Robust and efficient online auditory psychophysics Most human auditory psychophysics research has historically been conducted in carefully controlled environments with calibrated audio-equipment, and over potentially hours of repetitive testing with expert listeners. Here, we operationally define such conditions as having high  auditory hygiene. Hereof, conducting auditory psychophysical paradigms online presents a serious challenge, in that results may hinge on absolute sound presentation level, reliably estimated perceptual thresholds, and sustained motivation and attention. We introduce a set of procedures that address these challenges and facilitate auditory hygiene for online auditory psychophysics. First, we establish a simple means of setting sound presentation levels. Across a set of four level-setting conditions conducted in person, we demonstrate the stability and robustness of this amplitude setting procedure in open air and controlled settings. Second, we test participants tone-in-noise thresholds using widely adopted online experiment platforms and demonstrate that reliable threshold estimates can be derived online in approximately one minute of testing. Third, using these amplitude and threshold setting procedures to establish participant-specific stimulus conditions, we show that an online implementation of the classic probe-signal paradigm can be used to demonstrate frequency-selective attention on an individual participant basis, using a third of the trials used in recent in-lab experiments. Finally, we show how threshold and attentional measures relate to well-validated assays of online participants in-task motivation, fatigue, and confidence. This demonstrates the promise of online auditory psychophysics for asking new auditory neuroscience questions quickly, efficiently, and with more diverse samples. Code for the tests is publicly available through Pavlovia and Gorilla.
Low-level Action Schemas Support Gaze Guidance Behavior for Action Planning and Execution in Novel Tasks AO_SCPLOWBSTRACTC_SCPLOWEye movements in the natural environment have primarily been studied for over-learned activities such as tea-making, sandwich-making, and hand-washing, which have a fixed sequence of actions associated with them. These studies indicate an interplay of low-level cognitive schemas that facilitate task completion. However, it is unclear if this strategy is also in play when the task is novel and a sequence of actions must be planned. Here, to study attention mechanisms, we recorded gaze and body movements in a naturalistic sorting task. In a virtual environment, subjects moved objects on a life-size shelf to achieve a given order. To study the action planning and execution related gaze guidance behavior we also controlled the complexity of the tasks, i.e. the objects were to be sorted based on a single or two different object features. We show that subjects are close to optimal while performing the easy tasks and are more sub-optimal while performing the more complex tasks. Fixations aligned with action onset showed gaze is tightly coupled with the action sequence and less so with task complexity. Task complexity elicited differences at specific time points where oculomotor behavior could be categorized into searching, guiding, checking fixations. Task complexity also affected the scan-paths on the task-relevant regions of interest during action planning and execution where subjects exhibit a greater search and action monitoring behavior in more complex tasks. Furthermore, task complexity also affected the timing of first fixations on the task-relevant regions of interest for action planning but not for action execution. Finally, the scan paths and the temporal sequence of first fixations suggest a just-in-time strategy of gaze allocation. Based on the spatio-temporal features of gaze control, we can conclude that eye movements during novel tasks correspond closely to low-level cognitive schemas that support the production and execution of actions.

Author SummaryEye movements in the natural environment have primarily been studied for over-learned and habitual everyday activities (tea-making, sandwich-making, hand-washing) which have a fixed sequence of actions associated with them. These studies typically categorize eye movements related to low level action plans of locating the object of interest, directing the body or hand to the object, and monitoring the action execution. In this study, we were interested in generalizing these task-oriented eye movements for novel tasks that are not associated with an inherent action sequence. To that end, we asked subjects to sort objects based on object features on a life-size shelf in a virtual environment as we recorded their eye and body movements. We then analyzed gaze features relative to the action onset i.e. when the hand made contact with an object. By only studying the sequence of fixations based on past, current and next actions, we aimed to elucidate the general characteristics of gaze behavior relative to action planning and execution and independent of object identity. With our analysis, we could categorize fixations into searching, guiding, monitoring fixations in a data-driven manner. Based on the spatio-temporal features of gaze control, we can conclude that novel tasks predominantly elicit eye movements corresponding to low-level action plans that support the production and execution of actions.
Natural variations in reproductive timing of brood size and X-chromosome nondisjunction in Caenorhabditis elegans Reproductive timing underlies self-sperm production and X-chromosome nondisjunction rate in Caenorhabditis elegans. These traits can be measured as brood size and male-production rate, and brood size decreases and male-production rate increases as worms age. These phenotypic changes occur simultaneously; however, whether these changes share common underlying genetic architectures still remains unclear. To enhance our understanding of reproductive timing in C. elegans, we measured and compared these two reproductive timing-related traits of 96 wild strains during early, late and total reproductive periods. We found that the two traits exhibited natural phenotypic variation with few outliers and a similar reproductive timing pattern as previous reports. The brood size and the male-production rate were not genetically correlated, implying that they might have different genetic architectures and that meiosis quality can be maintained despite increased progeny-production in the late reproductive period caused by more self-sperm. We also identified loci and candidate genetic variants significantly associated with male-production rate in the late and total reproductive periods. Our results provide an insight into life history traits in wild C. elegans strains. The Functional Nanopore Screen: A Versatile High-throughput Assay to Study and Engineer Protein Nanopores in Escherichia coli Nanopores comprise a versatile class of membrane proteins that carry out a range of key physiological functions and are increasingly developed for different biotechnological applications. Yet, a capacity to study and engineer protein nanopores by combinatorial means has so far been hampered by a lack of suitable assays that combine sufficient experimental resolution with throughput. Addressing this technological gap, the Functional Nanopore (FuN) screen now provides a quantitative and dynamic read-out of nanopore assembly and function in E. coli. The assay is based on genetically-encoded fluorescent protein sensors that resolve the nanopore-dependent influx of Ca2+ across the inner membrane of E. coli. Illustrating its versatile capacity, the FuN screen is first applied to dissect the molecular features that underlie the assembly and stability of nanopores formed by the S2168 holin. In a subsequent step, nanopores are engineered by recombining the transmembrane module of S2168 with different ring-shaped oligomeric protein structures that feature defined hexa-, hepta- and octameric geometries. Library screening highlights substantial plasticity in the ability of the S2168 transmembrane module to oligomerize in alternative geometries while the functional properties of the resultant nanopores can be fine-tuned through the identity of the connecting linkers. Overall, the FuN screen is anticipated to facilitate both fundamental studies and complex nanopore engineering endeavors with many potential applications in biomedicine, biotechnology and synthetic biology. 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. Mitochondrial genomes of two parasitic Cuscuta species lack clear evidence of horizontal gene transfer and retain unusually fragmented ccmFC genes BackgroundThe intimate association between parasitic plants and their hosts favours the exchange of genetic material, potentially leading to horizontal gene transfer (HGT) between plants. With the recent publication of several parasitic plant nuclear genomes, there has been considerable focus on such non-sexual exchange of genes. To enhance the picture on HGT events in a widely distributed parasitic genus, Cuscuta (dodders), we assembled and analyzed the organellar genomes of two recently sequenced species, C. australis and C. campestris, making this the first account of complete mitochondrial genomes (mitogenomes) for this genus.

ResultsThe mitogenomes are 265,696 and 275,898 bp in length and contain a typical set of mitochondrial genes, with ten missing or pseudogenized genes often lost from angiosperm mitogenomes. Each mitogenome also possesses a structurally unusual ccmFC gene, which exhibits splitting of one exon and a shift to trans-splicing of its intron. Based on phylogenetic analysis of mitochondrial genes from across angiosperms and similarity-based searches, there is little to no indication of HGT into the Cuscuta mitogenomes. A few candidate regions for plastome-to-mitogenome transfer were identified, with one suggestive of possible HGT.

ConclusionsThe lack of HGT is surprising given examples from the nuclear genomes, and may be due in part to the relatively small size of the Cuscuta mitogenomes, limiting the capacity to integrate foreign sequences. Sensing their plasma membrane curvature allows migrating cells to circumvent obstacles Cell migration is a hallmark out-of-equilibrium process in biology. In addition to persistent self-propelled motion, many cells display remarkable adaptive behaviors when they navigate complex environments within the body. Combining theory and experiments, we identify a curvature-sensing mechanism underlying obstacle avoidance in immune-like cells. The genetic perturbation of this machinery leads to a reduced capacity to evade obstructions combined with faster and more persistent cell migration in obstacle-free environments. We propose that the active polymerization of the actin cytoskeleton at the advancing edge of migrating cells is locally inhibited by the curvature-sensitive BAR protein Snx33 in regions with inward plasma membrane curvature. This coupling between actin and membrane dynamics leads to a mechanochemical instability that generates complex protrusive patterns at the cellular front. Adaptive motility thus arises from two simultaneous curvature-dependent effects, i) the specific reduction of propulsion in regions where external objects deform the plasma membrane and ii) the intrinsic patterning capacity due to the membrane-actin coupling that promotes spontaneous changes in the cells protrusions. Our results show how cells utilize actin- and plasma membrane biophysics to sense their environment, allowing them to adaptively decide if they should move ahead or turn away. On the basis of our findings, we propose that the natural diversity of BAR proteins may allow cells to tune their curvature sensing machinery to match the shape characteristics in their environment. A Eukaryotic Circuit for Secrete-and-Sense Autonomy Cancers represent complex autonomous robust systems, displaying interconnectivity with feedback control. Autonomous growth is fueled by a cancer cells ability to  secrete-and-sense growth factors: a poorly understood phenomenon. Using an integrated systems and experimental approach, here we dissect the impact of a feedback-coupled GTPase circuit within the secretory pathway that imparts secretion-coupled autonomy. The circuit is assembled when the Ras-superfamily monomeric GTPase Arf1, and the heterotrimeric GTPase Giabg and their corresponding GAPs and GEFs are coupled by GIV/Girdin, a protein that is known to fuel aggressive traits in diverse cancers. One forward and two key negative feedback loops within the circuit create closed-loop control (CLC), allow the two GTPases to coregulate each other, and convert the expected switch-like behavior of Arf1-dependent secretion into an unexpected dose response alignment behavior of sensing and secretion. Such behavior translates into cell survival that is self-sustained by stimuli-proportionate secretion. Findings highlight how enhanced coupling of two biological switches in cancer cells is critical for multiscale feedback control to achieve secretion-coupled autonomy.

GRAPHIC ABSTRACT

O_FIG O_LINKSMALLFIG WIDTH=163 HEIGHT=200 SRC="FIGDIR/small/436048v3_ufig1.gif" ALT="Figure 1">
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org.highwire.dtl.DTLVardef@183ff5corg.highwire.dtl.DTLVardef@f21de7org.highwire.dtl.DTLVardef@1893683org.highwire.dtl.DTLVardef@159d966_HPS_FORMAT_FIGEXP  M_FIG C_FIG In BriefThis work delivers an experimentally validated dynamical systems model of the cooperativity between two distinct classes of biological switches in eukaryotic cells and reveals the basis of secretion-coupled cellular autonomy in cancer.

HIGHLIGHTSO_LIModeling and experimental approaches were used to dissect a coupled GTPase circuit
C_LIO_LICoupling enables closed loop feedback and mutual control of GTPases
C_LIO_LICoupling generates dose response alignment behavior of sensing and secretion.
C_LIO_LICoupling is critical for multiscale feedback control to achieve secretion-coupled autonomy.
C_LI Polycystins are upregulated in fibrotic kidneys and promote renal fibrosis Autosomal polycystic kidney disease (ADPKD) is caused by mutations in PKD1or PKD2 gene, which encodes polycystin-1 or polycystin-2 respectively. Paradoxically, polycystins are up-regulated in human ADPKD kidneys and overexpression of polycystin-1 even induced renal cyst formation in mice, suggesting that up-regulation of polycystin is pathogenic. In this study, we determined the role of polycystin in renal fibrosis and explored its underlying mechanisms. We found that the expression of polycystin-1 or polycystin-2 was positively correlated with the expression of collagen-I in the kidney of three different fibrotic mouse models. Pharmaceutical inhibition or genetic knockout of polycystin-2 reduced the expression of pro-fibrotic markers in fibrotic kidneys of two animal models. Moreover, conditional knockout of Pkd1 gene attenuated renal fibrosis in two mouse models. Enhancer of Zeste homolog 2 (EZH2) is a methyltransferase which plays important role in renal fibrosis. We found that EZH2 was positively correlated with the expression of polycystins in fibrotic kidneys. Conditional knockout of the Ezh2 attenuated the anti-fibrotic responses induced by Pkd1 deletion in fibrotic kidneys.

In conclusion, polycystins are up-regulated in injured kidneys and polycystins promote renal fibrosis through up-regulation of EZH2 in fibrotic kidneys, suggesting that renal fibrosis in polycystic kidneys is not the direct effect of PKD gene mutations.

SignificanceMutations of polycystins cause autosomal polycystic kidney disease (ADPKD) which is associated with increased renal fibrosis in cystic kidneys. However, the role of polycystin in renal fibrosis remains unclear. Here, we showed that the expression of polycystins is positively correlated with the development of renal fibrosis in injured kidneys. Pharmaceutical inhibition or genetic deletion of polycystin-2 attenuated renal fibrosis in fibrotic kidneys. Moreover, genetic deletion of polycystin-1 inhibited renal fibrosis in two different animal models. Finally, we showed that ablation of the methyltransferase EZH2 mitigated the anti-fibrotic effect of polycystin-1 deletion in fibrotic kidneys. We conclude that polycystin promotes through EZH2 and renal fibrosis in cystic kidney is not the direct effect of PKD gene mutations. A MALDI-TOF assay identifies nilotinib as an inhibitor of inflammation in acute myeloid leukaemia Inflammatory responses are important in cancer, particularly in the context of monocyte-rich aggressive myeloid neoplasm. We developed a label-free cellular phenotypic drug discovery assay to identify anti-inflammatory drugs in human monocytes derived from acute myeloid leukaemia (AML), by tracking several biological features ionizing from only 2,500 cells using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. A proof-of-concept screen showed that the BCR-ABL inhibitor nilotinib, but not the structurally similar imatinib, blocks inflammatory responses. In order to identify the cellular (off-)targets of nilotinib, we performed thermal proteome profiling (TPP). Unlike imatinib, nilotinib and other later generation BCR-ABL inhibitors inhibit the p38-MK2/3 signalling axis which suppressed the expression of inflammatory cytokines, cell adhesion and innate immunity markers in activated human monocytes derived from AML. Thus, our study provides a tool for the discovery of new anti-inflammatory drugs, which could contribute to the treatment of inflammation in myeloid neoplasms and other diseases.

Key PointsLabel-free cell-based assay identifies new anti-inflammatory drugs using MALDI-TOF MS. Nilotinib reduces inflammation by inhibition of MAPK14-MK2/3 signalling axis in AML. Evolution of thermal physiology alters the projected range of threespine stickleback under climate change Species distribution models (SDMs) are widely used to predict range shifts but could be unreliable under climate change scenarios because they do not account for evolution. The thermal physiology of a species is a key determinant of range and thus incorporating thermal trait evolution into SDMs might be expected to alter projected ranges. We identified a genetic basis for physiological and behavioural traits that evolve in response to temperature change in natural populations of threespine stickleback (Gasterosteus aculeatus). Using these data, we created geographic range projections using a mechanistic niche area approach under two climate change scenarios. Under both scenarios, trait data was either static ( no evolution models), allowed to evolve at observed evolutionary rates ( evolution models), or allowed to evolve at a rate of evolution scaled by the trait variance that is explained by quantitative trait loci (QTL;  scaled evolution models). We show that incorporating these traits and their evolution substantially altered the projected ranges for a widespread panmictic marine population, with over 7-fold increases in area under climate change projections when traits are allowed to evolve. Evolution-informed SDMs should improve the precision of forecasting range dynamics under climate change, and aid in their application to management and the protection of biodiversity. A novel two-generation approach to the population dynamics of gregarious parasitoids Parasitoids, as important natural enemies, help maintain balance in natural ecosystems. Their population dynamics is generally predicted from the number of individuals. Here, using gregarious parasitoids as models, we show that this traditional approach omits one important parameter: the mothers manipulation of offspring fertility due to the clutch size-body size- fertility correlation. As a result of this correlation, when females deliberately adjust the number of offspring laid in a host, they determine not only the number but also the body sizes and reproductive potentials of those offspring. For the first time in the model species Anaphes flavipes, we determined the parasitoids offspring fertility from clutch size. Using this, we experimentally clarified the advantage of specific clutch size combinations and we show that identical fertility in the F1 generation can lead to distinctly different fertility values in the F2 generation. Even with the same number of hosts, lower fertility in the F1 generation can cause higher fertility in the F2 generation. Based on these results, we propose a novel two-generation approach which includes the clutch size-body size-fertility correlation. Our novel approach provides a new perspective for determining the individual fitness levels of gregarious parasitoids with new options for the modelling of parasitoid population dynamics. Large protein complex interfaces have evolved to promote cotranslational assembly Assembly pathways of protein complexes should be precise and efficient to minimise misfolding and unwanted interactions with other proteins in the cell. One way to achieve this efficiency is by seeding assembly pathways during translation via the cotranslational assembly of subunits. While recent evidence suggests that such cotranslational assembly is widespread, little is known about the properties of protein complexes associated with the phenomenon. Here, using a combination of proteome-specific protein complex structures and publicly available ribosome profiling data, we show that cotranslational assembly is particularly common between subunits that form large intermolecular interfaces. To test whether large interfaces have evolved to promote cotranslational assembly, as opposed to cotranslational assembly being a non-adaptive consequence of large interfaces, we compared the sizes of first and last translated interfaces of heteromers in bacterial, yeast, and human complexes. We detect a clear preference in N-terminal interfaces, which will be translated first and thus more likely to form cotranslationally, to be larger than C-terminal interfaces, suggesting that large interfaces have evolved as a means to maximise the chance of successful cotranslational subunit binding. Naming of human diseases on the wrong side of history BackgroundIn the medical sphere, understanding naming conventions strengthen the integrity of naming human diseases remains nominal rather than substantial yet. Since the current nosology-based standard for human diseases could not offer a one-size-fits-all corrective mechanism, many idiomatic but flawed names frequently appear in scientific literature and news outlets at the cost of sociocultural impacts.

ObjectiveWe attempt to examine the ethical oversights of current naming practices and propose heuristic rationales and approaches to determine a pithy name instead of an inopportune nosology.

MethodsFirst, we examined the compiled global online news volumes and emotional tones on some inopportune nosology like German measles, Middle Eastern Respiratory Syndrome, Spanish flu, Hong Kong flu, and Huntingtons disease in the wake of COVID-19. Second, we prototypically scrutinize the lexical dynamics and pathological differentials of German measles and common synonyms by leveraging the capacity of the Google Books Ngram Corpus. Third, we demonstrated the empirical approaches to curate an exclusive substitute for an anachronistic nosology German measles based on deep learning models and post-hoc explanations.

ResultsThe infodemiological study shows that the public informed the offensive names with extremely negative tones in textual and visual narratives. The findings of the historiographical study indicate that many synonyms of German measles did not survive, while German measles became an anachronistic usage, and rubella has taken the dominant place since 1994. The PubMedBERT model could identify rubella as a potential substitution for German measles with the highest semantic similarity. The results of the semantic drift experiments further indicate that rubella tends to survive during the ebb and flow of semantic drift.

ConclusionsOur findings indicate that the nosological evolution of anachronistic names could result in sociocultural impacts without a corrective mechanism. To mitigate such impacts, we introduce some ethical principles for formulating an improved naming scheme. Based on deep learning models and post-hoc explanations, our illustrated experiments could provide hallmark references to the remedial mechanism of naming practices and pertinent credit allocations. Combined Lineage Tracing and scRNA-seq Reveals Unexpected First Heart Field Predominance of Human iPSC Differentiation During mammalian development, the left and right ventricles arise from early populations of cardiac progenitors known as the first and second heart fields, respectively. While these populations have been extensively studied in non-human model systems, their identification and study in vivo human tissues have been limited due to the ethical and technical limitations of accessing gastrulation stage human embryos. Human induced pluripotent stem cells (hiPSCs) present an exciting alternative for modeling early human embryogenesis due to their well-established ability to differentiate into all embryonic germ layers. Here, we describe the development of a TBX5/MYL2 lineage tracing reporter system that allows for the identification of FHF-progenitors and their descendants including left ventricular cardiomyocytes. Furthermore, using single cell RNA sequencing (scRNA-seq) with oligonucleotide-based sample multiplexing, we extensively profiled differentiating hiPSCs across 12 timepoints in two independent iPSC lines. Surprisingly, our reporter system and scRNA-seq analysis revealed a predominance of FHF differentiation using the small molecule Wnt-based 2D differentiation protocol. We compared this data with existing murine and 3D cardiac organoid scRNA-seq data and confirmed the dominance of left ventricular cardiomyocytes (>90%) in our hiPSC-derived progeny. Together, our work provides the scientific community with a powerful new genetic lineage tracing approach as well as a single cell transcriptomic atlas of hiPSCs undergoing cardiac differentiation. Competitive evolution of H1N1 and H3N2 influenza viruses in the United States: A mathematical modeling study Seasonal influenza causes vast public health and economic impact globally. The prevention and control of the annual epidemics remain a challenge due to the antigenic evolution of the viruses. Here, we presented a novel modeling framework based on changes in amino acid sequences and relevant epidemiological data to retrospectively investigate the competitive evolution and transmission of H1N1 and H3N2 influenza viruses in the United States during October 2002 and April 2019. To do so, we estimated the time-varying disease transmission rate from the reported influenza cases and the time-varying antigenic change rate of the viruses from the changes in amino acid sequences. By incorporating the time-varying antigenic change rate into the transmission models, we found that the models could capture the evolutionary transmission dynamics of influenza viruses in the United States. Our modeling results also showed that the antigenic change of the virus plays an essential role in seasonal influenza dynamics. Different contribution of the monkey prefrontal and premotor dorsal cortex in decision making during a transitive inference task Several studies have reported similar neural modulations between brain areas of the frontal cortex, such as the dorsolateral prefrontal (DLPFC) and the premotor dorsal (PMd) cortex, in tasks requiring encoding of the abstract rules for selecting the proper action. Here we compared the neuronal modulation of the DLPFC and PMd of monkeys trained to choose the higher rank from a pair of abstract images (target item), selected from an arbitrarily rank-ordered set (A>B>C>D>E>F) in the context of a transitive inference task. Once acquired by trial-and-error, the ordinal relationship between pairs of adjacent images (i.e., A>B; B>C; C>D; D>E; E>F), monkeys were tested in indicating the ordinal relation between items of the list not paired during learning. During these decisions, we observed that the choice accuracy increased and the reaction time decreased as the rank difference between the compared items enhanced. This result is in line with the hypothesis that after learning, the monkeys built an abstract mental representation of the ranked items, where rank comparisons correspond to the items position comparison on this representation. In both brain areas, we observed higher neuronal activity when the target item appeared in a specific location on the screen with respect to the opposite position and that this difference was particularly enhanced at lower degrees of difficulty. By comparing the time evolution of the activity of the two areas, we observed that the neural encoding of target item spatial position occurred earlier in the DLPFC than in the PMd. Nanocrown electrodes for reliable and robust intracellular recording of cardiomyocytes and cardiotoxicity screening Drug-induced cardiotoxicity arises primarily when a compound alters the electrophysiological properties of cardiomyocytes. Features of intracellular action potentials (iAPs) are powerful biomarkers that predict proarrhythmic risks. However, the conventional patch clamp techniques for measuring iAPs are either laborious and low throughput or not suitable for measuring electrically connected cardiomyocytes. In the last decade, a number of vertical nanoelectrodes have been demonstrated to achieve parallel and minimally-invasive iAP recordings. Nanoelectrodes show great promise, but the large variability in success rate, signal strength, and the low throughput of device fabrication have hindered them from being broadly adopted for proarrhythmia drug assessment. In this work, we developed vertically-aligned and semi-hollow nanocrown electrodes that are mechanically robust and made through a scalable fabrication process. Nanocrown electrodes achieve >99% success rates in obtaining intracellular access through electroporation, allowing reliable and simultaneous iAP recordings from up to 57 human pluripotent stem-cell-derived cardiomyocytes (hPSC-CMs). The accuracy of nanocrown electrode recordings is validated by simultaneous patch clamp recording from the same cell. Nanocrown electrodes enable prolonged iAP recording for continual monitoring of the same cells upon the sequential addition of four to five incremental drug doses. In this way, the dose-response data is self-referencing, which avoids the cell-to-cell variations inherent to hPSC-CMs. We are hopeful that this technology development is a step towards establishing an iAP screening assay for preclinical evaluation of drug-induced arrhythmogenicity. Effect of a low concentration in chloride, sodium and potassium on oocyte maturation, oocyte hydration, ovulation and egg quality in rainbow trout. Water salinity is an important environmental factor known to have detrimental effects on salmonid reproduction, mostly when migrating female broodfish are held in sea water. In contrast, data obtained in freshwater are scarce and the impact of low water salinity, and more specifically of low water concentrations in sodium, chloride and potassium, during reproduction in freshwater is currently unknown. For this reason, and because ion and water fluxes are critical for the final steps of the female gamete formation, including oocyte hydration and ovulation, the aim of the present study was to investigate the impact of low salinity water on final oocyte maturation, ovulation and, ultimately, on egg quality, using rainbow trout as a physiological model and relevant aquaculture species.

Fish from the same commercial strain were raised either in a site characterized by low concentrations of Na+, K+, and Cl- ions in the water or in a closely located control site exhibiting higher concentration in these elements. Egg quality and duration of final oocyte maturation were investigated using innovative phenotyping tools such as automatic assessment of egg viability using the VisEgg system and non-invasive echograph-based monitoring of final oocyte maturation duration, respectively. Oocyte hydration during final oocyte maturation and after ovulation was also investigated. Finally, molecular phenotyping was performed using real-time PCR-based monitoring of several key players of final oocyte maturation and ovulation associated with ion and water transport, inflammation, proteolytic activity, and coagulation. Oocyte hydration and gene expression data were analyzed in the light of the duration of final oocyte maturation.

Here we show that low water salinity (i.e., low water concentration in chloride, sodium and potassium) negatively influences final oocyte maturation, ovulation and, ultimately, egg quality. Low water salinity triggered delayed ovulation and lower oocyte viability. When investigating the impact of low water salinity on final oocyte maturation duration, individuals presenting the most severe phenotypes exhibited impaired oocyte hydration and abnormally reduced gene expression levels of several key players of the ovulatory process. While the under expression of water (i.e., aquaporins) and ion (i.e., solute carriers) transporters is consistent with impaired oocyte hydration, our observations also indicate that the overall ovulatory gene expression program is disrupted. Our results raise the question of the mechanisms underlying the negative influence of low salinity water on the dynamics of the preovulatory phase, on the control of the oocyte homeostasis, including hydration, and on the overall success of the maturation-ovulation process.

HighlightsO_LILow water salinity impairs final oocyte maturation and egg quality in rainbow trout
C_LIO_LILow water salinity induces delayed ovulation and impaired oocyte hydration
C_LIO_LILow water salinity induces a dysregulation of several key ovulatory genes
C_LIO_LIMonitoring of final oocyte maturation can be performed using ultrasound staging
C_LI Pipeline to detect the positional and directional relationship between transposable elements and adjacent genes in host genome AO_SCPLOWBSTRACTC_SCPLOWUnderstanding the relationship between transposable elements (TEs) and their closest positional genes in the host genome is a key point to explore their potential role in genome evolution. Transposable elements can regulate and affect gene expression not only because of their mobility within the genome but also because of their transcriptional activity. A comprehensive knowledge of structural organization between transposable elements and neighboring genes is important to study TE functional role in gene regulation. We implemented a pipeline to display the relationship between the distribution of TEs and adjacent genes in host genomes. Our tool is freely available here: https://github.com/marieBvr/TEs_genes_relationship_pipeline. Egress of human cytomegalovirus through multivesicular bodies Human Cytomegalovirus (HCMV) can infect a variety of cell types by using virions of varying glycoprotein compositions. It is still unclear how this diversity is generated, but spatio-temporally separated envelopment and egress pathways might play a role. So far, one egress pathway has been described in which HCMV particles are individually enveloped into small vesicles and are subsequently exocytosed continuously. However, some studies have also found enveloped virus particles inside multivesicular structures but could not link them to productive egress or degradation pathways.

We used a novel 3D-CLEM workflow allowing us to investigate these structures in HCMV morphogenesis and egress at high spatio-temporal resolution. We found that multiple envelopment events occurred at individual vesicles leading to multiviral bodies (MViBs), which subsequently traversed the cytoplasm to release virions as intermittent bulk pulses at the plasma membrane to form extracellular virus accumulations (EVAs). Our data support the existence of a novel bona fide HCMV egress pathway, which opens the gate to evaluate divergent egress pathways in generating virion diversity. An Immune Cell Atlas Reveals Dynamic COVID-19 Specific Neutrophil Programming Amenable to Dexamethasone Therapy SARS-CoV-2 is a novel coronavirus that causes acute respiratory distress syndrome (ARDS), death and long-term sequelae. Innate immune cells are critical for host defense but are also the primary drivers of ARDS. The relationships between innate cellular responses in ARDS resulting from COVID-19 compared to other causes of ARDS, such as bacterial sepsis is unclear. Moreover, the beneficial effects of dexamethasone therapy during severe COVID-19 remain speculative, but understanding the mechanistic effects could improve evidence-based therapeutic interventions. To interrogate these relationships, we developed an scRNA-Seq and plasma proteomics atlas (biernaskielab.ca/COVID_neutrophil). We discovered that compared to bacterial ARDS, COVID-19 was associated with distinct neutrophil polarization characterized by either interferon (IFN) or prostaglandin (PG) active states. Neutrophils from bacterial ARDS had higher expression of antibacterial molecules such as PLAC8 and CD83. Dexamethasone therapy in COVID patients rapidly altered the IFNactive state, downregulated interferon responsive genes, and activated IL1R2+ve neutrophils. Dexamethasone also induced the emergence of immature neutrophils expressing immunosuppressive molecules ARG1 and ANXA1, which were not present in healthy controls. Moreover, dexamethasone remodeled global cellular interactions by changing neutrophils from information receivers into information providers. Importantly, male patients had higher proportions of IFNactive neutrophils, a greater degree of steroid-induced immature neutrophil expansion, and increased mortality benefit compared to females in the dexamethasone era. Indeed, the highest proportion of IFNactive neutrophils was associated with mortality. These results define neutrophil states unique to COVID-19 when contextualized to other life-threatening infections, thereby enhancing the relevance of our findings at the bedside. Furthermore, the molecular benefits of dexamethasone therapy are also defined, and the identified pathways and plasma proteins can now be targeted to develop improved therapeutics. Discovery of highly potent pancoronavirus fusion inhibitors that also effectively inhibit COVID-19 variants from the UK (Alpha), South Africa (Beta), and India (Delta) We report here the discovery of several highly potent small molecules that showed low nM potency against SARS-CoV (IC50: as low as 13 nM), SARS-CoV-2 (IC50: as low as 23 nM), and MERS-CoV (IC50: as low as 76 nM) in pseudovirus based assays with excellent selectivity indices (SI: as high as > 5000) demonstrating their pancoronavirus inhibition. Some compounds also show 100% inhibition of CPE (IC100) at 1.25 {micro}M against an authentic SARS-CoV-2 (US_WA-1/2020). Furthermore, the most active inhibitors also potently inhibited variants of concerns (VOCs), such as the UK (B.1.1.7), South Africa (B.1.351), and Delta variant (B.1.617.2), originated in India. We confirmed that one of the potent inhibitors binds to the prefusion spike protein trimer of SARS-CoV-2 by SPR. Besides, we showed that they inhibit virus-mediated cell-cell fusion. The ADME data of one of the most active inhibitors, NBCoV1, show drug-like properties. In vivo PK of NBCoV1 in rats demonstrated excellent half-life (t1/2) of 11.3 h, mean resident time (MRT) of 14.2 h, and oral bioavailability. We expect the lead inhibitors to pave the way for further development to preclinical and clinical candidates. A new hypothesis-testing model for phyllochron based on a stochastic process - application to analysis of genetic and environment effects in maize The times between appearance of successive leaves or phyllochron characterize the vegetative development of annual plants. Hypothesis testing models, which enables to compare phyllochron between genetic groups or conditions, are usually based on regression of thermal time on the number of leaves, most of the time assuming a constant leaf appearance rate. However these models are both statistically biased and inappropriate in terms of modelling. We propose a stochastic process model in which the emergence of new leaves is considered as successive time-to-events, which provides a flexible and more accurate modelling as well as unbiased testing procedures. The model was applied on an original maize dataset collected in fields for three years on plants originating from two divergent selection experiments for flowering time conducted in two maize inbred lines. We showed that the main differences in phyllochron were not observed between selection populations (Early or Late), but rather between ancestral lines, years of experimentation, and leaf ranks. Our results highlight a strong departure from the assumption of a constant leaf appearance rate in one year that could be related to climate variations, even if the impact of each climatic variables individually was not clearly elucidated. Etmopterus lantern sharks use coelenterazine as the substrate for their luciferin-luciferase bioluminescence system The lantern shark genus Etmopterus is a group of deep-sea bioluminescent fishes. They emit blue light mainly from the ventral body surface, and the primary biological function is considered to be for camouflage by counterillumination. In this study, we detected both coelenterazine and coelenterazine-specific luciferase activity in the ventral photophore tissues. The results suggested that bioluminescence in lantern sharks is produced using coelenterazine as the substrate for the luciferin-luciferase reaction. P2X2 receptor subunit interfaces are missense variant hotspots where mutations tend to increase apparent ATP affinity Background and PurposeP2X receptors (P2XRs) are trimeric ligand-gated ion channels (LGICs) that open a cation-selective pore in response to ATP binding to their large extracellular domain (ECD). The seven known P2XR subtypes can assemble as homo- or heterotrimeric complexes and contribute to numerous physiological functions, including nociception, inflammation and hearing. The overall structure of P2XRs is well established, but little is known about the spectrum and prevalence of human genetic variations and the functional implications in specific domains.

Experimental ApproachHere we examine the impact of P2X2 receptor (P2X2R) inter-subunit interface missense variants identified in the human population or through structural predictions. We test both single and double mutants through electrophysiological and biochemical approaches.

Key resultsWe demonstrate that predicted ECD inter-subunit interfaces display a higher-than-expected density of missense variations and that the majority of mutations that disrupt putative inter-subunit interactions result in channels with higher apparent ATP affinity. Lastly, we show that double mutants at the subunit interface show significant energetic coupling, especially if located in close proximity.

Conclusions and ImplicationsWe provide the first structural mapping of the mutational burden across the human population in a LGIC and show that the density of missense mutations is constrained between protein domains, indicating evolutionary selection at the domain level. Our data may indicate that, unlike other LGICs, P2X2Rs have evolved an intrinsically high threshold for activation, possibly to allow for additional modulation or as a cellular protection mechanism against overstimulation.

Bullet point summary What is already known: O_LIP2X2 receptors are ATP-activated ion channels implicated in hearing and nociceptice pathways
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 What this study adds: O_LIA structural mapping of missense variants observed in the human population
C_LIO_LIWe identify the intersubunit-interface as a variant hotspot and decipher functional impact of mutations
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 Clinical significance: O_LIThe development of both inhibitors and activators of P2X2 receptor function may be required
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