title: Palmitoylation of PD-L1 Regulates Its Membrane Orientation and Immune Evasion

abstract: Recently identified palmitoylation of PD-L1 is essential for immune regulation. To elucidate the underlying molecular mechanism, we performed giant plasma membrane vesicle (GPMV) experiments, {micro}s-scale all-atom molecular dynamics (MD) simulations and immune killing experiments. GPMV experiments indicated that PD-L1 palmitoylation enhanced its lipid raft affinity. MD simulations revealed dramatically different membrane orientation states of PD-L1 in liquid-ordered (Lo, lipid raft) compared to liquid-disordered (Ld, non-raft) membrane environments. Ld region promoted the "lie-down" orientation of PD-L1, which could inhibit its association with the PD-1 protein on immune cells and thus promote the immune killing of cancer cells. This hypothesis was supported by immune killing experiments using{gamma}{delta} T cells as effector cells and NCI-H1299 lung cancer cells as target cells. In short, our study demonstrates that the palmitoylation affects PD-L1s membrane localization and then membrane orientation, which thus regulates its binding with T cell PD-1 and the immune regulation. These observations may guide therapeutic strategies by explicating the regulation of immune checkpoint proteins by post-translational modifications and membrane environments.


title: Neurexins Regulate GABA Co-release by Dopamine Neurons

abstract:
Midbrain dopamine (DA) neurons are key regulators of basal ganglia functions. The axonal domain of these neurons is highly complex, with a large subset of non-synaptic release sites and a smaller subset of synaptic terminals from which glutamate or GABA are released. The molecular mechanisms regulating the connectivity of DA neurons and their neurochemical identity are unknown. Here we tested the hypothesis that the trans-synaptic cell adhesion molecules neurexins (Nrxns) regulate DA neuron neurotransmission. Conditional deletion of all Nrxns in DA neurons (DAT::Nrxns KO) showed that loss of Nrxns does not impair the basic development and ultrastructural characteristics of DA neuron terminals. However, loss of Nrxns caused an impairment of DA transmission revealed as a reduced rate of DA reuptake following activity-dependent DA release, decreased DA transporter levels, increased vesicular monoamine transporter expression, and impaired amphetamine-induced locomotor activity. Strikingly, electrophysiological recording revealed an increase of GABA co-release from DA neuron axons in the striatum of the KO mice. These findings suggest that Nrxns act as key regulators of DA neuron connectivity and DA-mediated functions.

HighlightsO_LIThe study provides the first direct evidence of the role of neurexins in dopaminergic neurons.
C_LIO_LIThe synaptic adhesion molecules, neurexins, are not required for maintaining the structure of dopamine neuron terminals.
C_LIO_LINeurexins regulate dopaminergic neurotransmission through regulation of dopamine reuptake, impacting amphetamine-induced locomotion.
C_LIO_LIDeletion of Nrxns in DA neurons causes a region-specific increase of GABA release by DA neurons.
C_LI

category: neuroscience


title: Wild cereal grain consumption among Early Holocene foragers of the Balkans predates the arrival of agriculture

abstract: Forager focus on wild cereal plants has been documented in the core zone of domestication in southwestern Asia, while evidence for forager use of wild grass grains remains sporadic elsewhere. In this paper, we present starch grain and phytolith analyses of dental calculus from 61 Mesolithic and Early Neolithic individuals from five sites in the Danube Gorges of the central Balkans. This zone was inhabited by likely complex Holocene foragers for several millennia before the appearance of the first farmers [~]6200 cal BC. We also analyzed forager ground stone tools for evidence of plant processing. Our results based on the study of dental calculus show that certain species of Poaceae (species of the genus Aegilops) were used since the Early Mesolithic, while ground stone tools exhibit traces of a developed grass grain processing technology. The adoption of domesticated plants in this region after [~]6500 cal BC might have been eased by the existing familiarity with wild cereals.

category: ecology


title: How low can you go? Driving down the DNA input requirements for nanopore sequencing

abstract:
The requirement for large amounts of purified DNA limits many sequencing experiments, especially when seeking to avoid pre-amplification or when using third generation technology to sequence molecules directly. We wanted to test the limits of current nanopore sequencing input requirements and devised a set of experiments to evaluate extraction and library preparation approaches for low inputs.

We found an optimised bead beating approach combined with a magnetic bead protocol, rather than traditional spin columns for DNA extraction, improved both molecule length, integrity score and DNA yield. Through reducing the DNA input to as little as 6.25 % of recommended (25 ng versus 400 ng) and reaction volumes in half, library construction can be completed, and sequencing begun within 20 minutes of sample collection.

Applying these approaches, we demonstrated that our pipeline can be used as a cheap and effective method to de novo assemble a genome and identify genes from low quantities and quality of DNA. With our rapid extraction protocol using transportable equipment and low input library construction we were able to generate a de novo assembly from a single insect (Drosophila melanogaster) spanning 125 Mbp / 85 % of the reference genome, over 96.9% complete BUSCO genes, with a contig N50 over 1.2 Mbp, including chromosome arm sized contigs, for a modest consumable cost under {pound}600.

category: genomics


title: Comparison of dipsogenic responses of adult offspring as a function of different perinatal programming models

abstract: The perinatal environment interacts with the genotype of the developing organism resulting in a unique phenotype through a developmental or perinatal programming phenomenon. However, it remains unclear how this phenomenon differentially affects particular targets expressing specific drinking responses depending on the perinatal conditions. The main goal of the present study was to compare the dipsogenic responses induced by different thirst models as a function of two perinatal manipulation models, defined by the maternal free access to hypertonic sodium solution and a partial aortic ligation (PAL-W/Na) or a sham-ligation (Sham-W/Na). The programmed adult offspring of both perinatal manipulated models responded similarly when was challenged by overnight water dehydration or after a sodium depletion showing a reduced water intake in comparison to the non-programmed animals. However, when animals were evaluated after a body sodium overload, only adult Sham-W/Na offspring showed drinking differences compared to PAL and control offspring. By analyzing the central neurobiological substrates involved, a significant increase in the number of Fos + cells was found after sodium depletion in the subfornical organ of both programmed groups and an increase in the number of Fos + cells in the dorsal raphe nucleus was only observed in adult depleted PAL-W/Na. Our results suggest that perinatal programming is a phenomenon that differentially affects particular targets which induce specific dipsogenic responses depending on matching between perinatal programming conditions and the osmotic challenge in the latter environment. Probably, each programmed-drinking phenotype has a particular set point to elicit specific repertoires of mechanisms to reestablish fluid balance.


title: The functional differences between paralogous regulators define the control of the General Stress Response in Sphingopyxis granuli TFA

abstract:
Sphingopyxis granuli TFA is a contaminant degrading alphaproteobacterium that responds to adverse conditions by inducing the General Stress Response (GSR), an adaptive response that controls the transcription of a variety of genes to overcome adverse conditions. The GSR triggered by TFA is driven by two extracytoplasmic function {sigma} factors (ECFs), EcfG1 and EcfG2, whose functional differences have been addressed previously, being EcfG2 the main activator. Upstream in this cascade, NepR anti-{sigma} factors directly inhibit EcfG activity under non-stress conditions, whereas PhyR response regulators sequester the NepR elements upon stress sensing to relieve EcfG inhibition. These elements, which are essential mediators of the GSR regulation, are duplicated in TFA, being NepR1 and NepR2, and PhyR1 and PhyR2. Here, based on multiple genetic, phenotypical and biochemical evidences including in vitro transcription assays, we have assigned distinct functional features to each of these paralogs and assessed their contribution to the GSR regulation, dictating its timing and the intensity. We show that different stress signals are differentially integrated into the GSR by PhyR1 and PhyR2, therefore producing different levels of GSR activation. We demonstrate in vitro that both NepR1 and NepR2 bind EcfG1 and EcfG2, although NepR1 produces a more stable interaction than NepR2. Conversely, NepR2 interacts with phosphorylated PhyR1 and PhyR2 more efficiently than NepR1. We propose an integrative model where NepR2 would play a dual negative role: it would directly inhibit the {sigma} factors upon activation of the GSR and it would modulate the GSR activity indirectly by titrating the PhyR regulators.

IMPORTANCEIn Alphaproteobacteria, the General Stress Response (GSR) aims at protecting against a variety of stresses. Needing to integrate different signals, its modulation is capital to produce a proportionate response according to the environmental conditions. Individual alphaproteobacterial species have evolved distinct GSR cascades in which the information flow is usually straightforward to ascertain due to the presence of a single copy of at least one of its main regulators (PhyR, NepR and EcfG), restricting the regulatory possibilities. However, Sphingopyxis granuli TFA encodes two paralogs of each regulator, multiplying the possible regulatory interplays. We demonstrate that functional differences between paralogous GSR regulators allow an intrinsic feedback regulation in this pathway. We provide evidence of a NepR anti-{sigma} factor that exerts a dual negative feedback regulation on the GSR by interacting with the EcfG {sigma} factors and with the PhyR regulators. This would attune its output to the actual needs of the cell.

category: microbiology


title: A second view on the evolution of flight in stick and leaf insects (Phasmatodea)

abstract: The re-evolution of complex characters is generally considered impossible, yet, studies of recent years have provided several examples of phenotypic reversals shown to violate Dollos law. Along these lines, the regain of wings in stick and leaf insects (Phasmatodea) was hypothesised to have occurred several times independently after an ancestral loss, a scenario controversially discussed among evolutionary biologists. Here, we revisit the recovery of wings by reconstructing a phylogeny based on a comprehensive taxon sample of over 500 representative phasmatodean species to infer the evolutionary history of wings. We additionally explored the presence of ocelli, the photoreceptive organs used for flight stabilisation in winged insects, which might provide further information for interpreting flight evolution. Our findings support an ancestral loss of wings and that the ancestors of most major lineages were wingless. While the evolution of ocelli was estimated to be dependent on the presence of (fully-developed) wings, ocelli are nevertheless absent in the majority of all examined winged species and only appear in the members of few subordinate clades, albeit winged and volant taxa are found in every lineage. The disjunct distribution of ocelli substantiates the hypothesis on trait reacquisition and that wings were regained in Phasmatodea.

category: evolutionary biology


title: Alone Again: Altered Activation in the Observation Execution System during Synchronization in High Loneliness Individuals

abstract: 1AO_SCPLOWBSTRACTC_SCPLOWLonely people evaluate social exchanges negatively and display difficulties in interactions. Interpersonal synchronization is crucial for achieving positive interactions, promoting affinity, closeness, and satisfaction. However, little is known about lonely individuals ability to synchronize and about their brain activity while synchronizing. Following the screening of 303 participants we recruited 32 low and 32 high loneliness participants. They were scanned while engaged in movement synchronization, using a novel dyadic interaction paradigm. Results showed that high loneliness individuals exhibited a reduced ability to adapt their movement to their partners movement. Intriguingly, during movement adaptation periods, high loneliness individuals showed increased activation in the observation-execution (OE) system, specifically in the inferior frontal gyrus and the inferior parietal lobule. They did not show increased activation in the dorsomedial prefrontal cortex, which in the context of synchronization was suggested to be related to gap-monitoring. Based on these findings, we propose a model according to which lonely people may require stronger activation of their OE system for alignment, to compensate for some deficiency in their synchronization ability. Despite this hyper-activation, they still suffer from reduced synchronization capacity. Consequently, synchronization may be a relevant intervention area for the amelioration of chronic loneliness.


title: The role of oxytocin in modulating self-other distinction brain: a pharmacological fMRI study

abstract: The self-other distinction is crucial in human social cognition and social interaction. Studies have found that oxytocin (OT) sharpens the self-other perceptual boundary but with mixed results. Further, little is known if the effect of OT on self-resemblance facial perception exists, especially on its neural basis. Moreover, it is unclear if OT would influence the judgment in self-other discrimination when the other is a child or an adult. In the current double-blinded, placebo-controlled study, we investigated the effect of OT on self-face perception at both behavioral and neural levels. We morphed participants faces and strangers faces to create four stimuli conditions. After being treated by either OT or placebo (PL), participants reported whether a morphed face resembles themselves, or was morphed with their own faces, while being scanned with fMRI. Behavioral results showed that people judged adult-morphed faces better than child-morphed faces. fMRI results showed that the OT group exhibited generally increased activities in the visual area and IFG for self-morphed faces. Such difference was more pronounced in the adult face compared to child face conditions. Multivariate fMRI analysis revealed that the OT group showed better classification between self-morphed versus other-morphed faces, indicating that OT increased self-other distinction, especially for adult faces and in the left hemisphere. Our study shows the significant effect of OT on self-referential brain processes, providing evidence for the potential OTs effect on a left hemisphere self network.


title: Preparatory attention incorporates contextual expectations

abstract:
Humans are remarkably proficient at finding objects within a complex visual world. Current theories of attentional selection propose that this ability is mediated by target-specific preparatory activity in visual cortex, biasing visual processing in favor of the target object. In real-world situations, however, the retinal image that any object will produce is unknown in advance; its size, for instance, varies dramatically with the objects distance from the observer. Using fMRI, we show that preparatory activity is systematically modulated by expectations derived from scene context. Human participants searched for objects at different distances in scenes. Activity patterns in object-selective cortex during search preparation (while no objects were presented), resembled activity patterns evoked by viewing targets object in isolation. Crucially, this preparatory activity was modulated by distance, reflecting the predicted retinal image of the object at each distance. These findings reconcile current theories of attentional selection with the challenges of real-world vision.

HighlightsO_LIVisual cortex contains object-specific representations during search preparation.
C_LIO_LIWe demonstrate this for the first time during concurrent visual scene processing.
C_LIO_LIPreparatory object representations are scaled to account for viewing distance.
C_LIO_LIPreparatory biases reflect the predicted retinal image inferred from scene context.
C_LI

eTOC blurbAttentional selection is thought to be mediated by target-specific preparatory activity in visual cortex. Gayet and Peelen provide evidence that such preparatory biases incorporate contextual expectations about object appearance, reconciling attention theories with the challenges of naturalistic vision.


title: Source-sink connectivity: a novel resting-state EEG marker of the epileptogenic zone

abstract:
Over 15 million epilepsy patients worldwide have drug-resistant epilepsy (DRE). Successful surgery is a standard of care treatment for DRE but can only be achieved through complete resection or disconnection of the epileptogenic zone (EZ), the brain region(s) where seizures originate. Surgical success rates vary between 20-80% because no clinically validated biological markers of the EZ exist. Localizing the EZ is a costly and time-consuming process beginning with non-invasive neuroimaging and often followed by days to weeks of intracranial EEG (iEEG) monitoring. Clinicians visually inspect iEEG data to identify abnormal activity (e.g., low-voltage high frequency activity) on individual channels occurring immediately before seizures or spikes that occur on interictal iEEG (i.e., between seizures). In the end, the clinical standard mainly relies on a small proportion of the iEEG data captured to assist in EZ localization (minutes of seizure data versus days of recordings), missing opportunities to leverage these largely ignored interictal data to better diagnose and treat patients.

Intracranial EEG offers a unique opportunity to observe epileptic cortical network dynamics but waiting for seizures increases patient risks associated with invasive monitoring. In this study, we aim to leverage interictal iEEG data by developing a new network-based interictal iEEG marker of the EZ. We hypothesize that when a patient is not clinically seizing, it is because the EZ is inhibited by other regions. We developed an algorithm that identifies two groups of nodes from the interictal iEEG network: those that are continuously inhibiting a set of neighboring nodes ("sources") and the inhibited nodes themselves ("sinks"). Specifically, patient-specific dynamical network models (DNMs) were estimated from minutes of iEEG and their connectivity properties revealed top sources and sinks in the network, with each node being quantified by source-sink metrics (SSMs). We validated the SSMs in a retrospective analysis of 65 patients by using the SSMs of the annotated EZ to predict surgical outcomes. The SSMs predicted outcomes with an accuracy of 79% compared to an accuracy of 43% for clinicians predictions (surgical success rate of this dataset). In failed outcomes, we identified regions of the brain with high SSMs that were untreated. When compared to high frequency oscillations, the most commonly proposed interictal iEEG feature for EZ localization, SSMs outperformed in predictive power (by a factor of 1.2) suggesting SSMs may be an interictal iEEG fingerprint of the EZ.

category: bioengineering


title: Source-sink connectivity: A novel interictal EEG marker for seizure localization


title: Mitoxantrone modulates a glycosaminoglycan-spike complex to inhibit SARS-CoV-2 infection

abstract: Spike-mediated entry of SARS-CoV-2 into human airway epithelial cells is an attractive therapeutic target for COVID-19. In addition to protein receptors, the SARS-CoV-2 spike (S) protein also interacts with heparan sulfate, a negatively charged glycosaminoglycan (GAG) attached to certain membrane proteins on the cell surface. This interaction facilitates the engagement of spike with a downstream receptor to promote viral entry. Here, we show that Mitoxantrone, an FDA-approved topoisomerase inhibitor, targets a spike-GAG complex to compromise the fusogenic function of spike in viral entry. As a single agent, Mitoxantrone inhibits the infection of an authentic SARS-CoV-2 strain in a cell-based model and in human lung EpiAirway 3D tissues. Gene expression profiling supports the plasma membrane as a major target of Mitoxantrone but also underscores an undesired activity targeting nucleosome dynamics. We propose that Mitoxantrone analogs bearing similar GAG-binding activities but with reduced affinity for DNA topoisomerase may offer an alternative therapy to overcome breakthrough infections in the post-vaccine era.

category: cell biology


title: SIMBA: SIngle-cell eMBedding Along with features

abstract: Recent advances in single-cell omics technologies enable the individual and joint profiling of cellular measurements. Currently, most single-cell analysis pipelines are cluster-centric and cannot explicitly model the interactions between different feature types. In addition, single-cell methods are generally designed for a particular task as distinct single-cell problems are formulated differently. To address these current shortcomings, we present SIMBA, a graph embedding method that jointly embeds single cells and their defining features, such as genes, chromatin accessible regions, and transcription factor binding sequences into a common latent space. By leveraging the co-embedding of cells and features, SIMBA allows for the study of cellular heterogeneity, clustering-free marker discovery, gene regulation inference, batch effect removal, and omics data integration. SIMBA has been extensively applied to scRNA-seq, scATAC-seq, and dual-omics data. We show that SIMBA provides a single framework that allows diverse single-cell analysis problems to be formulated in a unified way and thus simplifies the development of new analyses and integration of other single-cell modalities. SIMBA is implemented as an efficient, comprehensive, and extensible Python library (https://simba-bio.readthedocs.io) for the analysis of single-cell omics data using graph embedding.

category: bioinformatics


title: Improving MHC Class I antigen processing predictions using representation learning and cleavage site-specific kernels

abstract: In this work, we propose a new deep learning model, MHCrank, to predict the probability that a peptide will be processed for presentation within the MHC Class I pathway. We find that the performance of our model is significantly higher than two previously published baseline methods: MHCflurry and netMHCpan. Gains in performance result from the utilization of cleavage site-specific kernels and learned representations for amino acids. By visualizing the site-specific amino acid enrichment among top-ranked peptides, we find MHCranks top-ranked peptides are enriched at biologically relevant positions with amino acids that are consistent with previous work. Furthermore, the cosine similarity matrix derived from MHCranks learned embeddings for amino acids correlate highly with physiochemical properties that have been experimentally shown to be important in determining a peptides favorability to be processed. Altogether, the results reported in this work indicate that the proposed MHCrank demonstrates strong performance compared to existing methods and could have vast applicability to aid drug and vaccine development.


title: RTX-KG2: a system for building a semantically standardized knowledge graph for translational biomedicine

abstract:
BackgroundBiomedical translational science is increasingly leveraging computational reasoning on large repositories of structured knowledge (such as the Unified Medical Language System (UMLS), the Semantic Medline Database (SemMedDB), ChEMBL, DrugBank, and the Small Molecule Pathway Database (SMPDB)) and data in order to facilitate discovery of new therapeutic targets and modalities. Since 2016, the NCATS Biomedical Data Translator project has been working to federate autonomous reasoning agents and knowledge providers within a distributed system for answering translational questions. Within that project and within the field more broadly, there is an urgent need for an open-source framework that can efficiently and reproducibly build an integrated, standards-compliant, and comprehensive biomedical knowledge graph that can be either downloaded in standard serialized form or queried via a public application programming interface (API) that accords with the FAIR data principles.

ResultsTo create a knowledge provider system within the Translator project, we have developed RTX-KG2, an open-source software system for building--and hosting a web API for querying--a biomedical knowledge graph that uses an Extract-Transform-Load (ETL) approach to integrate 70 knowledge sources (including the aforementioned sources) into a single knowledge graph. The semantic layer and schema for RTX-KG2 follow the standard Biolink metamodel to maximize interoperability within Translator. RTX-KG2 is currently being used by multiple Translator reasoning agents, both in its downloadable form and via its SmartAPI-registered web interface. Serializations of RTX-KG2 are available for download in both the pre-canonicalized form and in canonicalized form (in which synonym concepts are merged). The current canonicalized version (KG2.7.3) of RTX-KG2 contains 6.4M concept nodes and 39.3M relationship edges with a rich set of 77 relationship types.

ConclusionRTX-KG2 is the first open-source knowledge graph of which we are aware that integrates UMLS, SemMedDB, ChEMBL, DrugBank, SMPDB, and 65 additional knowledge sources within a knowledge graph that conforms to the Biolink standard for its semantic layer and schema at the intersections of these databases. RTX-KG2 is publicly available for querying via its API at arax.rtx.ai/api/rtxkg2/v1.2/openapi.json. The code to build RTX-KG2 is publicly available at github:RTXteam/RTX-KG2.


title: Maintenance of ankle power and decreased inter-limb asymmetry during sprint running in fatigued multi-speed athletes

abstract:
The ability to shift from walking and jogging to sprinting gaits, even when fatigued after prolonged effort, would have been as useful to our hunter-gatherer ancestors as it is to modern athletes. During prolonged jogging, joint moment and work decrease in the distal (ankle) joint but increase at proximal (hip/knee) joints as fatigue progresses, and such adaptations might be expected to occur in sprinting. Fatigue is also thought to increase inter-limb kinematic and force production asymmetries, which are speculated to influence injury risk. However, the effects of running-related fatigue on sprint running gait have been incompletely studied, so these hypotheses remain untested. We studied 3-D kinematics and ground reaction force production in dominant (DL) and non-dominant (NDL) legs during both non-fatigued and fatigued sprinting in habitual but uncoached running athletes. Contrary to the tested hypotheses, relative between-leg differences were greater in non-fatigued than fatigued sprinting. When not fatigued, DL produced greater propulsive impulse through both greater positive and negative work being done at the ankle, whilst NDL produced more vertical impulse, possibly resulting from the greater hip flexion observed prior to the downwards acceleration of the foot towards the ground. Whilst few changes were detected in DL once fatigued, NDL shifted towards greater horizontal force production, largely resulting from an increase in plantarflexion (distal-joint) moments and power. After fatiguing running, therefore, inter-limb asymmetry was reduced during sprinting and no distal-to-proximal shift in work/power was detected. Speculatively, these adaptations may help to attenuate decreases in running speed whilst minimising injury risk.

SignificanceThe ability to attain fast running speeds may critically determine success in tasks such as prey chase- and-capture in hunter-gatherer societies as well as success in modern sports competitions. At times, sprint running may have to be performed whilst fatigued from previous, longer-distance running, when speeds are reduced, and injury risk may be higher. Previous work indicated that fatigue prompts a proximal shift in joint work and power production and an increase in inter-limb asymmetry. On the contrary, we show that relative ankle positive and negative joint work was maintained in the face of fatigue and that inter-limb asymmetry was reduced in a group of runners experienced, but not formally instructed, in both long-distance and sprint running.

category: biophysics


title: Fatigue does not increase limb asymmetry or induce proximal joint power shift during sprinting in habitual, multi-speed runners


title: Elevated islet prohormone ratios as indicators of insulin dependency in islet transplant recipients

abstract: Autologous pancreatic islet transplantation is an established therapy for patients with chronic pancreatitis. However, the long-term transplant outcomes are modest. Identifying indicators of graft function will aid the preservation of transplanted islets and glycemic control. To this end, we analyzed beta cell prohormone peptide levels in a retrospective cohort of total pancreatectomy autologous islet transplant patients (n=28). Proinsulin-to-C-peptide (PI/C) and proIAPP-to-total IAPP (proIAPP/IAPP) ratios measured at 3 months post-transplant were significantly higher in patients who remained insulin dependent at 1 year follow-up. In a mouse model of human islet transplantation, recipient mice that later became hyperglycemic displayed significantly higher PI/C ratios than mice that remained normoglycemic. Histological analysis of islet grafts showed reduced insulin- and proinsulin-positive area, but elevated glucagon-positive area in grafts that experienced greater secretory demand. Increased prohormone convertase 1/3 was detected in glucagon-positive cells, and glucagon-like peptide 1 (GLP-1) area was elevated in grafts from mice that displayed hyperglycemia or elevated plasma PI/C ratios, demonstrating intra-islet incretin production in metabolically challenged human islet grafts. These data indicate that in failing grafts, alpha cell prohormone processing is likely altered, and incomplete beta cell prohormone processing may be an early indicator of insulin dependency.

category: pathology


title: HEAT SHOCK FACTOR BINDING PROTEIN limits meiotic crossovers by repressing HEI10 transcription

abstract: The number of meiotic crossovers is tightly controlled and most depend on pro-crossover ZMM proteins, such as the E3 ligase HEI10. Despite the importance of HEI10 dosage for crossover formation, how HEI10 transcription is controlled remains unexplored. In a forward genetic screen using a sensitive fluorescent seed crossover reporter in Arabidopsis thaliana we identify heat shock factor binding protein (HSBP) as a repressor of HEI10 transcription and crossover numbers. Using genome-wide crossover mapping and cytogenetics, we show that hsbp mutations or meiotic HSBP knockdowns increase ZMM-dependent crossovers towards the telomeres, mirroring the effects of HEI10 overexpression. Through RNA sequencing, DNA methylome and chromatin immunoprecipitation analysis, we reveal that HSBP directly represses HEI10 transcription by binding with heat shock factors (HSFs) at the HEI10 promoter and maintaining DNA methylation over the HEI10 5' untranslated region. Our findings provide insights into how the temperature response regulator HSBP restricts meiotic HEI10 transcription and crossover number by attenuating HSF activity.

category: plant biology


title: A common neural currency account for social and non-social decisions

abstract: To date, social and non-social decisions have been studied in isolation. Consequently, the extent to which social and non-social forms of decision uncertainty are integrated using shared neurocomputational resources remains elusive. Here, we address this question using simultaneous EEG-fMRI and a task in which decision evidence in social and non-social contexts varies along comparable scales. First, we identify comparable time-resolved build-up of activity in the EEG, akin to a process of evidence accumulation. We then use the endogenous trial-by-trial variability in the slopes of these accumulating signals to construct parametric fMRI predictors. We show that a region of the posterior-medial frontal cortex (pMFC) uniquely explains trial-wise variability in the process of evidence accumulation in both the social and non-social contexts. We further demonstrate a task-dependent coupling between the pMFC and regions of the human valuation system in dorso- and ventro-medial prefrontal cortex (dmPFC/vmPFC) across both contexts. Finally, we report domain-specific representations in regions known to encode the early decision evidence for each context. These results are suggestive of a domain-general decision-making architecture, whereupon domain-specific information is likely converted into a "common currency" in the dmPFC/vmPFC and accumulated for the decision in the pMFC.


title: Neural dynamics of retrieval suppression in abolishing item-specific cortical pattern of unwanted emotional memories

abstract: When reminded of an unpleasant experience, people often try to exclude the unwanted memory from awareness, a process known as retrieval suppression. Despite the importance of this form of mental control to mental health, the ability to track, in real time, individual memories as they are suppressed remains elusive. Here we used multivariate decoding on EEG data to track how suppression unfolds in time and to reveal its impact on cortical patterns related to individual memories. We presented reminders to aversive scenes and asked people to either suppress or to retrieve the scene. During suppression, mid-frontal theta power within the first 500 ms distinguished suppression from passive viewing of the reminder, indicating that suppression rapidly recruited control. During retrieval, we could discern EEG cortical patterns relating to individual memories-initially, based on theta-driven, visual perception of the reminders (0-500 ms) and later, based on alpha-driven, reinstatement of the aversive scene (500-3000 ms). Critically, suppressing retrieval weakened (during 420-600 ms) and eventually abolished item-specific cortical patterns, a robust effect that persisted until the reminder disappeared (1200-3000 ms). Actively suppressing item-specific cortical patterns, both during an early (300-680 ms) window and during sustained control, predicted later episodic forgetting. Thus, both rapid and sustained control contribute to abolishing cortical patterns of individual memories, limiting awareness, and precipitating later forgetting. These findings reveal how suppression of individual memories from awareness unfolds in time, presenting a precise chronometry of this process.


title: Observing the suppression of individual aversive memories from conscious awareness


title: Extrasynaptic NMDA receptors bidirectionally modulate intrinsic excitability of inhibitory neurons

abstract: The NMDA subtype glutamate receptors (NMDARs) play important roles in both physiological and pathological processes in the brain. Comparing to their critical roles in synaptic modifications and excitotoxicity in the excitatory neurons, much less is understood about the functional contributions of NMDARs to the inhibitory/GABAergic neurons. By using selective NMDAR inhibitors and potentiators, we here show that NMDARs bi-directionally modulate the intrinsic excitability (defined as spontaneous/evoked spiking activity and EPSP-spike coupling) in the inhibitory/GABAergic neurons. This modulation depends on GluN2C/2D-but not GluN2A/2B-containing NMDARs. We further show that NMDAR modulator EU1794-4 mostly enhances extrasynaptic NMDAR activity, and by using it we demonstrate a significant contribution of extrasynaptic NMDARs to the modulation of intrinsic excitability in the inhibitory neurons. Altogether, this bidirectional modulation of intrinsic excitability reveals a previously less appreciated importance of NMDARs in the second-to-second functioning of inhibitory/GABAergic neurons.


title: VISTA as a ligand downregulates LPS-mediated inflammation in macrophages and neutrophils

abstract: V-domain immunoglobulin suppressor of T-cell activation (VISTA) has emerged as a unique immunoregulatory receptor on cells of the myeloid lineage. Agonizing VISTA on myeloid cells has recently been demonstrated to have a profound effect on dampening inflammatory responses. VISTA has been proposed to function both as a ligand and as a receptor. In this context, the role of VISTA as a ligand has been largely ignored. Using a high-avidity agonist of the VISTA receptor (VISTA-COMP), we investigated the effect of exogenous VISTA, as a ligand, on macrophages and neutrophil cellular pathways in an acute inflammatory setting. RNA sequencing analysis demonstrated that VISTA-COMP downregulates pro-inflammatory cytokines and chemokines and upregulates immunoregulatory genes in both LPS-stimulated macrophages and neutrophils ex vivo. Interestingly, unlike VISTA itself, the receptor is only expressed following LPS stimulation of these cell populations. Furthermore, the administration of VISTA-COMP attenuated the rise in circulating TNF levels in LPS-treated mice in vivo. These results suggest that VISTA serves a redundant role on macrophages and neutrophils acting as both a ligand and a receptor in the context of an acute inflammatory event.

category: immunology


title: Boosting of SARS-CoV-2 immunity in nonhuman primates using an oral rhabdoviral vaccine

abstract: An orally active vaccine capable of boosting SARS-CoV-2 immune responses in previously infected or vaccinated individuals would help efforts to achieve and sustain herd immunity. Unlike mRNA-loaded lipid nanoparticles and recombinant replication-defective adenoviruses, replicating vesicular stomatitis viruses with SARS-CoV-2 spike glycoproteins (VSV-SARS2) were poorly immunogenic after intramuscular administration in clinical trials. Here, by G protein trans-complementation, we generated VSV-SARS2(+G) virions with expanded target cell tropism. Compared to parental VSV-SARS2, G-supplemented viruses were orally active in virus-naive and vaccine-primed cynomolgus macaques, powerfully boosting SARS-CoV-2 neutralizing antibody titers. Clinical testing of this oral VSV-SARS2(+G) vaccine is planned.


title: Aerosolization of Mycobacterium tuberculosis by tidal breathing

abstract:
RationaleInterrupting tuberculosis (TB) transmission requires an improved understanding of how - and when - the causative organism, Mycobacterium tuberculosis (Mtb), is aerosolized. Although Cough is commonly assumed to be the dominant source of Mtb aerosols, recent evidence of Cough-independent Mtb release implies the contribution of alternative mechanisms.

ObjectiveTo compare the aerosolization of Mtb and particulate matter from GeneXpert-positive patients during three separate respiratory manoeuvres: Tidal Breathing (TiBr), Forced Vital Capacity (FVC), and Cough.

MethodologyBioaerosol sampling and Mtb detection were combined with real-time assessments of CO2 production and particle counts from 39 confirmed TB patients.

Measurements and Main ResultsTiBr and FVC produced comparable numbers of particles, with Cough producing >4-fold more. For all manoeuvres, the proportions of particles detected across size categories from 0.5 - 5 m were similar, with minor differences observed only in particles between 1.5 - 2 m (p = 0.014) and >5 m (p = 0.020). Viable Mtb bacilli were detected in 66%, 70%, and 65% of TiBr, FVC, and Cough samples, respectively. Notably, while Cough produced 3-fold more Mtb than TiBr, the relative infrequency of coughing compared to breathing implies that TiBr likely contributes >90% of the daily aerosolised Mtb across a range of Cough frequencies.

ConclusionsOur results suggest that, while Cough increases particle aerosolization compared to TiBr, this is not associated with increased Mtb aerosolization. Instead, TiBr produces more Mtb per particle than Cough. Assuming the number of viable Mtb organisms detected provides a proxy measure of patient infectiousness, these observations imply a significant contribution of TiBr to TB transmission.


title: A fluorescence-based viability assay for Phytophthora agathidicida oospores

abstract: Viability staining is an essential tool in many fields of microbiology. In this study, we aimed to establish a dual fluorescence method for detecting the viability of Phytophthora oospores. Phytophthora is a genus of plant pathogens in the class Oomycete that cause disease in a wide range of agriculturally and ecologically important plants. During the Phytophthora disease cycle, thick-walled oospores can be produced via sexual reproduction. These oospores are essential for long-range dispersal and long-term survival of the pathogen, and therefore methods for the study of oospores are of great interest. In this study, we tested five fluorescent dyes for their ability to stain Phytophthora agathidicida oospores: SYTO 9, FUN-1, fluorescein diacetate, propidium iodide, and TOTO-3 iodide. These dyes represented a selection of total, viable and non-viable cell stains Each dye was assessed individually, and then the best dyes were combined for dual viability staining. In this study, we have identified three dyes - SYTO 9, fluorescein diacetate and TOTO-3 - that can be used to report total, viable and non-viable oospores respectively. We further demonstrate that fluorescein diacetate and TOTO-3 can be used together for dual viability staining. This new method is quantitative and compatible with automated image analysis, allowing oospores to be rapidly and accurately assessed for viability.


title: Adaptation of peristaltic pumps for laminar flow experiments

abstract: Endothelial cells (ECs) are the primary cellular constituent of blood vessels that are in direct contact with hemodynamic forces over the course of a lifetime. Throughout the body, vessels experience different types of blood flow patterns and rates that alter vascular architecture and cellular behavior. Because of the complexities of studying blood flow in an intact organism, particularly during development, modeling of blood flow in vitro has become a powerful technique for studying hemodynamic dependent signaling mechanisms in ECs. While commercial flow systems that recirculate fluids exist, many commercially available pumps are peristaltic and best model pulsatile flow conditions. However, there are many important in vivo situations in which ECs experience laminar flow conditions, such as along long, straight stretches of the vasculature. To understand EC function under these situations, it is important to be able to consistently model laminar flow conditions in vitro. Here, we outline a method to reliably adapt commercially available peristaltic pumps to reproducibly study laminar flow conditions. Our proof of concept study focuses on 2-dimensional (2D) models but could be further adapted to 3-dimensional (3D) environments to better model in vivo scenarios such as organ development. Our studies make significant inroads into solving technical challenges associated with flow modeling, and allow us to conduct functional studies towards understanding the mechanistic role of flow forces on vascular architecture, cellular behavior, and remodeling during a variety of physiological contexts.


title: Evaluation role of miR-124 in neurodegenerative diseases: literature review and in silico analysis

abstract: Neurodegenerative diseases (ND) are characterized by loss of function and structure of neurons. NDs like Alzheimers disease (AD) and Parkinsons disease (PD) have high burden on the society and patients. Currently microRNAs (miRNAs) approach is growing. miRNAs express in different tissues, especially in the central neuron systems (CNS). miRNAs have a dynamic role in the CNS among this miRNAs, miR-124 significantly express in the CNS. Studies on miR-124 have shown that miR-124 improves ND. In this study, we evaluated the role of miR-124 in the ND by literature review and in silico analysis. We used Pubmed database to find miR-124 function in the Alzheimers disease, Parkinsons disease, Multiple sclerosis, Huntingtons disease and amyotrophic lateral sclerosis. To better understand the role of miR-124 in the neurons, RNA-seq data form miR-124-deleted neuronal cells extracted from GEO database and analyzed in Galaxy platform. According literature review miR-124 attenuates inflammation and apoptosis in the ND by target NF-kb signaling pathway and regulation of BAX/BCL-2. miR-124 targets BACE1 and decreases level of A{beta}. RNA-seq data showed miR-124 downregulation, an increase in chemokine gene like CCL1 and cytokine-cytokine receptor-interaction, as well as MAPK-signaling pathway. Our study shows that miR-124 can be promising therapeutic approaches to ND.

category: genetics


title: Physarum polycephalum: Establishing an Assay for Testing Decision-making Under Shifting Somatic Boundaries

abstract: Prior studies of decision-making generally assume a fixed agent which maximizes utility among its various options. Physarum polycephalum is a popular model for basal cognition that can be cut into pieces that may or may not re-join. We exploited this capacity to develop a novel assay in which radical changes to the agent itself are among the options of the decision-making process. Specifically, we transected a Physarum culture in the presence of a food reward that was located closer to the new smaller piece. In this scenario, the newly created branch must choose between exploiting the reward itself, or first re-connecting with the original mass (and sharing the nutrient reward across a large body). We report a pilot study establishing a protocol in which the number of agents is part of the decision-making process. We observed that despite the presence of food, new branches strongly prefer to merge back to the syncytium before exploiting the reward. Many improvements to the protocol are possible, to extend this effort to understand the interplay between behavioral options and the structure and boundary of the individual making choices in its environment.

category: animal behavior and cognition


title: Calling the Amino Acid Sequence of a Protein/Peptide from the Nanospectrum Produced by a Sub-nanometer Diameter Pore

abstract: The blockade current that develops when a protein translocates across a thin membrane through a sub-nanometer diameter pore (i.e., a nanospectrum) informs with extreme sensitivity on the sequence of amino acids that constitute the protein. Whereas mass spectrometry (MS) is still the dominant technology for protein identification, it suffers limitations. In proteome-wide studies, MS fails to sequence proteins de novo, but merely classifies a protein and it is not very sensitive requiring about a femtomole to do that. Compared with MS, a sub-nanometer diameter pore (i.e. a sub-nanopore) directly reads the amino acids constituting a single protein molecule, but efficient computational tools are still required for processing and interpreting the blockade current. Here, we delineate computational methods for processing sub-nanopore nanospectra and predicting electrical blockade currents from protein sequences, which are essential for protein identification.


title: Nanoscopic resolution within a single imaging frame

abstract: Mean-Shift Super Resolution (MSSR) is a principle based on the Mean Shift theory that improves the spatial resolution in fluorescence images beyond the diffraction limit. MSSR works on low- and high-density fluorophore images, is not limited by the architecture of the detector (EM-CCD, sCMOS, or photomultiplier-based laser scanning systems) and is applicable to single images as well as temporal series. The theoretical limit of spatial resolution, based on optimized real-world imaging conditions and analysis of temporal image series, has been measured to be 40 nm. Furthermore, MSSR has denoising capabilities that outperform other analytical super resolution image approaches. Altogether, MSSR is a powerful, flexible, and generic tool for multidimensional and live cell imaging applications.


title: Transfer of Knowledge from Model Organisms to Evolutionarily Distant Non-Model Organisms: The Coral Pocillopora damicornis Membrane Signaling Receptome

abstract: With the ease of gene sequencing and the technology available to study and manipulate non-model organisms, the need to translate our understanding of model organisms to non-model organisms has become an urgent problem. For example, mining of large coral and their symbiont sequence data is a challenge, but also provides an opportunity for understanding functionality and evolution of these and other non-model organisms. Much more information than for any other eukaryotic species is available for humans, especially related to signal transduction and diseases. However, the coral cnidarian host and human have diverged over 700 million years ago and homologies between proteins are therefore often in the gray zone or undetectable with traditional BLAST searches. We introduce a two-stage approach to identifying putative coral homologues of human proteins. First, through remote homology detection using Hidden Markov Models, we identify candidate human homologues in the cnidarian genome. However, for many proteins, the human genome alone contains multiple family members with similar or even more divergence in sequence. In the second stage, therefore, we filter the remote homology results based on the functional and structural plausibility of each coral candidate, shortlisting the coral proteins likely to be true human homologues. We demonstrate our approach with a pipeline for mapping membrane receptors in humans to membrane receptors in corals, with specific focus on the stony coral, P. damicornis. More than 1000 human membrane receptors mapped to 335 coral receptors, including 151 G protein coupled receptors (GPCRs). To validate specific sub-families, we chose opsin proteins, representative GPCRs that confer light sensitivity, and Toll-like receptors, representative non-GPCRs, which function in the immune response, and their ability to communicate with microorganisms. Through detailed structure-function analysis of their ligand-binding pockets and downstream signaling cascades, we selected those candidate remote homologues likely to carry out related functions in the corals. This pipeline may prove generally useful for other non-model organisms, such as to support the growing field of synthetic biology.


title: Identification of age-associated proteins and functional alterations in human primary retinal pigment epithelium cells

abstract:
BackgroundRetinal pigmented epithelium (RPE) has essential functions to nourish and support the neural retina, and is of vital importance in the pathogenesis of age-related retinal degeneration. However, the exact molecular changes of RPE in aging remain poorly defined.

MethodsWe isolated human primary RPE (hRPE) cells from 18 eye donors distributed over a wide age range (10 - 67 years). A quantitative proteomic analysis was performed to analyze their intracellular and secreted protein changes, and potential age-associtated mechanisms were validated by ARPE-19 and hRPE cells.

ResultsAge-stage related subtypes and age-associtated proteins and functional alterations were revealed. Proteomic data and verifications showed that RNF123 and RNF149 related ubiquitin-mediated proteolysis might be an important clearance mechanism in elimination of oxidative damaged proteins in aged hRPE. In older hRPE cells, apoptotic signaling related pathways were up-regulated and endoplasmic reticulum organization was down-regulated both in intracellular and secreted proteome.

ConclusionsThis work paints a detailed molecular picture of human RPE in aging process and provides new insights for molecular characteristics of RPE in aging and related clinical retinal conditions.


title: Active reconfiguration of cytoplasmic lipid droplets governs migration of nutrient-limited phytoplankton

abstract:
As open oceans continue to warm, modified currents and enhanced stratification exacerbate nitrogen and phosphorus limitation, constraining primary production. The ability to migrate vertically bestows motile phytoplankton a crucial-albeit energetically expensive-advantage toward vertically redistributing for optimal growth, uptake and resource storage in nutrient-limited water columns. However, this traditional view discounts the possibility that the phytoplankton migration strategy may be actively selected by the storage dynamics when nutrients turn limiting. Here we report that storage and migration in phytoplankton are coupled traits, whereby motile species harness energy storing lipid droplets (LDs) to biomechanically regulate migration in nutrient limited settings. LDs grow and translocate-directionally-within the cytoplasm to accumulate below the cell nucleus, tuning the speed, trajectory and stability of swimming cells. Nutrient reincorporation reverses the LD translocation, restoring the homeostatic migratory traits measured in population-scale millifluidic experiments. Combining intracellular LD tracking and quantitative morphological analysis of red-tide forming alga, Heterosigma akashiwo, along with a model of cell mechanics, we discover that the size and spatial localization of growing LDs govern the ballisticity and orientational stability of migration. The strain-specific shifts in migration which we identify here are amenable to a selective emergence of mixotrophy in nutrient-limited phytoplankton. We rationalize these distinct behavioral acclimatization in an ecological context, relying on concomitant tracking of the photophysiology and reactive oxygen species (ROS) levels, and propose a dissipative energy budget for motile phytoplankton alleviating nutrient limitation. The emergent resource acquisition strategies, enabled by distinct strain-specific migratory acclimatizing mechanisms, highlight the active role of the reconfigurable cytoplasmic LDs in guiding vertical movement. By uncovering the mechanistic coupling between dynamics of intracellular changes to physiologically-governed migration strategies, this work offers a tractable framework to delineate diverse strategies which phytoplankton may harness to maximize fitness and resource pool in nutrient-limited open oceans of the future.

One sentence summaryPhytoplankton harness reconfigurable lipid droplets to biomechanically tune migratory strategies in dynamic nutrient landscapes.


title: A mathematical model for electrical activity in pig atrial tissue

abstract:
Atrial fibrillation (AF) is the most common sustained form of cardiac arrhythmia occurring in humans. Its effective treatment requires a detailed understanding of the underlying mechanisms at the genetic, molecular, cellular, tissue and organ levels. To study the complex mechanisms underlying the development, maintenance and termination of cardiac arrhythmias, we need preclinical research models. These models range from in vitro cell cultures to in vivo small and large animal hearts. However, translational research requires that the results of these animal experiments are understood in the context of human subjects. Currently, this is achieved through simulations with state-of-the-art mathematical models for human and animal heart tissue. In the context of AF, a model that is extensively used by experimentalists, is that of the pig atria. However, until now, an ionically detailed mathematical model for pig atrial tissue has been lacking, and researchers have been forced to rely on mathematical models from other animal species to understand their experimental observations.

In this paper, we present the first ionically detailed mathematical model of porcine atrial electrophysiology. To build the model, we first fitted experimental patch-clamp data from literature to describe the individual currents flowing across the cell membrane. Later, we fine-tuned the model by fitting action potential duration restitution (APDR) curves for different repolarisation levels. The experimental data for the APDR studies was produced in N. Voigts lab.

We extended our model to the tissue level and demonstrated the ability to maintain stable spiral waves. In agreement with previous experimental results, our model shows that early repolarisation is primarily driven by a calcium-mediated chloride current, IClCa, which is completely inactivated at high pacing frequencies. This is a condition found only in porcine atria. The model shows spatiotemporal chaos with reduced repolarisation.

Author summaryState-of-the-art mathematical models of cardiac electrophysiology play an important role in bridging the gap between animal research conducted in the laboratory and preclinical research being considered for translation into the clinic. Using computer simulations, these models enable detailed studies of the behaviour of ion channels and ion transfer at the cellular level, the propagation of electrical waves at the tissue level and the visualisation of the excitation pattern within the heart wall at the organ level. Thus, they contribute to a better understanding of the mechanisms underlying cardiac arrhythmias.

Here, we present the first ionically detailed mathematical model for porcine atrial electrophysiology. The individual membrane currents were modelled by fitting experimental data obtained from literature. The overall electrical response of the tissue was adjusted by fitting action potential duration restitution (APDR) curves obtained from in-house patch-clamp measurements. Our model accounts for an early repolarisation phase of the AP that is primarily Ca2+-dependent, a feature that is consistent with experiments and is identified to be unique to pigs. In extended media, our model is capable of sustaining stable spiral waves, and spatiotemporal chaos, when the repolarisation reserve is reduced.


title: Nuclear PHGDH promotes neutrophil recruitment to drive liver cancer progression

abstract: Metabolic dysregulation and the communications between cancer and immune cells are emerging as two essential features of malignant tumors. In this study, we observed that nuclear localization of phosphoglycerate dehydrogenase (PHGDH) associates with poor prognosis of liver cancer patients, and Phgdh is required for liver cancer progression in a mouse model. Unexpectedly, the impairment of Phgdh enzyme activity exerts a slight effect on liver cancer model, indicating PHGDH contributes to liver cancer progression mainly depending on its non-metabolic roles with nuclear location. PHGDH uses its ACT domain to bind cMyc in nuclear and forms a transactivation axis "PHGDH/p300/cMyc/AF9" which drives CXCL1/8 gene expression. Chemokines CXCL1/8 promotes neutrophils recruitment and then supports tumor associated macrophages (TAMs) filtration in liver, thereby urging liver cancer into advanced stages. Forced cytosolic location of PHGDH or destruction of the PHGDH/cMyc interaction abolishes the oncogenic function of nuclear PHGDH. Collectively, this study reveals a non-metabolic role of PHGDH with altered cellular location in liver cancer, and suggests a promising drug target for liver cancer therapy by targeting the interaction between PHGDH and "undruggable" cMyc.

category: cancer biology


title: PathoClock and PhysioClock in mice recapitulate human multimorbidity and heterogeneous aging

abstract: Multimorbidity is a public health concern and an essential component of aging and healthspan but understudied because investigative tools are lacking that can be translatable to capture similarities and differences of the aging process across species and variability between individuals and individual organs. To help address this need, body organ disease number (BODN) borrowed from human studies was applied to C57BL/6 (B6) and CB6F1 mouse strains at 8, 16, 24 and 32 months of age, as a measure of systems morbidity based on pathology lesions to develop a mouse PathoClock resembling clinically based Body Clock in humans, using Bayesian inference. A mouse PhysioClock was also developed based on measures of physiological domains including cardiovascular, neuromuscular, and cognitive function in the same two mouse strains so that alignment with BODN was predictable. The results revealed between- and within-age variabilities in PathoClock and PhysioClock, as well as between-strain variabilities. Both PathoClock and PhysioClock correlated with chronological age more strongly in CB6F1 than C57BL/6. Prediction models were then developed, designated as PathoAge and PhysioAge, using regression models of pathology and physiology measures on chronological age. PathoAge better predicted chronological age than PhysioAge as the predicted chronological and observed chronological age for PhysioAge were complex rather than linear. In conclusion, PathoClock and PhathoAge can be used to capture biological changes that predict BODN, a metric developed in human, and compare multimorbidity across species. These mouse clocks are potential translational tools that could be used in aging intervention studies.


title: Colon Innervating TRPA1 Positive Nociceptors Influence Mucosal Health In Mice

abstract:
IntroductionTransient receptor potential ankyrin-1 positive (TRPA1+ve) nociceptors, primarily present as peptidergic neuronal afferents in the colon are sensors of disturbance in lower gastrointestinal tract including pain induced by different pathologies. Their therapeutic role in the alleviation of chronic pain (receptor antagonism and receptor desensitization) associated with inflammatory bowel diseases (IBD) is reported. However, there is limited literature available about their role in formation and sustenance of the mucosal layer, and its interaction with host physiology as well as luminal microbial community. The aim of this study focuses on the effects of nociceptive TRPA1 channel desensitization on colonic mucus production and gut health.

MethodsTRPA1+ve nociceptors were desensitized by rectal administration of capsazepine. Ileum, colon was harvested and cecum content was collected. We performed morphological/histological analysis, gut permeability alteration, gene expression changes, colon metabolite profiling, and gut microbial abundance in these animals.

ResultsWe found that presence of TRPA1-positive nociceptors is required for mucus layer integrity, using an intra-rectal capsazepine-induced TRPA1 desensitization model. Desensitization of TRPA1 positive nociceptors resulted in damaged mucosal lining, resultant increase in gut permeability and altered transcriptional profile of genes for goblet cell markers, mucus regulation, immune response and tight junction proteins. The damage to mucosal lining prevented its role in enterosyne (short chain fatty acids) actions.

ConclusionThese results suggest that caution may need to be exercised before employing TRPA1 desensitization as a therapeutic option to alleviate pain caused due to IBD.

category: pharmacology and toxicology


title: The intestinal circadian clock drives microbial rhythmicity to maintain gastrointestinal homeostasis

abstract: Diurnal (i.e., 24-hour) oscillations of the gut microbiome have been described in various species including mice and humans. However, the driving force behind these rhythms remains less clear. In this study, we differentiate between endogenous and exogenous time cues driving microbial rhythms. Our results demonstrate that fecal microbial oscillations are maintained in mice kept in the absence of light, supporting a role of the hosts circadian system rather than representing a diurnal response to environmental changes. Intestinal epithelial cell-specific ablation of the core clock gene Bmal1 disrupts rhythmicity of microbiota. Targeted metabolomics functionally link intestinal clock-controlled bacteria to microbial-derived products, in particular branched-chain fatty acids and secondary bile acids. Microbiota transfer from intestinal clock-deficient mice into germ-free mice altered intestinal gene expression, enhanced lymphoid organ weights and suppressed immune cell recruitment. These results highlight the importance of functional intestinal clocks for circadian microbiota composition and function, which is required to balance the hosts gastrointestinal homeostasis.

category: physiology


title: The power of peer networking for improving STEM faculty job applications: a successful pilot program

abstract: In order to successfully obtain a faculty position, postdoctoral fellows or  postdocs, must submit an application which requires considerable time and effort to produce. These job applications are often reviewed by mentors and colleagues, but rarely are postdocs offered the opportunity to solicit feedback multiple times from reviewers with the same breadth of expertise often found on an academic search committee. To address this gap, this manuscript describes an international peer reviewing program for small groups of postdocs with a broad range of expertise to reciprocally and iteratively provide feedback to each other on their application materials. Over 145 postdocs have participated, often multiple times, over three years. A survey of participants in this program revealed that nearly all participants would recommend participation in such a program to other faculty applicants. Furthermore, this program was more likely to attract participants who struggled to find mentoring and support elsewhere, either because they changed fields or because of their identity as a woman or member of an underrepresented population in STEM. Participation in programs like this one could provide early career academics like postdocs with a diverse and supportive community of peer mentors during the difficult search for a faculty position. Such psychosocial support and encouragement has been shown to prevent attrition of individuals from these populations and programs like this one target the largest  leak in the pipeline, that of postdoc to faculty. Implementation of similar peer reviewing programs by universities or professional scientific societies could provide a valuable mechanism of support and increased chances of success for early-career academics in their search for independence.

category: scientific communication and education


title: Brain hubs defined in the group do not overlap with regions of high inter-individual variability

abstract: Connector  hubs are brain regions with links to multiple networks. These regions are hypothesized to play a critical role in brain function. While hubs are often identified based on group-average functional magnetic resonance imaging (fMRI) data, there is considerable inter-subject variation in the functional connectivity profiles of the brain, especially in association regions where hubs tend to be located. Here we investigated how group hubs are related to locations of inter-individual variability, to better understand if hubs are (a) relatively conserved across people, (b) locations with malleable connectivity, leading individuals to show variable hub profiles, or (c) artifacts arising from cross-person variation. To answer this question, we compared the locations of hubs and regions of strong idiosyncratic functional connectivity ("variants") in both the Midnight Scan Club and Human Connectome Project datasets. Group hubs defined based on the participation coefficient did not overlap strongly with variants. These hubs have relatively strong similarity across participants and consistent cross-network profiles. Consistency across participants was further improved when participation coefficient hubs were allowed to shift slightly in local position. Thus, our results demonstrate that group hubs defined with the participation coefficient are generally consistent across people, suggesting they may represent conserved cross-network bridges. More caution is warranted with alternative hub measures, such as community density, which are based on spatial proximity and show higher correspondence to locations of individual variability.

category: neuroscience


title: Different brain systems support the aversive and appetitive sides of human pain-avoidance learning

abstract: Both unexpected pain and unexpected pain absence can drive avoidance learning, but whether they do so via shared or separate neural and neurochemical systems is largely unknown. To address this issue, we combined an instrumental pain-avoidance learning task with computational modeling, functional magnetic resonance imaging (fMRI) and pharmacological manipulations of the dopaminergic (100 mg levodopa) and opioidergic (50 mg naltrexone) systems (N=83). Computational modeling provided evidence that untreated participants learned more from received than avoided pain. Our dopamine and opioid manipulations negated this learning asymmetry by selectively increasing learning rates for avoided pain. Furthermore, our fMRI analyses revealed that pain prediction errors were encoded in subcortical and limbic brain regions, whereas no-pain prediction errors were encoded in frontal and parietal cortical regions. However, we found no effects of our pharmacological manipulations on the neural encoding of prediction errors. Together, our results suggest that human pain-avoidance learning is supported by separate threat- and safety-learning systems, and that dopamine and endogenous opioids specifically regulate learning from successfully avoided pain.


title: Type 1 piliated uropathogenic Escherichia coli hijack the host immune response by binding to CD14

abstract: A key attribute of persistent or recurring bacterial infections is the ability of the pathogen to evade the hosts immune response. Many Enterobacteriaceae express type 1 pili, a pre-adapted virulence trait, to invade host epithelial cells and establish persistent infections. However, the molecular mechanisms and strategies by which bacteria actively circumvent the immune response of the host remain poorly understood. Here, we identified CD14, the major co-receptor for lipopolysaccharide detection, on dendritic cells as a previously undescribed binding partner of FimH, the protein located at the tip of the type 1 pilus of Escherichia coli. The FimH amino acids involved in CD14 binding are highly conserved across pathogenic and non-pathogenic strains. Binding of pathogenic bacteria to CD14 lead to reduced dendritic cell migration and blunted expression of co-stimulatory molecules, both rate-limiting factors of T cell activation. While defining an active molecular mechanism of immune evasion by pathogens, the interaction between FimH and CD14 represents a potential target to interfere with persistent and recurrent infections, such as urinary tract infections or Crohns disease.


title: The pathobiology of Mycobacterium abscessus revealed through phenogenomic analysis

abstract: The medical and scientific response to emerging pathogens is often severely hampered by ignorance of the genetic determinants of virulence, drug resistance, and clinical outcomes that could be used to identify therapeutic drug targets and forecast patient trajectories 1-5. Taking the newly emergent multidrug-resistant bacteria Mycobacterium abscessus as an example 6, we show that combining high dimensional phenotyping with whole genome sequencing in a phenogenomic analysis can rapidly reveal actionable systems-level insights into bacterial pathobiology. Using in vitro and in vivo phenotyping, we discovered three distinct clusters of isolates, each associated with a different clinical outcome. We combined genome-wide association studies (GWAS) with proteome-wide computational structural modelling 7 to define likely causal variants, and employed direct coupling analysis (DCA) 8 to identify co-evolving, and therefore potentially epistatic, gene networks. We then used in vivo CRISPR-based silencing to validate our findings, defining a novel secretion system controlling virulence in M. abscessus, and illustrating how phenogenomics can reveal critical pathways within emerging pathogenic bacteria.


title: Rvb1/Rvb2 proteins couple transcription and translation during glucose starvation

abstract: During times of unpredictable stress, organisms must adapt their gene expression to maximize survival. Along with changes in transcription, one conserved means of gene regulation during conditions that quickly represses translation is the formation of cytoplasmic phase-separated mRNP granules such as P-bodies and stress granules. Previously, we identified that distinct steps in gene expression can be coupled during glucose starvation as promoter sequences in the nucleus are able to direct the subcellular localization and translatability of mRNAs in the cytosol. Here, we report that Rvb1 and Rvb2, conserved ATPase proteins implicated as protein assembly chaperones and chromatin remodelers, were enriched at the promoters and mRNAs of genes involved in alternative glucose metabolism pathways that we previously found to be transcriptionally upregulated but translationally downregulated during glucose starvation in yeast. Engineered Rvb1/Rvb2-binding on mRNAs was sufficient to sequester the mRNAs into phase-separated granules and repress their translation. Additionally, this Rvb-tethering to the mRNA drove further transcriptional upregulation of the target genes. Overall, our results point to Rvb1/Rvb2 coupling transcription, mRNA granular localization, and translatability of mRNAs during glucose starvation. This Rvb-mediated rapid gene regulation could potentially serve as an efficient recovery plan for cells after stress removal.

category: molecular biology


title: Strip1 regulates retinal ganglion cell survival by suppressing Jun-mediated apoptosis to promote retinal neural circuit formation

abstract: In the vertebrate retina, an interplay between retinal ganglion cells (RGCs), amacrine, and bipolar cells establishes a synaptic layer called the inner plexiform layer (IPL). This circuit conveys signals from photoreceptors to visual centers in the brain. However, the molecular mechanisms involved in its development remain poorly understood. Striatin-interacting protein 1 (Strip1) is a core component of the STRIPAK complex, and it has shown emerging roles in embryonic morphogenesis. Here, we uncover the importance of Strip1 in inner retina development. Using zebrafish, we show that loss of Strip1 causes defects in IPL formation. In strip1 mutants, RGCs undergo dramatic cell death shortly after birth. Amacrine and bipolar cells subsequently invade the degenerating RGC layer, leading to a disorganized IPL. Mechanistically, zebrafish Strip1 interacts with its STRIPAK partner, Striatin3, and both show overlapping functions in RGC survival. Furthermore, loss of Strip1 or Striatin 3, leads to activation of the pro-apoptotic marker, Jun, within RGCs, and Jun knockdown rescues RGC survival in strip1 mutants. In addition to its function in RGC maintenance, Strip1 is required for RGC dendritic patterning, which likely contributes to proper IPL formation. Taken together, we propose that a series of Strip1-mediated regulatory events coordinates inner retinal circuit formation by maintaining RGCs during development, which ensures proper positioning and neurite patterning of inner retinal neurons.

category: developmental biology


title: Doors and corners of variance partitioning in statistical ecology

abstract: Variance partitioning is a common tool for statistical analysis and interpretation in both observational and experimental studies in ecology. Its popularity has led to a proliferation of methods with sometimes confusing or contradicting interpretations. Here, we present variance partitioning as a general tool in a model based Bayesian framework for summarizing and interpreting regression-like models. To demonstrate our approach we present a case study comprising of a simple occupancy model for a metapopulation of the Glanville fritillary butterfly. We pay special attention to the thorny issue of correlated covariates and random effects, and highlight uncertainty in variance partitioning. We recommend several alternative measures of variance, which jointly can be used to better interpret variance partitions. Additionally, we extend the general approach to encompass partitioning of variance within and between groups of observations, an approach very similar to analysis of variance. While noting that many troublesome issues relating to variance partitioning, such as uncertainty quantification, have been neglected in the literature, we likewise feel that the rather general applicability of the methods as an extension of statistical model-based analyses has not been fully utilized by the ecological research community either.

category: ecology


title: Outsourced hearing in spiders by using their webs as auditory sensors

abstract: Hearing is a fundamental sense of many animals, including all mammals, birds, some reptiles, amphibians, fish, and arthropods. The auditory organs of these animals are extremely diverse in anatomy after hundreds of millions of years of evolution, yet all are made up of cellular tissues and are morphologically part of bodies of animals. Here we show hearing in the orb-weaving spider, Larinioides sclopetarius is not constrained by the organisms body but is extended through outsourcing hearing to its extended phenotype, the proteinaceous, self-manufactured orb-web. We find the wispy, wheel-shaped orb-web acts as a hyperacute acoustic "antenna" to capture the sound-induced air particle movements that approach the maximum physical efficiency, better than the acoustic responsivity of all previously known eardrums. By sensing the motion of web threads, the spider remotely detects and localizes the source of an incoming airborne acoustic wave such as those emitted by approaching prey or predators. By outsourcing its acoustic sensors to its web, the spider is released from body size constraints and permits the araneid spider to increase its sound-sensitive surface area enormously, up to 10,000 times greater than the spider itself. The spider also enables the flexibility to functionally adjust and regularly regenerate its "external eardrum" according to its needs. The "outsourcing" and "supersizing" of auditory function in spiders provides unique features for studying extended and regenerative sensing, and designing novel acoustic flow detectors for precise fluid dynamic measurement and manipulation.


title: Outsourced hearing in an orb-weaving spider that uses its web as an auditory sensor


title: Circulatory neutrophils exhibit enhanced neutrophil extracellular trap formation in early puerperium: NETs at the nexus of thrombosis and immunity?

abstract: Pregnancy is associated with elevated maternal levels of cell-free DNA of neutrophil extracellular trap (NET) origin, as circulatory neutrophils exhibit increased spontaneous NET formation, mainly driven by G-CSF and finely modulated by sex hormones. The postpartum period, on the other hand, involves physiological alterations consistent with the need for protection against infections and fatal haemorrhage. Our findings indicate that all relevant serum markers of neutrophil degranulation and NET release are substantially augmented postpartum. Neutrophil pro-NETotic activity in vitro is also upregulated particularly in post-delivery neutrophils. Moreover, maternal puerperal neutrophils exhibit a strong pro-NETotic phenotype, associated with increased levels of all key players in the generation of NETs - citH3, MPO, NE and ROS, compared to non-pregnant and pregnant controls. Intriguingly, post-delivery NET formation is independent of G-CSF in contrast to late gestation and complemented by the presence of TF on the NETs, alterations in the platelet activity status, and activation of the coagulation cascade, triggered by circulating microparticles. Taken together, our results reveal the highly pro-NETotic and potentially procoagulant nature of postpartum neutrophils, bridging an overt immune activation with possible harmful thrombotic incidence.


title: Widespread Utilization of Diverse Organophosphate Pollutants by Marine Bacteria

abstract:
Anthropogenic organophosphates (AOPs), such as phosphotriesters, are used extensively as plasticizers, flame retardants, nerve agents and pesticides. Soil bacteria bearing a phosphotriesterase (PTE) can degrade AOPs, but whether bacteria are capable of utilizing AOPs as a phosphorus source, and how widespread PTEs are in nature, remains unclear. Here, we report the utilization of diverse AOPs by four model marine bacteria and seventeen bacterial isolates from seawater samples. To unravel the details of AOP utilization, two novel PTEs from marine bacteria were isolated and characterized. When expressed in E. coli, these PTEs enabled growth on a pesticide analog as the sole phosphorus source. Utilization of AOPs provides bacteria with a source of phosphorus in depleted environments and offers a new prospect for the bioremediation of a pervasive class of anthropogenic pollutants.

One sentence summaryWidespread utilization of diverse organophosphate pollutants by over 20 marine bacterial strains represents a new hope for ocean bioremediation.

category: microbiology


title: ASF1 activation PI3K/AKT pathway regulates sexual and asexual development in filamentous ascomycete

abstract: Sexual and asexual reproduction is ubiquitous in eukaryotes. PI3K/AKT signaling pathway can modulate sexual reproduction in mammals. However, this signaling pathway modulating sexual and asexual reproduction in fungi is scarcely understood. SeASF1, a SeH4 chaperone, could manipulate sexual and asexual reproduction of Stemphylium eturmiunum. SeDJ-1, screened from Se{Delta}asf1 transcriptome, was confirmed to regulate sexual and asexual development by RNAi, of which the mechanism was demonstrated by detecting transcriptional levels and protein interactions of SeASF1, SeH4 and SeDJ-1 by qRT-PCR, and Y2H, Co-IP and Pull-down, respectively. SeASF1 coupling SeH4 bound SeDJ-1 to arouse the sexual and asexual activity. In S. eturmiunum genome, SeDJ-1 was upstream while SeGSK3 was downstream in PI3K/AKT signaling pathway. Moreover, SeDJ-1 interacted with SePI3K or SeGSK3 in vivo and in vitro. Significantly, SeDJ-1 or SePI3K could effectively stimulate sexual activity alone, but SePI3K could recover the sexual development of SiSeDJ-1. Meanwhile, SeDJ-1-M6 was a critical segment for interaction of SeDJ-1 with SePI3K. SeDJ-1-M6 played a critical role in irritating sexual reproduction in SiSePI3K, which further uncovered the regulated mechanism of SeDJ-1. Summarily, SeASF1 coupling SeH4 motivates SeDJ-1 to arouse SePI3K involved in sexual reproduction. Thus, SeASF1 can activate PI3K/AKT signaling pathway to regulate sexual and asexual development in filamentous ascomycete.


title: A peroxide-responding sRNA evolved from a peroxidase mRNA

abstract:
Small RNAs (sRNAs) are critical regulators of gene expression in bacteria, but we lack a clear understanding of how new sRNAs originate and get integrated into regulatory networks. A major obstacle to elucidating their evolution is the difficulty in tracing sRNAs across large phylogenetic distances. To overcome this roadblock, we investigated the prevalence of sRNAs in more than a thousand genomes across Enterobacterales, a bacterial order with a rare confluence of factors that allows robust genome-scale sRNA analyses: several well-studied organisms with fairly conserved genome structures, an established phylogeny, and substantial nucleotide diversity within a narrow evolutionary space. Using a covariance modeling-based approach, we analyzed the presence of hundreds of sRNAs and discovered that a majority of sRNAs arose recently, and uncovered protein-coding genes as a potential source for the generation of new sRNA genes. A detailed investigation of the emergence of OxyS, a peroxide-responding sRNA, demonstrated that it evolved from a 3' end fragment of a peroxidase mRNA. Collectively, our data show that the erosion of protein-coding genes can result in the formation of new sRNAs that continue to be part of the original proteins regulon. This novel insight provides a fresh framework for understanding how new sRNAs originate and get incorporated into preexisting regulatory networks.

AUTHOR SUMMARYSmall RNAs (sRNAs) are important gene regulators in bacteria, but it is unclear how new sRNAs originate and become part of regulatory networks that coordinate bacterial response to environmental stimuli. Here, we show that new sRNAs could arise from protein-coding genes and potentially be incorporated into the ancestral proteins regulatory networks. We illustrate this process by defining the origin of OxyS. This peroxide-responding sRNA evolved from and replaced a peroxidase gene, but continues to be part of the peroxide-response regulon. In sum, we describe the source from which OxyS, one of the most well-studied sRNAs, arose, identify protein-coding genes as a potential raw material from which new sRNAs could emerge, and suggest a novel evolutionary path through which new sRNAs could get incorporated into pre-existing regulatory networks.


title: Bees can be trained to identify SARS-CoV-2 infected samples.

abstract: The COVID-19 pandemic has illustrated the need for the development of fast and reliable testing methods for novel, zoonotic, viral diseases in both humans and animals. Pathologies lead to detectable changes in the Volatile Organic Compound (VOC) profile of animals, which can be monitored, thus allowing the development of a rapid VOC-based test. In the current study, we successfully trained honeybees (Apis mellifera) to identify SARS-CoV-2 infected minks (Neovison vison) thanks to Pavlovian conditioning protocols. The bees can be quickly conditioned to respond specifically to infected minks odours and could therefore be part of a wider SARS-CoV-2 diagnostic system. We tested two different training protocols to evaluate their performance in terms of learning rate, accuracy and memory retention. We designed a non-invasive rapid test in which multiple bees are tested in parallel on the same samples. This provided reliable results regarding a subjects health status. Using the data from the training experiments, we simulated a diagnostic evaluation trial to predict the potential efficacy of our diagnostic test, which yielded a diagnostic sensitivity of 92% and specificity of 86%. We suggest that a honeybee-based diagnostics can offer a reliable and rapid test that provides a readily available, low-input addition to the currently available testing methods. A honeybee-based diagnostic test might be particularly relevant for remote and developing communities that lack the resources and infrastructure required for mainstream testing methods.


title: Differential nuclear import sets the timing of protein access to the embryonic genome

abstract:
The development of a fertilized egg to an embryo requires the proper temporal control of gene expression1-6. During cell differentiation, timing is often controlled via cascades of transcription factors (TFs)7,8. However, in early development, transcription is often inactive, and many TF levels are constant, suggesting that unknown mechanisms govern the observed rapid and ordered onset of gene expression9. Here, we find that in early embryonic development, access of maternally deposited nuclear proteins to the genome is temporally ordered via importin affinities, thereby timing the expression of downstream targets. We quantify changes in the nuclear proteome during early development and find that nuclear proteins, such as TFs and RNA polymerases, enter nuclei sequentially. Moreover, we find that the timing of the access of nuclear proteins to the genome corresponds to the timing of downstream gene activation. We show that the affinity of proteins to importin is a major determinant in the timing of protein entry into embryonic nuclei. Thus, we propose a mechanism by which embryos encode the timing of gene expression in early development via biochemical affinities. This process could be critical for embryos to organize themselves before deploying the regulatory cascades that control cell identities.

Graphical abstract

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title: Behavioral phenotyping of Zip8 393T-KI mice for in vivo study of schizophrenia pathogenesis

abstract: Genetic studies have informed on the genetic landscape of schizophrenia, and the next challenge is to link the genetic associations to mechanistic studies. A common single nucleotide polymorphism in the zinc and manganese transporter ZIP8 (rs13107325; ZIP8 A391T) is a top candidate to prioritize for functional studies because it is a missense mutation that results in hypomorphic protein function. With this goal, we have established a mouse model (Zip8 393T-knock-in (KI)), and here, we report the results of brain necropsy and initial behavioral phenotyping experiments in the KI mice using open field testing, elevated plus maze, Y-maze, and trace fear conditioning. Overall, male, homozygous KI mice may exhibit subtle defects in cognition and spatial learning, otherwise the baseline testing supports minimal behavioral differences between wild-type and Zip8 393T-KI mice. There were no genotype-specific alterations of gross or microscopic neuroanatomy. These experiments are important to establish the baseline characteristics of the Zip8 393T-KI mice that may be perturbed in animal models of schizophrenia and position the Zip8 393T-KI mouse as an important model for translational studies of schizophrenia pathogenesis.

category: genomics


title: Conserved chromatin and repetitive patterns reveal slow genome evolution in frogs

abstract: Frogs are an ecologically diverse and phylogenetically ancient group of living amphibians that include important vertebrate cell and developmental model systems, notably the genus Xenopus. Here we report a high-quality reference genome sequence for the western clawed frog, Xenopus tropicalis, along with draft chromosome-scale sequences of three distantly related emerging model frog species, Eleutherodactylus coqui, Engystomops pustulosus and Hymenochirus boettgeri. Frog chromosomes have remained remarkably stable since the Mesozoic Era, with limited Robertsonian (i.e., centric) translocations and end-to-end fusions found among the smaller chromosomes. Conservation of synteny includes conservation of centromere locations, marked by centromeric tandem repeats associated with Cenp-a binding, surrounded by pericentromeric LINE/L1 elements. We explored chromosome structure across frogs, using a dense meiotic linkage map for X. tropicalis and chromatin conformation capture (HiC) data for all species. Abundant satellite repeats occupy the unusually long ([~]20 megabase) terminal regions of each chromosome that coincide with high rates of recombination. Both embryonic and differentiated cells show reproducible association of centromeric chromatin, and of telomeres, reflecting a Rabl configuration similar to the "bouquet" structure of meiotic cells. Our comparative analyses reveal 13 conserved ancestral anuran chromosomes from which contemporary frog genomes were constructed.


title: Topological analysis of a bacterial DedA protein associated with alkaline tolerance and antimicrobial resistance.

abstract: Maintaining membrane integrity is of paramount importance to the survival of bacteria as the membrane is the site of multiple crucial cellular processes including energy generation, nutrient uptake, and antimicrobial efflux. The DedA family of integral membrane proteins are widespread in bacteria and are associated with maintaining the integrity of the membrane. In addition, DedA proteins have been linked to resistance to multiple classes of antimicrobials in various microorganisms. Therefore, the DedA family are attractive targets for the development of new antibiotics. Despite DedA family members playing a key physiological role in many bacteria, their structure, function and physiological role remain unclear. To help illuminate the structure of the bacterial DedA proteins, we have performed substituted cysteine accessibility method (SCAM) analysis on the most comprehensively characterized bacterial DedA protein, YqjA from Escherichia coli. By probing the accessibility of 15 cysteine residues across the length of YqjA using thiol reactive reagents, we have mapped the topology of the protein. Using these data, we have experimentally validated a structural model of YqjA generated using evolutionary co-variance, which consists of an -helical bundle with two re-entrant hairpin loops reminiscent of several secondary active transporters. In addition, our cysteine accessibility data suggests that YqjA forms an oligomer wherein the protomers are arranged in a parallel fashion. This experimentally verified model of YqjA lays the foundation for future work in understanding the function and mechanism of this interesting and important family.

category: biochemistry


title: Two distinct classes of co-chaperones compete for the EEVD motif in heat shock protein 70 (Hsp70) to tune its activity

abstract: Chaperones of the heat shock protein 70 (Hsp70) family engage in protein-protein interactions (PPIs) with many co-chaperones. One hotspot for co-chaperone binding is the EEVD motif that is found at the extreme C-terminus of cytoplasmic Hsp70s. This motif is known to bind tetratricopeptide repeat (TPR) domain co-chaperones, such as the E3 ubiquitin ligase CHIP, and Class B J-domain proteins (JDPs), such as DnaJB4. Although complexes between Hsp70-CHIP and Hsp70-DnaJB4 are both important for chaperone functions, the molecular determinants that dictate the competition between these co-chaperones are not clear. Using a collection of EEVD-derived peptides, we find that DnaJB4 binds to the IEEVD motif of Hsp70s, but not the related MEEVD motif of cytoplasmic Hsp90s. Then, we explored which residues are critical for binding to CHIP and DnaJB4, revealing that they rely on some shared features of the IEEVD motif, such as the C-terminal carboxylate. However, they also had unique preferences, especially at the isoleucine position. Finally, we observed a functionally important role for competition between CHIP and DnaJB4 in vitro, as DnaJB4 can limit the ubiquitination activity of the Hsp70-CHIP complex, while CHIP suppresses the chaperone activities of Hsp70-DnaJB4. Together, these results suggest that the EEVD motif has evolved to support diverse PPIs, such that competition between co-chaperones could help guide whether Hsp70-bound proteins are folded or degraded.


title: Plant-based production of SARS-CoV-2 antigens for use in a subunit vaccine

abstract: The COVID-19 pandemic has brought to the forefront an urgent need for the rapid development of highly efficacious vaccines, particularly in light of the ongoing emergence of multiple variants of concern. Plant-based recombinant protein platforms are emerging as cost-effective and highly scalable alternatives to conventional protein production. Viral glycoproteins, however, are historically challenging to produce in plants. Herein, we report the production of plant-expressed wild-type glycosylated SARS-CoV-2 Spike RBD (receptor-binding domain) protein that is recognized by anti-RBD antibodies and exhibits high-affinity binding to the SARS-CoV-2 receptor ACE2 (angiotensin-converting enzyme 2). Moreover, our plant-expressed RBD was readily detected by IgM, IgA, and IgG antibodies from naturally infected convalescent, vaccinated, or convalescent and vaccinated individuals. We further demonstrate that RBD binding to the ACE2 receptor was efficiently neutralized by antibodies from sera of SARS-CoV-2 convalescent and partially and fully vaccinated individuals. Collectively, these findings demonstrate that recombinant RBD produced in planta exhibits suitable biochemical and antigenic features for use in a subunit vaccine platform.

category: bioengineering


title: Pathogen-derived 9-methyl sphingoid base is perceived by a lectin receptor kinase in Arabidopsis

abstract:
In plants, many invading microbial pathogens are recognized by cell-surface pattern recognition receptors (PRRs), inducing defense responses; yet how PRRs perceive pathogen sphingolipids remains unclear. Here, we show that the ceramide Pi-Cer D from a plant pathogenic oomycete Phytophthora infestans triggers defense responses in Arabidopsis. Pi-Cer D is cleaved by an Arabidopsis apoplastic ceramidase, NCER2, and the resulting 9-methyl-branched sphingoid base is recognized by a plasma membrane lectin receptor-like kinase, RDA2. Importantly, 9-methyl-branched sphingoid base, which is unique to microbes, induces plant immune responses by interacting with RDA2. Loss of RDA2 or NCER2 function compromised Arabidopsis resistance against an oomycete pathogen, indicating that these are crucial for defense. We provide new insights that help elucidate the recognition mechanisms of pathogen-derived lipid molecules in plants.

One Sentence SummaryOomycete-derived ceramide is cleaved into sphingoid base by ceramidase and recognized by an Arabidopsis receptor kinase.