Unbiased integration of single cell transcriptome replicates Single cell transcriptomic approaches are becoming mainstream, with replicate experiments commonly performed with the same single cell technology. Methods that enable integration of these datasets by removing batch effects while preserving biological information are required for unbiased data interpretation. Here we introduce Canek for this purpose. Canek leverages information from mutual nearest neighbor to combine local linear corrections with cell-specific non-linear corrections within a fuzzy logic framework. Using a combination of real and synthetic datasets, we show that Canek corrects batch effects while introducing the least amount of bias compared with competing methods. Canek is computationally efficient and can easily integrate thousands of single-cell transcriptomes from replicated experiments.
Identifying complex sequence patterns in massive omics data with a variable-convolutional layer in deep neural network Motif identification is among the most common and essential computational tasks for bioinformatics and genomics. Here we proposed a novel convolutional layer for deep neural network, named Variable Convolutional (vConv) layer, for effective motif identification in high-throughput omics data by learning kernel length from data adaptively. Empirical evaluations on DNA-protein binding and DNase footprinting cases well demonstrated that vConv-based networks have superior performance to their convolutional counterparts regardless of model complexity. Meanwhile, vConv could be readily integrated into multi-layer neural networks as an "in-place replacement" of canonical convolutional layer. All source codes are freely available on GitHub for academic usage.
Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. Recombination marks the evolutionary dynamics of a recently endogenized retrovirus All vertebrate genomes have been colonized by retroviruses along their evolutionary trajectory. While endogenous retroviruses (ERVs) can contribute important physiological functions to contemporary hosts, such benefits are attributed to long-term co-evolution of ERV and host because germline infections are rare and expansion is slow, because the host effectively silences them. The genomes of several outbred species including mule deer (Odocoileus hemionus) are currently being colonized by ERVs, which provides an opportunity to study ERV dynamics at a time when few are fixed. Because we have locus-specific data on the distribution of cervid endogenous retrovirus (CrERV) in populations of mule deer, in this study we determine the molecular evolutionary processes acting on CrERV at each locus in the context of phylogenetic origin, genome location, and population prevalence. A mule deer genome was de novo assembled from short and long insert mate pair reads and CrERV sequence generated at each locus. CrERV composition and diversity have recently measurably increased by horizontal acquisition of a new retrovirus lineage. This new lineage has further expanded CrERV burden and CrERV genomic diversity by activating and recombining with existing CrERV. Resulting inter-lineage recombinants endogenized and subsequently retrotransposed. CrERV loci are significantly closer to genes than expected if integration were random and gene proximity might explain the recent expansion by retrotransposition of one recombinant CrERV lineage. Thus, in mule deer, retroviral colonization is a dynamic period in the molecular evolution of CrERV that also provides a burst of genomic diversity to the host population. Brominated Lipid Probes Expose Structural Asymmetries in Constricted Membranes Lipids in biological membranes are thought to be functionally organized, but few experimental tools can probe nanoscale membrane structure. Using brominated lipids as contrast probes for cryo-EM and a model ESCRT-III membrane remodeling system, we observed leaflet-level and protein-localized lipid structural patterns within highly constricted and thinned membrane nanotubes. These nanotubes differed markedly from protein-free, flat bilayers in leaflet thickness, lipid diffusion rates, and lipid compositional and conformational asymmetries. Simulations and cryo-EM imaging of brominated SDPC showed how a pair of phenylalanine residues scored the outer leaflet with a helical hydrophobic defect where polyunsaturated docosahexaenoyl (DHA) tails accumulated at the bilayer surface. Combining cryo-EM of halogenated lipids with molecular dynamics thus enables new characterizations of the composition and structure of membranes on molecular length scales.

One-Sentence SummaryCryo-EM imaging of halogenated lipids and MD simulations provide molecular-scale insight into constricted bilayer structure. The renal inflammatory network of nephronophthisis STRUCTURED ABSTRACTO_ST_ABSBACKGROUNDC_ST_ABSThe majority of genetic kidney disease leading to kidney failure is caused by mutations in ciliary genes. How cilia malfunction leads to progressive kidney damage is poorly understood, but recent evidence links ciliopathy genes to CCL2 dependent macrophage recruitment in autosomal dominant polycystic kidney disease (ADPKD), the most studied renal ciliopathy. Whether or not renal inflammation is involved in other renal ciliopathies is unclear.

METHODSWe combined mice models with kidney biopsies and renal epithelial cells sampled from human urine to characterize the renal inflammatory network of nephronophthisis (NPH), the most frequent renal ciliopathy in children.

RESULTSIn human, mutations in cilia genes involved in NPH enhance urine excretion of the chemokine CCL2, causing abnormal macrophage recruitment in kidney tissues from NPH patients. Differing from ADPKD, inactivating Ccl2 specifically in mouse tubular cells does not rescue the NPH phenotype, suggesting that other inflammatory mediators are involved. Using transcriptional data from 2 NPH models, we identify a set of pro-inflammatory cytokines upregulated in this disease, independently of CCL2. The majority of detectable transcripts from this set are specifically upregulated in kidney cells from NPH patients. In line with the function of these cytokines, NPH kidneys show disproportionate neutrophils and T cells infiltrates compared to healthy subject or hypertensive and diabetic chronic kidney disease patients.

CONCLUSIONSThis study reveals that inflammation is a central aspect in human NPH and delineates a specific set of inflammatory mediators that regulates immune cell recruitment in human NPH.

SIGNIFICANCE STATEMENTMutations in genes encoding primary cilia proteins are the leading cause of genetic kidney failure. In autosomal dominant polycystic kidney disease (ADPKD), deregulated cilia signaling leads to kidney infiltration by macrophages through the chemokine CCL2. Little is known about renal inflammation in nephronophthisis (NPH), the most frequent pediatric renal ciliopathy. Using NPH mice models, tissues and cells from NPH patients, we unveil renal inflammation as preeminent feature of NPH. Remarkably, the renal inflammatory evoked by ciliary gene mutations in NPH does not overlap with ADPKD: it is CCL2 independent, involves a prominent recruitment of neutrophils and T cells and a specific cytokine signature. This unforeseen findings strengthen the link between primary cilia and renal inflammation. Clonal Dynamics of Cancer-Initiating Cells in a Mouse Genetic Mosaic Model of Serous Ovarian Cancer High-grade serous ovarian cancer (HGSOC) is the most lethal gynecologic malignancy. Early detection remains the most promising strategy to decrease mortality and requires a deep understanding of the cancer initiation process. Recent breakthroughs identified the fallopian tube (FT) Pax8+ cells as a cell-of-origin for HGSOC. However, the progressive trajectory from an initiated Pax8+ cell toward malignancy remains elusive. Here, we use a mouse genetic mosaic system called MADM (Mosaic Analysis with Double Markers), which generates sporadic mutant cells labeled with GFP, to trace the clonal dynamics of HGSOC development. Surprisingly, we observed dichotomous clonal expansion kinetics of individual cancer-initiating Pax8+ cells; while the vast majority stalled immediately, a few significantly expanded. Clonal analysis of wildtype cells showed concordant dichotomy, suggesting that a small fraction of Pax8+ cells intrinsically carry higher expansion potential. Furthermore, although all wildtype clones stalled shortly after development, some mutant clones continued to progress, indicating that oncogenic mutations prolong clonal expansion beyond the cell-intrinsic potential. Finally, intra-clonal fate mapping revealed that progressing mutant clones have a higher propensity to produce Pax8+ cells, linking imbalanced cell differentiation to disease progression. In summary, our studies reveal that the earliest clonal expansion during HGSOC initiation emerges when oncogenic mutations exaggerate the intrinsic expansion potential of a rare Pax8+ subset and bias its differentiation potential, warranting future investigation as a readout for early detection and cancer prevention.

SIGNIFICANCE STATEMENTEffective early detection is the most promising approach to manage cancer, but it requires a thorough understanding of cancer initiation and early progression. Serous ovarian cancer has been found to originate from Pax8+ cells in the fallopian tube. To assess the clonal dynamics of cancer-initiating cells, we established a mouse genetic mosaic model that enables clonal analysis of mutant Pax8+ cells in the fallopian tube. We uncovered that the earliest clonal event in ovarian cancer development emerges from a rare subset of stem/progenitor-like cells with intrinsic expansion potential, and that oncogenic mutations exaggerate and prolong this potential to promote cancer initiation. Our findings pinpoint a precise target for future early-detection and cancer prevention studies.

HIGHLIGHTSO_LIOvarian cancer initiation revealed by a mouse genetic mosaic model.
C_LIO_LIDichotomous expansion capacity of initiated cells at the clonal level.
C_LIO_LISynergies between cell-intrinsic properties and onco-mutations fuels clonal expansion.
C_LIO_LIImbalanced differentiation within highly expansive clones leads to Pax8 clusters.
C_LI Computation of the mitochondrial age distribution along the axon length We describe a compartmental model of mitochondrial transport in axons, which we apply to compute mitochondrial age at different distances from the soma. The model predicts that the average mitochondrial age at the tip of an axon with a length of 1 cm is approximately 22 hours. The mitochondria are youngest closest to the soma and their age scales approximately linearly with distance from the soma. To the best of the authors knowledge, this is the first attempt to predict the spatial distribution of mitochondrial age within an axon. A sensitivity study of the mean age of mitochondria to various model parameters is also presented. A Hierarchy of Biomolecular Proportional-Integral-Derivative Feedback Controllers for Robust Perfect Adaptation and Dynamic Performance Proportional-Integral-Derivative (PID) feedback controllers have been the most widely used controllers in industry for almost a century due to their good performance, simplicity, and ease of tuning. Motivated by their success in various engineering disciplines, PID controllers recently found their way into synthetic biology, where the design of feedback molecular control systems has been identified as an important goal. In this paper, we consider the mathematical realization of PID controllers via biomolecular interactions. We propose an array of topologies that offer a compromise between simplicity and high performance. We first demonstrate that different Proportional-Integral (PI) controllers exhibit different capabilities for enhancing the dynamics and reducing variance (cell-to-cell variability). Next, we introduce several derivative controllers that are realized based on incoherent feedforward loops acting in a feedback configuration. Alternatively, we show that differentiators can be realized by placing molecular integrators in a negative feedback loop--an arrangement that can then be augmented by PI components to yield PID feedback controllers. We demonstrate that the derivative component can be exploited for enhancing system stability, dramatically increasing the molecular control systems dynamic performance, and for reducing the noise effect on the output of interest. The PID controller features are established through various deterministic and stochastic analyses as well as numerical simulations. Finally, we provide an experimental demonstration using a recently developed hybrid setup, the cyberloop, where the controller is implemented in silico to control a biological genetic circuit in vivo. The large array of novel biomolecular PID controllers introduced here forms a basis for the design and construction of advanced high-performance biomolecular control systems that robustly regulate the dynamics of living systems. Second Heart Field-derived Cells Contribute to Angiotensin II-mediated Ascending Aortopathies BackgroundThe ascending aorta is a common location for aneurysm and dissection. This aortic region is populated by a mosaic of medial and adventitial cells that are embryonically derived from either the second heart field (SHF) or the cardiac neural crest. SHF-derived cells populate areas that coincide with the spatial specificity of thoracic aortopathies. The purpose of this study was to determine whether and how SHF-derived cells contribute to ascending aortopathies.

MethodsAscending aortic pathologies were examined in patients with sporadic thoracic aortopathies and angiotensin II (AngII)-infused mice. Ascending aortas without overt pathology from AngII-infused mice were subjected to mass spectrometry assisted proteomics, and molecular features of SHF-derived cells were determined by single cell transcriptomic analyses. Genetic deletion of either low-density lipoprotein receptor-related protein 1 (Lrp1) or transforming growth factor-{beta} receptor 2 (Tgfbr2) in SHF- derived cells was conducted to examine the impact of SHF-derived cells on vascular integrity.

ResultsPathologies in human ascending aortic aneurysmal tissues were predominant in outer medial layers and adventitia. This gradient was mimicked in mouse aortas following AngII infusion that was coincident with the distribution of SHF-derived cells. Proteomics indicated that brief AngII infusion, prior to overt pathology, evoked downregulation of SMC proteins and differential expression of extracellular matrix proteins, including several LRP1 ligands. LRP1 deletion in SHF-derived cells augmented AngII-induced ascending aortic aneurysm and rupture. Single cell transcriptomic analysis revealed that brief AngII infusion decreased Lrp1 and Tgfbr2 mRNA abundance in SHF-derived cells and induced a unique fibroblast population with low abundance of Tgfbr2 mRNA. SHF-specific Tgfbr2 deletion led to embryonic lethality at E12.5 with dilatation of the outflow tract and retroperitoneal hemorrhage. Integration of proteomic and single cell transcriptomics results identified plasminogen activator inhibitor 1 (PAI1) as the most increased protein in SHF-derived SMCs and fibroblasts during AngII infusion. Immunostaining revealed a transmural gradient of PAI1 in both ascending aortas of AngII-infused mice and human ascending aneurysmal aortas that mimicked the gradient of medial and adventitial pathologies.

ConclusionSHF-derived cells exert a critical role in maintaining vascular integrity through LRP1 and TGF-{beta} signaling associated with increases of aortic PAI1.



O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/930917v6_ufig1.gif" ALT="Figure 1">
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org.highwire.dtl.DTLVardef@185e3d9org.highwire.dtl.DTLVardef@131d702org.highwire.dtl.DTLVardef@11afa42org.highwire.dtl.DTLVardef@242786_HPS_FORMAT_FIGEXP  M_FIG C_FIG Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABS- SHF-derived SMCs and fibroblasts associate with AngII-induced aortic pathologies.
- AngII induces a distinct fibroblast sub-cluster that is less abundant for mRNAs related to major extracellular components and TGF{beta} ligands and receptors, but more abundant for proliferative genes.
- TGFBR2 deletion in SHF-derived cells are embryonic lethal with significant dilatation of the outflow tract in mice.
- SHF-specific deletion of LRP1 leads to aortic pathologies in mice, supporting the importance of SHF-derived cells in maintaining ascending aortic wall integrity.


What are the clinical implications?- Heterogeneity of the embryonic origins of SMCs and fibroblasts contributes to complex mechanisms of vasculopathy formation, which should be considered when investigating the pathogenesis of thoracic aortopathies. Disruption of the microphysiological niche alters matrix deposition and causes loss of mitochondrial Ca2+ control in skeletal muscle fibers Cells rapidly lose their physiological phenotype upon disruption of their extracellular matrix (ECM)-intracellular cytoskeleton interactions. Here, we investigated acute effects of ECM disruption on cellular and mitochondrial morphology, transcriptomic signatures, and Ca2+ handling in adult mouse skeletal muscle fibers. Adult skeletal muscle fibers were isolated from mouse toe muscle either by collagenase-induced dissociation of the ECM or by mechanical dissection that leaves the contiguous ECM intact. Experiments were generally performed four hours after cell isolation. At this time, there were striking differences in the gene expression patterns between fibers isolated with the two methods; 24h after cell isolation, enzymatically dissociated fibers had transcriptomic signatures resembling dystrophic phenotypes. Mitochondrial appearance was grossly similar in the two groups, but 3D electron microscopy revealed shorter and less branched mitochondria in enzymatically dissociated than in mechanically dissected fibers. Similar increases in free cytosolic [Ca2+] during repeated tetanic stimulation were accompanied by marked mitochondrial Ca2+ uptake only in enzymatically dissociated muscle fibers. The aberrant mitochondrial Ca2+ uptake was partially prevented by the mitochondrial Ca2+ uniporter inhibitor Ru360 and by cyclosporine A and NV556, which inhibit the mitochondrial protein Ppif (also called cyclophilin D). Importantly, inhibition of Ppif with NV556 significantly improved survival of mice with mitochondrial myopathy in which muscle mitochondria take up excessive amounts of Ca2+ even with an intact ECM. In conclusion, skeletal muscle fibers isolated by collagenase-induced dissociation of the ECM display aberrant mitochondrial Ca2+ uptake, which involves a Ppif-dependent mitochondrial Ca2+ influx resembling that observed in mitochondrial myopathies. A constraints-based theory of the primary cause of senescence: imbalance of epigenetic and non-epigenetic information in histone crosstalk Cellular aging has been progressively elucidated by science. However, the fundamental cause of senescence--i.e., why organisms age at the multicellular-individual level--remains unclear. A recent theory of individuated multicellularity describes the emergence and growth of crucial information content for cell differentiation. This information is mostly conveyed in the non-epigenetic (i.e., transcription uncorrelated) histone crosstalk near transcription start sites. According to this theory, the non-epigenetic content emerges and grows at the expense of the information capacity for epigenetic content. If this "reassignment" of information capacity continues after adulthood, it may explain the senescence phenomenon. Here, I present a novel, falsifiable theory describing an uninterrupted growth of capacity for non-epigenetic information at the expense of that for epigenetic information not only during ontogeny but also throughout adulthood. As a byproduct, this continuous "reassignment" of capacity effectively creates an information imbalance in histone crosstalk, which in turn overregulates transcriptional levels. This overregulation is to be understood as transcriptional levels becoming more and more accurate but also less and less precise with respect to the needs of the multicellular individual--up to the point of dysfunctionality. This epigenetic/non-epigenetic information imbalance is proposed to be the primary reason why individuated multicellular organisms senesce. Alteration of gut microbiota with a broad-spectrum antibiotic does not impair maternal care in the European earwig. AO_SCPLOWBSTRACTC_SCPLOWThe microbes residing within the gut of an animal host often increase their own fitness by modifying their hosts physiological, reproductive, and behavioural functions. Whereas recent studies suggest that they may also shape host sociality and therefore have critical effects on animal social evolution, the impact of the gut microbiota on maternal care remains unexplored. This is surprising, as this behaviour is widespread among animals, often determines the fitness of both juveniles and parents, and is essential in the evolution of complex animal societies. Here, we tested whether life-long alterations of the gut microbiota with rifampicin - a broad-spectrum antibiotic - impair pre- and post-hatching maternal care in the European earwig. Our results first confirm that rifampicin altered the mothers gut microbial communities and indicate that the composition of the gut microbiota differs before and after egg care. Contrary to our predictions, however, the rifampicin-induced alterations of the gut microbiota did not modify pre- or post-hatching care. Independent of maternal care, rifampicin increased the females feces production and resulted in lighter eggs and juveniles. By contrast, rifampicin altered none of the other 21 physiological, reproductive and longevity traits measured over the 300 days of a females lifetime. Overall, these findings reveal that altering the gut microbiota with a large spectrum antibiotic such as rifampicin does not necessarily affect host sociality. They also emphasize that not all animals have evolved a co-dependence with their microbiota and call for caution when generalizing the central role of gut microbes in host biology. First detection of herpesvirus and prevalence of mycoplasma infection in free-ranging Hermann s tortoises (Testudo hermanni), and in potential pet vectors AO_SCPLOWBSTRACTC_SCPLOWTwo types of pathogens cause highly contagious upper respiratory tract diseases (URTD) in Chelonians: testudinid herpesviruses (TeHV) and a mycoplasma (Mycoplasma agassizii). In captivity, these infections are frequent and can provoke outbreaks. Pet trade generates international flow of tortoises, often without sanitary checking; individuals intentionally or accidentally released in the wild may spread pathogens. A better understanding of the transmission of infectious agents from captivity to wild tortoises is needed. Many exotic individuals have been introduced in populations of the endangered western Hermanns tortoise (Testudo hermanni hermanni), notably spur-thighed tortoises (Testudo graeca). We assessed the presence of TeHV and mycoplasma in native western Hermanns tortoises and in potential pet vectors in south-eastern France. Using a large sample (N=572 tortoises), this study revealed, by PCR, the worrying presence of herpesvirus in 7 free-ranging individuals (3 sub-populations). Additionally, Mycoplasma agassizii was detected, by PCR, in 15 of the 18 populations sampled with a frequency ranging from 3.4% (1 of 29 tortoises) to 25% (3 of 12 tortoises). Exotic spur-thighed tortoises showed high frequency of mycoplasma infection in captivity (18.2%) and in individuals (50%) found in native Hermanns tortoise sub-populations, suggesting that this species could be a significant vector. The paucity of information of TeHV on European tortoise URTD in natural settings, especially in combination with mycoplasma, prompts for further studies. Indeed, sick tortoises remain concealed and may not be easily detected in the field. Our results indicate a good health for most infected tortoise but it should be screened in the field as well as in captivity. Towards a unified model of naive T cell dynamics across the lifespan Naive CD4 and CD8 T cells are cornerstones of adaptive immunity, but the dynamics of their establishment early in life and how their kinetics change as they mature following release from the thymus are poorly understood. Further, due to the diverse signals implicated in naive T cell survival, it has been a long-held and conceptually attractive view that they are sustained by active homeostatic control as thymic activity wanes. Here we employ multiple experimental systems to identify a unified model of naive CD4 and CD8 T cell population dynamics across the mouse lifespan. We infer that both subsets divide rarely and progressively increase their survival capacity with cell age. Strikingly, this simple model captures naive CD4 T cell dynamics throughout life. In contrast, we find that newly generated naive CD8 T cells are lost more rapidly during the first 3-4 weeks of life, likely due to increased recruitment into memory. We find no evidence for elevated division rates in neonates, or for feedback regulation of naive T cell numbers at any age. We show how confronting mathematical models with diverse datasets can reveal a quantitative and remarkably simple picture of naive T cell dynamics from birth into old age. Late-acting self-incompatible system, preferential allogamy and delayed selfing in the heteromorphic invasive populations of Ludwigia grandiflora subsp. hexapetala AO_SCPLOWBSTRACTC_SCPLOWBreeding system influences local population genetic structure, effective size, offspring fitness and functional variation. Determining the respective importance of self- and cross-fertilization in hermaphroditic flowering plants is thus important to understand their ecology and evolution. The worldwide invasive species, Ludwigia grandiflora subsp. hexapetala (Lgh) presents two floral morphs: one self-compatible short-styled morph (S-morph) and one self-incompatible long-styled morph (L-morph). In this study, we identified the breeding systems of western European experimental and natural populations of Lgh by comparing structural characteristics of pollen and style, by studying self- and cross-pollen tube elongations and the viability of the resulting seeds and seedlings in both floral morphs. Our results showed no differences in pollen shape and stigma surfaces no matter the floral morph. In the self-incompatible L-morph flowers, self-pollen tubes were stopped tardily, in the ovarian area, and were unable to fertilize the ovules. This first formal identification of a late-acting, prezygotic self-incompatible system (LSI) in Ludwigia genus contributes a case of LSI in an additional family within the Myrtales order. In the self-compatible S-morph flowers, self-pollen always succeeded to self-fertilize the ovules that nearly all developed into viable seedlings. However, cross-pollen tubes always elongated faster than self-pollen tubes in both morphs. S-morph individuals may thus advantage preferential allogamy over selfing when cross-pollen is available despite its self-compatibility. As expected in late-acting self-incompatible systems, L-morph flowers authorised 0.2{per thousand} of selfed seeds during the uppermost flowering season, that increased to 1{per thousand} at the end of the flowering season. Such delayed selfing resulted in a significant quantity of viable floating seeds. They may contribute to the local regeneration, seed bank and propagation of the L-morph, which may contribute to explain its invasion success worldwide. Management plans of Lgh would gain to consider the breeding systems we identified. Synchronized Neural Activity Indexes Engagement with Spoken Stories Despite Acoustic Masking Older people with hearing problems often experience difficulties understanding speech in the presence of background sound. As a result, they may disengage in social situations, which has been associated with negative psychosocial health outcomes. Measuring listening (dis-)engagement during challenging listening situations has received little attention thus far. We recruit young, normal-hearing human adults (both sexes) and investigate how speech intelligibility and engagement during naturalistic story listening is affected by the level of acoustic masking (12-talker babble) at different signal-to-noise ratios (SNRs). In Experiment 1, we observed that word-report scores were above 80% for all but the lowest SNR (-3 dB SNR) we tested, at which performance dropped to 54%. In Experiment 2, we calculated inter-subject correlation (ISC) using electroencephalography (EEG) data to identify dynamic spatial patterns of shared neural activity evoked by the stories. ISC has previously been used as a neural measure of participants engagement with naturalistic materials. Our results show that ISC was stable across all but the lowest SNRs, despite reduced speech intelligibility. Comparing ISC and intelligibility demonstrated that word-report performance declined more strongly with decreasing SNR compared to ISC. Our measure of neural engagement suggests that individuals remain engaged in story listening despite missing words due to background noise. Our work provides a potentially fruitful approach to investigate listener engagement with naturalistic, spoken stories that may be used to investigate (dis)engagement in older adults with hearing impairment. Quercetin handles cellular oxidant / antioxidant systems and mitigates immunosenescence hallmarks in human PBMCs: an in vitro study Immunosenescence, oxidative stress, and low vaccine efficacy are important symptoms of aging. The goal of our study was to see if quercetin had anti-aging and stimulating effects on PBMC immune cells in vitro. In the presence of concanavalin a, PBMCs were isolated from healthy elderly and young people and cultured in a complete RPMI-1640 medium supplemented with quercetin. Cell proliferation was assessed using the MTT colorimetric assay after a 48-hour incubation period. Spectrophophotometric assays were used to assess oxidative biomarkers (PC, MDA, and GSH). The ELISA method was used to determine the amount of IL-2 released. A Griess reagent was used to investigate iNOS activity. When compared to young control cells, aged PBMCs had lower proliferation potency, lower IL-2 and NO release, and higher MDA and PC levels. Importantly, quercetin-treated aged PBMCs have a high proliferative response comparable to young cells, restored iNOS activity, and increased levels of GSH antioxidant defences. In comparison to untreated aged PBMCs, treated PBMCs have lower lipo-oxidative damage but higher PC levels. Quercetin may be used as a promising dietary vaccinal adjuvant in the elderly, it has significant effects in reducing immunosenescence hallmarks, as well as mitigating the lipo-oxidative stress in PBMCs cells.

Practical abstractQuercetin is a common dietary polyphenol found in fruits and vegetables that is well known for its antioxidant activity in radical scavenging and anticancer properties characterized by immune system stimulation. In vitro, quercetin boosts immune responses in the elderly and reduces immunosenescence symptoms, according to our findings. As a result, this molecule could be a safe and promising vaccine adjuvant for boosting immunity and reducing aging complications in immune cells as well as reducing oxidative stress in PBMC cells. Protein cost minimization promotes the emergence of coenzyme redundancy Coenzymes distribute a variety of chemical moieties throughout cellular metabolism, participating in group (e.g., phosphate, acyl) and electron transfer. For a variety of reactions requiring acceptors or donors of specific resources, there often exist degenerate sets of molecules (e.g., NAD(H) and NADP(H)) that carry out similar functions. Although the physiological roles of various coenzyme systems are well established, it is unclear what selective pressures may have driven the emergence of coenzyme redundancy. Here we use genome-wide metabolic modeling approaches to decompose the selective pressures driving enzymatic specificity for either NAD(H) or NADP(H) in the metabolic network of Escherichia coli. We found that few enzymes are thermodynamically constrained to using a single coenzyme, and in principle, a metabolic network relying on only NAD(H) is feasible. However, structural and sequence analyses revealed widespread conservation of residues that retain selectivity for either NAD(H) or NADP(H), suggesting that additional forces may shape specificity. Using a model accounting for the cost of oxidoreductase enzyme expression, we found that coenzyme redundancy universally reduces the minimal amount of protein required to catalyze coenzyme-coupled reactions, inducing individual reactions to strongly prefer one coenzyme over another when reactions are near thermodynamic equilibrium. We propose that protein minimization generically promotes coenzyme redundancy, and that coenzymes typically thought to exist in a single pool (e.g., CoA) may exist in more than one form (e.g., dephospho-CoA).

Significance statementMetabolism relies on a small class of molecules (coenzymes) that serve as universal donors and acceptors of key chemical groups and electrons. Although metabolic networks crucially depend on structurally redundant coenzymes (e.g., NAD(H) and NADP(H)) associated with different enzymes, the criteria that led to the emergence of this redundancy remain poorly understood. Our combination of modeling, and structural and sequence analysis indicates that coenzyme redundancy is not essential for metabolism, but rather an evolved strategy promoting efficient usage of enzymes when biochemical reactions are near equilibrium. Our work suggests that early metabolism may have operated with fewer coenzymes, and that adaptation for metabolic efficiency may have driven the rise of coenzyme diversity in living systems. Comparison of hybrid learning and remote education in the implementation of the 'Adopt a Microorganism' methodology The Internet has changed the way teachers and students access information and build knowledge. The recent COVID-19 pandemic has created challenges for both teachers and students and a demand for new methodologies of remote learning. In the life sciences, mixing online content with practical activities represents an even greater challenge. In microbiology, the implementation of an active teaching methodology, the #Adopt project, based on the social network Facebook(R), represents an excellent option for connecting remote education with classroom activities. In 2020, the version applied in high school, "Adopt a Microorganism", was adapted to meet the demands of emergency remote education owing to the suppression of face-to-face activities caused by the pandemic. In the present study, we assessed how the change in methodology impacted the discourse richness of students from high school integrated with technical education in the Business Administration program of the Federal Institute of Sao Paulo, Sorocaba Campus. Three questionnaires related to the groups of microorganisms (Archaea, Bacteria, Virus, Fungi, and Protozoan) were applied. The students responses in the 2019 and 2020 classes were compared concerning content richness and multiplicity of concepts through the application of the Shannon diversity index, an approach that is generally used to assess biodiversity in different environments. The observed results suggest that remote learning provided students with a conceptual basis and richness of content equivalent to that achieved by students subjected to the hybrid teaching model. In conclusion, this study suggests that the #Adopt project methodology increases students discourse richness in microbiology even without face-to-face traditional classes. S-acylation stabilizes ligand-induced receptor kinase complex formation during plant pattern-triggered immune signalling. Plant receptor kinases are key transducers of physical extracellular stimuli, such as the presence of beneficial or pathogenic microbes or secreted signalling molecules. Receptor kinases are regulated by numerous post-translational modifications. Here, using the bacterial flagellin perceiving receptor kinase FLS2, we show that S-acylation at an evolutionarily conserved cysteine is crucial for function. S-acylation involves the addition of long-chain fatty acids to cysteine residues within proteins, altering their biophysical properties and behaviour within the membrane environment. We observe S-acylation of FLS2 at C-terminal kinase domain cysteine residues within minutes of treatment with flg22 ligand, and in a BAK1 co-receptor dependent manner. We demonstrate that S-acylation is essential for FLS2-mediated immune signalling, including anti-bacterial immunity. Similarly, mutating the corresponding conserved cysteine residue in the immune receptor kinase EFR supressed elf18 mediated signalling. Biochemical analysis of unstimulated and activated FLS2 containing complexes using detergents and native membrane DIBMA nanodiscs indicates that S-acylation assists the stabilization of activated receptor kinase complexes withing the membrane environment to increase signalling efficiency. Plasma proteome analyses in individuals of European and African ancestry identify cis-pQTLs and models for proteome-wide association studies Improved understanding of genetic regulation of proteome can facilitate the identification of causal mechanisms for complex traits. We analyzed data on 4,657 plasma proteins from 7,213 European American (EA) and 1,871 African American (AA) individuals from the ARIC study, and further replicated findings on 467 AA individuals from the AASK study. Here we identified 2,004 proteins in EA and 1,618 in AA, with majority overlapping, which showed associations with common variants in cis-regions. Availability of AA samples led to smaller credible sets and significant number of population-specific cis-pQTLs. Elastic-net produced powerful models for protein prediction in both populations. An application of proteome-wide association studies (PWAS) to serum urate and gout, implicated several proteins, including IL1RN, revealing the promise of the drug anakinra to treat acute gout flares. Our study demonstrates the value of large and diverse ancestry study for genetic mechanisms of molecular phenotypes and their relationship with complex traits. Sound localization in world and head-centered space in ferrets The location of sounds can be described in multiple coordinate systems that are defined relative to ourselves, or the world around us. Evidence from neural recordings in animals point towards the existence of both head-centered and world-centered representations of sound location in the brain; however, it is unclear whether such neural representations have perceptual correlates in the sound localization abilities of non-human listeners. Here, we establish novel behavioral tests to determine the coordinate systems in which ferrets can localize sounds. We found that ferrets could learn to discriminate between sound locations that were fixed in either world-centered or head-centered space, across wide variations in sound location in the alternative coordinate system. Using probe sounds to assess broader generalization of spatial hearing, we demonstrated that in both head and world-centered tasks, animals used continuous maps of auditory space to guide behavior. Single trial responses of individual animals were sufficiently informative that we could then model sound localization using speaker position in specific coordinate systems and accurately predict ferrets actions in held-out data. Our results indicate that an animal model in which neurons are known to be tuned to sound location in egocentric and allocentric reference frames can also localize sounds in multiple head and world-centered spaces.

Significance StatementHumans can describe the location of sounds either relative to themselves, or in the world, independent of their momentary position. These different spaces are also represented in the activity of neurons in animals, but its not clear whether non-human listeners also perceive both head and world-centered sound location. Here, we designed two behavioral tasks in which ferrets had to discriminate between two sounds using their position in the world, or relative to the head. Subjects learnt to solve both problems and showed the ability to generalize sound location in each space when presented with infrequent probe sounds. These findings reveal a perceptual correlate of neural sensitivity previously observed in the ferret brain and establish that, like humans, ferrets can access an auditory map of their local environment. Active antithrombin glycoforms are selectively physiosorbed on plasma extracellular vesicles Antithrombin (AT) is a glycoprotein produced by the liver and a principal antagonist of active clotting proteases. A deficit in AT function leads to AT qualitative deficiency, challenging to diagnose. Here we report that active AT may travel physiosorbed on the surface of plasma extracellular vesicles (EVs), contributing to form the "EV-protein corona". The corona is enriched in specific AT glycoforms, thus suggesting glycosylation to play a key role in AT partitioning between EVs and plasma. Differences in AT glycoform composition of the corona of EVs separated from plasma of healthy and AT qualitative deficiency-affected subjects were also noticed. This suggests deconstructing the plasma into its nanostructured components, as extracellular vesicles, could help to unravel pathophysiological mechanisms otherwise undiscovered. Rapid genotyping of tilapia lake virus (TiLV) using Nanopore sequencing Infectious diseases represent one of the major challenges to sustainable aquaculture production. Rapid, accurate diagnosis and genotyping of emerging pathogens during early-suspected disease cases is critical to facilitate timely response to deploy adequate control measures and prevent or reduce spread. Currently, most laboratories use PCR to amplify partial pathogen genomic regions, occasionally combined with sequencing of PCR amplicon(s) using conventional Sanger sequencing services for confirmatory diagnosis. The main limitation of this approach is the lengthy turnaround time. Here, we report an innovative approach using a previously developed specific PCR assay for pathogen diagnosis combined with a new Oxford Nanopore Technologies (ONT)-based amplicon sequencing method for pathogen genotyping. Using fish clinical samples, we applied this approach for the rapid confirmation of PCR amplicon sequences identity and genotyping of tilapia lake virus (TiLV), a disease-causing virus affecting tilapia aquaculture globally. The consensus sequences obtained after polishing exhibit strikingly high identity to references derived by Illumina and Sanger methods (99.83-100%). This study suggests that ONT-based amplicon sequencing is a promising platform to deploy in regional aquatic animal health diagnostic laboratories in low and medium income countries, for fast identification and genotyping of emerging infectious pathogens from field samples within a single day. Swine H1N1 influenza virus variants with enhanced polymerase activity and HA stability promote airborne transmission in ferrets Understanding how animal influenza A viruses (IAVs) acquire airborne transmissibility in humans and ferrets is needed to prepare for and respond to pandemics. Here, we investigated in ferrets the replication and transmission of swine H1N1 isolates P4 and G15, whose majority population had decreased polymerase activity and poor HA stability, respectively. For both isolates, a minor variant was selected and transmitted in ferrets. Polymerase-enhancing variant PA-S321 airborne-transmitted and propagated in one ferret. HA-stabilizing variant HA1-S210 was selected in all G15-inoculated ferrets and was transmitted by contact and airborne routes. With an efficient polymerase and a stable HA, the purified minor variant G15-HA1-S210 had earlier and higher peak titers in inoculated ferrets and was recovered at a higher frequency after airborne transmission than P4 and G15. Overall, HA stabilization played a more prominent role than polymerase enhancement in the replication and transmission of these viruses in ferrets. The results suggest pandemic risk-assessment studies may benefit from deep sequencing to identify minor variants with human-adapted traits.

IMPORTANCEDiverse IAVs circulate in animals, yet few acquire the viral traits needed to start a human pandemic. A stabilized HA and mammalian-adapted polymerase have been shown to promote the adaptation of IAVs to humans and ferrets (the gold-standard model for IAV replication, pathogenicity, and transmissibility). Here, we used swine IAV isolates of the gamma lineage a model to investigate the importance of HA stability and polymerase activity in promoting replication and transmission in ferrets. These are emerging viruses that bind to both -2,6- and -2,3-linked receptors. Using isolates containing mixed populations, a stabilized HA was selected within days in inoculated ferrets. An enhanced polymerase was also selected and propagated after airborne transmission to a ferret. Thus, HA stabilization was a stricter requirement, yet both traits promoted transmissibility. Knowing the viral traits needed for pandemic potential, and the relative importance of each, will help identify emerging viruses of greatest concern.