing PomBase GO annotations and protein networks, with integrated analyses revealing 1675 novel GO predictions for 783 genes, including 47 predictions for 23 priority unstudied proteins. Experimental validation identified new proteins involved in cellular aging, showing that these predictions and phenomics data provide a rich resource to uncover new protein functions.

[Procalcitonin as sepsis predictor in cardiovascular surgery with cardiopulmonary bypass]. / Procalcitonina como predictor de sepsis en cirugía cardiovascular con circulación extracorpórea.

Background: Cardiopulmonary bypass generates an exacerbated response that may lead to sepsis. Objective: To describe the association between procalcitonin levels and sepsis diagnosis in cardiovascular surgery subjects with cardiopulmonary bypass. Methods: A case-series study was conducted in 142 patients. Serum procalcitonin levels were measured at 24 hours and at 72 hours after surgery using a point of care testing based on quantitative immunochromatographic method. To assess association between procalcitonin levels and sepsis status, we calculated area under the curve (AUC) and sensitivity, specificity, and predictive values for the best cut-off point. Results: From 142 patients studied, 7 developed sepsis after surgery (4.9%). For 24-hours procalcitonin levels AUC was 0.921 and best cut-off point was 3.8 ng/mL (sensitivity 0.857 and specificity 0.904). In the case of 72-hours procalcitonin levels, we observed a value of 0.868 for AUC and best cut-off point was 8.4 ng/mL (sensitivity 0.86 and specificity 0.97). Conclusions: Procalcitonin levels at 24 and 72 hours after cardiovascular surgery with cardiopulmonary bypass are associated with sepsis presence at cut-off points of 3.8 and 8.4 ng/mL respectively.

Introducción: la circulación extracorpórea durante la cirugía cardiovascular genera una respuesta exacerbada que puede asociarse con sepsis. Objetivo: describir la asociación entre los niveles de procalcitonina y el diagnóstico de sepsis en sujetos de cirugía cardiovascular con circulación extracorpórea. Material y métodos: se realizó un estudio de serie de casos en 142 pacientes. Los niveles de procalcitonina fueron medidos a las 24 horas y a las 72 horas después de la cirugía. Para evaluar la asociación entre los niveles de procalcitonina y la identificación de sepsis, se calculó el área bajo la curva (AUC) y la sensibilidad y especificidad identificando el mejor punto de corte. Resultados: de un total de 142 pacientes estudiados, 7 desarrollaron sepsis (4.9%). En los niveles de procalcitonina en las 24 horas, el AUC fue de 0.921 y el mejor punto de corte fue 3.8 ng/mL (sensibilidad de 0.857 y especificidad de 0.904). En el caso de los niveles de procalcitonina a las 72 horas, observamos un AUC de 0.868 y el mejor punto de corte fue 8.4 ng/mL (sensibilidad de 0.86 y especificidad de 0.97). Conclusiones: los niveles de procalcitonina a las 24 y 72 horas de la cirugía cardiovascular con circulación extracorpórea se asociaron con la presencia de sepsis con los puntos de corte de 3.8 ng/mL y 8.4 ng/mL respectivamente.

Functional profiling of long intergenic non-coding RNAs in fission yeast.

Eukaryotic genomes express numerous long intergenic non-coding RNAs (lincRNAs) that do not overlap any coding genes. Some lincRNAs function in various aspects of gene regulation, but it is not clear in general to what extent lincRNAs contribute to the information flow from genotype to phenotype. To explore this question, we systematically analysed cellular roles of lincRNAs in Schizosaccharomyces pombe. Using seamless CRISPR/Cas9-based genome editing, we deleted 141 lincRNA genes to broadly phenotype these mutants, together with 238 diverse coding-gene mutants for functional context. We applied high-throughput colony-based assays to determine mutant growth and viability in benign conditions and in response to 145 different nutrient, drug, and stress conditions. These analyses uncovered phenotypes for 47.5% of the lincRNAs and 96% of the protein-coding genes. For 110 lincRNA mutants, we also performed high-throughput microscopy and flow cytometry assays, linking 37% of these lincRNAs with cell-size and/or cell-cycle control. With all assays combined, we detected phenotypes for 84 (59.6%) of all lincRNA deletion mutants tested. For complementary functional inference, we analysed colony growth of strains ectopically overexpressing 113 lincRNA genes under 47 different conditions. Of these overexpression strains, 102 (90.3%) showed altered growth under certain conditions. Clustering analyses provided further functional clues and relationships for some of the lincRNAs. These rich phenomics datasets associate lincRNA mutants with hundreds of phenotypes, indicating that most of the lincRNAs analysed exert cellular functions in specific environmental or physiological contexts. This study provides groundwork to further dissect the roles of these lincRNAs in the relevant conditions.