title: ULTRASTRUCTURAL ANALYSIS OF CRF-DA INTERACTIONS IN MALE AND FEMALE MACAQUE: BEYOND THE CLASSIC VTA

abstract:
Dopamine (DA) is involved in stress and stress-related illnesses, including many psychiatric disorders. Corticotropin-releasing factor (CRF) plays a role in stress responses and targets the ventral midbrain DA system. This system is comprised of DA and non-DA cells and is divided into specific subregions. Although CRF inputs to the midline A10 nuclei of the DA system are well studied in rodents, in monkeys, CRF-containing terminals are highly enriched in the expanded A10 parabrachial pigmented nucleus (PBP) and in the A8 subregion (retrorubral field). In primates, the central extended amygdala, a rich source of CRF afferents across species, preferentially targets the PBP and A8 fields. We thus sought to characterize CRF terminals on DA (tyrosine hydroxylase, TH+) and non-DA (TH-) cell types in the PBP and A8 regions at the ultrastructural level using immuno-reactive electron microscopy (EM) for TH and CRF in male and female macaques. CRF labeling was present mostly in axon terminals, which mainly contacted non-DA dendrites in both subregions. Most CRF-positive terminals had inhibitory (symmetric) profiles. In the A8, CRF symmetric (inhibitory) contacts onto non-DA neurons were significantly greater than symmetric (excitatory) profiles; this pattern was also seen in the PBP, but did not reach statistical significance. No sex differences were found. Hormonal assays suggested that our animals were at similar developmental stages and experienced similar stress levels. Together our findings suggest that at baseline, CRF terminals in the primate PBP and A8 largely regulate DA indirectly through non-DA neurons.

KEY POINTSO_LIIn addition to midline A10 neurons, CRF terminals robustly target the enlarged A10 PBP and A8 regions in primate ventral midbrain
C_LIO_LICRF terminals in the Macaque PBP/A8 mainly contact non-DA neurons compared to DA neurons
C_LIO_LIRelatively more CRF-positive terminals are symmetric (inhibitory) versus asymmetric (excitatory) in both PBP and A8, suggesting indirect, possibly interneuronal, mechanisms of DA control.
C_LI


title: Genome-wide CRISPRi screening reveals regulators of Alzheimee's tau pathology shared between exosomal and vesicle-free tau seeds.

abstract: Aggregation of the microtubule-associated protein tau is a defining feature of Alzheimers disease and other tauopathies. Tau pathology is believed to be driven by both free tau aggregates and tau carried within exosomes, which propagate trans-synaptically and induce tau pathology in recipient neurons by a corrupting process of seeding. Here, we performed a genome-wide CRISPRi screen in tau biosensor cells and identified cellular regulators shared by both mechanisms of tau seeding. The top validated regulators are ANKLE2, BANF1, NUSAP1, EIF1AD, and VPS18, which work as factors that restrict tau aggregation initiated by both exosomal and vesicle-free tau seeds. Interestingly, ANKLE2 and BANF1 more robustly affected exosomal tau seeding than free aggregates. Lastly, validation studies revealed that several of the identified protein hits are downregulated in the brains of Alzheimers patients, suggesting that their decreased activity may be required for the emergence or progression of tau pathology in the human brain.

category: neuroscience


title: High Concentrations of floating life in the North Pacific Garbage Patch

abstract: Floating life (obligate neuston) is a core component of the ocean surface food web. However, only one region of high neustonic abundance is known so far, the Sargasso Sea in the Subtropical North Atlantic, where floating life provides critical habitat structure and ecosystem services. Here, we hypothesize that floating life is also concentrated in other gyres with converging surface currents. To test this hypothesis, we collected samples through the eastern North Pacific Subtropical Gyre in the area of the North Pacific "garbage patch" (NPGP) known to accumulate floating anthropogenic debris. We found that densities of floating life were significantly higher inside the central part of NPGP than on its periphery, and there was a significant positive relationship between neuston abundance and plastic abundance. This work has important implications for the ecology and human impact of subtropical oceanic gyre ecosystems.

category: ecology


title: The distribution of the Lansing Effect across animal species

abstract: Maternal senescence is a reduction in individual performance associated with an increase in its mothers age at conception. When manifested on adult lifespan, this is known as the  Lansing Effect. Single-species studies report both maternal age-related increases and decreases in adult lifespan, but no comprehensive review of the literature has yet determined if the Lansing Effect is a widespread phenomenon. To address this knowledge gap, we performed a meta-analysis of maternal aging rates taken from all available published studies. We recovered 74 estimates from 20 studies representing 14 species. All studies taken together suggest a propensity for a Lansing Effect with an estimated average effect of maternal age on adult lifespan of between -22% to -17% (the lifespan response to an increase in maternal age expressed in the same time units), depending upon our specific choice of model. We failed to find a significant effect of animal class or insect order, but given the oversampling of insect species in the published literature and the paucity of vertebrate studies, we infer that only rotifers and insects demonstrate a tendency for expressing the phenomenon.

category: evolutionary biology


title: Variable paralog expression underlies phenotype variation

abstract: Human faces are variable; we look different from one another. Craniofacial disorders further increase this variability. Here we used the zebrafish mef2ca mutant, which produces variable phenotypes, to understand craniofacial variation. Comparing different mef2ca alleles demonstrated that severity, measured by penetrance and expressivity, correlates with variation. Years of selective breeding for low and high penetrance produced strains that are either resilient, or sensitive, to the mef2ca mutation. Comparing these strains further demonstrates that severity correlates with variation. Gene expression studies indicated that selective breeding upregulated and downregulated mef2ca paralog expression in the low- and high-penetrance strains, respectively. We hypothesized that heritable paralog expression variation underlies mutant phenotype variation. In support, mutagenizing all mef2ca paralogs in the low-penetrance strain demonstrated modular buffering by paralogs. Specifically, some paralogs buffer severity while others buffer variability. We present a novel, mechanistic model for phenotypic variation where cryptic vestigial paralog expression modularly buffers development and contributes to evolution. These studies are a major step forward in understanding of the mechanisms of facial variation, including how some genetically resilient individuals can overcome a deleterious mutation.

category: genetics


title: A fast machine-learning-guided primer design pipeline for selective whole genome amplification

abstract: Addressing many of the major outstanding questions in the fields of microbial evolution and pathogenesis will require analyses of populations of microbial genomes. Although population genomic studies provide the analytical resolution to investigate evolutionary and mechanistic processes at fine spatial and temporal scales - precisely the scales at which these processes occur - microbial population genomic research is currently hindered by the practicalities of obtaining sufficient quantities of the relatively pure microbial genomic DNA necessary for next-generation sequencing. Here we present swga2.0, an optimized and parallelized pipeline to design selective whole genome amplification (SWGA) primer sets. Unlike previous methods, swga2.0 incorporates active and machine learning methods to evaluate the amplification efficacy of individual primers and primer sets. Additionally, swga2.0 optimizes primer set search and evaluate strategies, including parallelization at each stage of the pipeline, to dramatically decrease program runtime from weeks to minutes. Here we describe the swga2.0 pipeline, including the empirical data used to identify primer and primer set characteristics, that improve amplification performance. Additionally, we evaluated the novel swga2.0 pipeline by designing primers sets that successfully amplify Prevotella melaninogenica, an important component of the lung microbiome in cystic fibrosis patients, from samples dominated by human DNA.

category: genomics


title: Structural insights into the high selectivity of the anti-diabetic drug mitiglinide

abstract: Mitiglinide is a highly selective fast-acting anti-diabetic drug that inhibits pancreatic KATP channels to induce insulin secretion. However, how mitiglinide binds KATP channels remains unknown. Here, we show the cryo-EM structure of the SUR1 subunit in complex with mitiglinide. The structure reveals that mitiglinide binds inside the common insulin secretagogue-binding site in the transmembrane domain of SUR1, locking SUR1 in a NBD-separated inward-facing conformation. The detailed structural analysis uncovers the molecular basis of the high selectivity of mitiglinide.

category: biophysics


title: neuroWalknet, a controller for hexapod walking allowing for context dependent behavior

abstract:
Decentralized control has been established as a key control principle in insect walking and has been successfully leveraged to account for a wide range of walking behaviors in the proposed neuroWalknet architecture. This controller allows for walking patterns with different velocities in forward and backward direction -- quite similar to the behavior shown in stick insects --, for negotiation of curves, and for robustly dealing with various disturbances.

While these simulations focus on the cooperation of different, decentrally controlled legs, here we consider a set of biological experiments not yet been tested by neuroWalknet, that focus on the function of the individual leg and are context dependent. These intraleg studies deal with four groups of interjoint reflexes. The reflexes are elicited by either a stimulation of the femoral chordotonal organ (fCO) or a specific stimulation of campaniform sensilla (CS). Motor output signals are recorded from the alpha-joint, the beta-joint or the gamma-joint of the leg. Furthermore, such reflexes have been studied while the ganglion was treated with pilocarpine.

Although these biological data represent results obtained from different local reflexes in different contexts, they fit with and are embedded into the behavior shown by the global structure of neuroWalknet. In particular, a specific and intensively studied behavior, active reaction, has since long been assumed to represent a separate behavioral element, from which it is not clear why it occurs in some situations, but not in others. This question could now be explained as an emergent property of the holistic structure of neuroWalknet. When experimenting with pilocarpine, oscillations were induced in neuroWalknet even though this does not include an explicit central pattern generator and in this way provides a simpler model as a functional explanation. As the simulation data result from a holistic system, further results were obtained that could be used as predictions to be tested in further biological experiments.

AUTHOR SUMMARYBehavior of animals can be studied by detailed observation, but observation alone does not explain the function of the underlying neuronal controller structures. To better understand this function, an important tool can be to develop an artificial structure based on simulated neurons and a simulated or physical body. Although typical animal behavior appears complex, the corresponding neuronal structures may be comparatively simple.

The goal for such a hypothetical structure should be to include as many different behaviors as possible, and, at the same time, search for a simple explanation consisting of a minimum of neuronal elements. Furthermore, such a simulation system, e.g. an artificial neuronal network, should contain hypotheses that can be tested in biological experiments.

We propose an extension to such a network that is based on a decentralized neuronal structure, using a neural network as a scaffold, that enables various combinations of local neuronal elements that allow for emergent, i.e. not explicitly designed properties. Indeed, neuroWalknet contains further abilities not yet recognized in the earlier version. For instance, neither explicit structures like central pattern generators nor explicit Active Reaction are required to reproduce typical intraleg reactions. Therefore, neuroWalknet presents a holistic approach enabling emergent properties out of the cooperation of small neuronal elements that are context dependent instead of explicit, dedicated elements.


title: Characterizing dynamic neural representations of scene attractiveness

abstract: Aesthetic experiences during natural vision are varied: they can arise from viewing scenic landscapes, interesting architecture, or attractive people. Recent research in the field of neuroaesthetics has taught us a lot about where in the brain such aesthetic experiences are represented. Much less is known about when such experiences arise during the cortical processing cascade. Particularly, the dynamic neural representation of perceived attractiveness for rich natural scenes is not well understood. Here, I present data from an EEG experiment, in which participants provided attractiveness judgments for a set of diverse natural scenes. Using multivariate pattern analysis, I demonstrate that scene attractiveness is mirrored in early brain signals that arise within 200ms of vision, suggesting that the aesthetic quality of scenes is first resolved during perceptual processing. In more detailed analyses, I show that even such early neural correlates of scene attractiveness are partly related to inter-individual variation in aesthetic preferences and that they generalize across scene contents. Together, these results characterize the time-resolved neural dynamics that give rise to aesthetic experiences in complex natural environments.


title: Detecting epistatic interactions in genomic data using Random Forests

abstract:
Epistatic interactions can play an important role in the genetic mechanisms that control phenotypic variation. However, identifying these interactions in high dimensional genomic data can be very challenging due to the large computational burden induced by the high volume of combinatorial tests that have to be performed to explore the entire search space. Random Forests Decision Trees are widely used in a variety of disciplines and are often said to detect interactions. However, Random Forests models do not explicitly detect variable interactions. Most Random Forests based methods that claim to detect interactions rely on different forms of variable importance measures that suffer when the interacting variables have very small or no marginal effects. The proposed Random Forests based method detects interactions using a two-stage approach and is computationally efficient. The approach is demonstrated and validated through its application on several simulated datasets representing different data structures with respect to genomic data and trait heritabilities. The method is also applied to two high dimensional genomics data sets to validate the approach. In both cases, the method results were used to identify several genes closely positioned to the interacting markers that showed strong biological potential for contributing to the genetic control for the respective traits tested.

Contacthawlader.almamun@csiro.au

category: bioinformatics


title: Extension of PERMANOVA to Testing the Mediation Effect of the Microbiome

abstract:
BackgroundRecently, we have seen a growing volume of evidence linking the microbiome and human diseases or clinical outcomes, as well as evidence linking the microbiome and environmental exposures. Now comes the time to assess whether the microbiome mediated the effects of the exposures on the outcomes, which will enable researchers to develop interventions to modulate the outcomes by modifying the microbiome composition. Use of distance matrices is a popular approach to analyzing complex microbiome data that are high-dimensional, sparse, and compositional. However, the existing distance-based methods for mediation analysis of microbiome data, MedTest and MODIMA, only work well in limited scenarios.

ResultsPERMANOVA is currently the most commonly used distance-based method for testing microbiome associations. Using the idea of inverse regression, here we extend PER-MANOVA to testing microbiome mediation effects by including both the exposure and the outcome as covariates and basing the test on the product of their F-statistics. This extension of PERMANOVA, which we call PERMANOVA-med, naturally inherits all the flexible features of PERMANOVA, e.g., allowing adjustment of confounders, accommodating continuous, binary, and multivariate exposure and outcome variables including survival outcomes, and providing an omnibus test that combines the results from analyzing multiple distance matrices. Our extensive simulations indicated that PERMANOVA-med always controlled the type I error and had compelling power over MedTest and MODIMA. Frequently, MedTest had diminished power and MODIMA had inflated type I error. Using real data on melanoma immunotherapy response, we demonstrated the wide applicability of PERMANOVA-med through 16 different mediation analyses, only 6 of which could be performed by MedTest and 4 by MODIMA.

Availability and ImplementationPERMANOVA-med has been added to the existing function "permanovaFL" in our R package LDM, which is available on GitHub at https://github.com/yijuanhu/LDM.


title: Liposome-assisted in-situ cargo delivery to artificial cells and cellular subcompartments

abstract: We report on liposome-mediated targeted delivery of membrane-impermeable constituents into surface-adhered giant lipid compartments, employed as artificial cells. Soluble cargo compounds are delivered by means of an open-space microfluidic device, which perfuses selected lipid compartments with loaded small unilamellar vesicles (SUVs) composed of cationic lipids. The SUV membranes fuse with the surface-adhered containers, merging their contents. We monitored the fusion process via Forster resonance energy transfer (FRET) by labeling both the membranes of the SUVs and the target compartments with a fluorophore pair. We established that, upon fusion, water-soluble dyes, fluorescently labeled genetic polymers, sugars and proteins carried by the SUVs can be successfully internalized at high yield. Finally, by transferring carbonic anhydrase (CA) to the giant lipid compartments, enzymatic hydrolysis of the prefluorescent carboxyfluorescein diacetate (CFDA) is demonstrated by the emission intensity increase emanating from the product carboxyfluorescein (CF). Spontaneous subcompartmentalization occurred during liposomal delivery of the enzyme, leading to CF formation in an organelle-like subcompartment. The reported targeted delivery technique enables chemical reactions and cell-free gene expression in synthetic cell models with unprecedented ease and precision, and opens pathways to protocell architectures with distinct functional subcompartments in the context of origins of life research.


title: Truncated suPAR simultaneously causes kidney disease and autoimmune diabetes mellitus

abstract:
Soluble urokinase-type plasminogen activator receptor (suPAR) is a risk factor for kidney diseases. Here we report the presence of C-terminal suPAR fragment, D2D3, in patients with diabetic nephropathy. D2D3-positive human sera inhibited glucose-stimulated insulin release in human islets and were associated with patients requiring insulin therapy. D2D3 transgenic mice presented kidney disease and diabetes marked by decreased levels of insulin and C-peptide, impaired glucose-stimulated insulin secretion, decreased pancreatic {beta}-cell mass, and high fasting glucose. D2D3 fragment dysregulated glucose-induced cytoskeletal dynamics, impaired maturation and trafficking of insulin granules, and inhibited bioenergetics of {beta}-cells in culture. An anti-uPAR antibody restored {beta}-cell function in D2D3 transgenic mice. We show that the D2D3 fragment injures the kidney and pancreas, offering a unique dual therapeutic approach for kidney diseases and insulin-dependent diabetes.

SummaryProteolytic suPAR fragment, D2D3, simultaneously injures two organs, the kidney and pancreas, thus causing a dual organ disease.

category: physiology


title: Spleen voltammetry assesses in real time norepinephrine release elicited by autonomic stimulation

abstract: Bioelectronic medicine uses neurostimulation devices to modulate the activity of autonomic nerves and restore function in visceral organs. The autonomic, noradrenergic, innervation of the spleen is implicated in the neural control of inflammation; for that reason, the spleen is the target of autonomic stimulation therapies tested in rheumatoid arthritis and other inflammatory diseases. However, there are no real time markers for successful engagement of the spleen by autonomic stimulation. Here, we use fast-scan cyclic voltammetry (FSCV) in the spleen to assess, in real time, the release of norepinephrine (NE) in response to autonomic stimulation in mice. We show that spleen voltammograms demonstrate oxidative current peaks at a voltage of ~0.8 V that is proportional up to a NE blood concentration of 30 to 300nM. The oxidative current increases within seconds in response to electrical splenic nerve stimulation and is blocked by interventions that deplete NE or inhibit NE release. Oxidative current increases in response to electrical vagus nerve stimulation (VNS) and is elicited by efferent, but not afferent, fiber-selective optogenetic VNS. Spleen cyclic voltammetry can potentially provide a real-time physiological marker for dose calibration and monitoring of the anti-inflammatory effectiveness of autonomic stimulation therapies targeting the spleen.


title: Neural dynamics and architecture of the heading direction circuit in a vertebrate brain

abstract: Animals can use different strategies to navigate. They may guide their movements by relying on external cues in their environment or, alternatively, by using an internal cognitive map of the space around them and their position within it. An essential part of this representation are heading cells, neurons whose activity depends on the heading direction of the animal. Although those cells have been found in vertebrates, the full network has never been observed and there is very little mechanistic understanding of how these cells acquire their response properties. In this study, we use volumetric functional imaging in larval zebrafish to observe, for the first time in a vertebrate, a full network that encodes allocentric heading direction. This network of approximately one hundred inhibitory neurons is arranged in an anatomical circle in the anterior hindbrain. Its activity is driven purely by the integration of internally generated signals, indicating that a simple vertebrate brain can encode maps of how an animal moves within its surroundings. Single cell reconstructions of electron micrographs allow us to uncover how the connectivity pattern of neurons within the network supports the implementation of a ring attractor network. The neurons we identify share features with neurons in the dorsal tegmentum nucleus of rodents and the fly central complex, showing that similar connectivity and mechanistic principles underlie the generation of cognitive maps of heading direction across the animal kingdom.


title: Disease diagnostics using machine learning of immune receptors

abstract: Clinical diagnoses rely on a wide variety of laboratory tests and imaging studies, interpreted alongside physical examination and documentation of symptoms and patient history. However, the tools of diagnosis make little use of the immune systems internal record of specific disease exposures encoded by the antigen-specific receptors of memory B cells and T cells. We have combined extensive receptor sequence datasets with three different machine learning representations of the contents of immune repertoires to develop an interpretive framework, MAchine Learning for Immunological Diagnosis (Mal-ID), that screens for multiple illnesses simultaneously. This approach can already reliably distinguish a wide range of disease states, including specific acute or chronic infections, and autoimmune or immunodeficiency disorders, and could contribute to identifying new infectious diseases as they emerge. Importantly, many features of the model of immune receptor sequences are human-interpretable. They independently recapitulate known biology of the responses to infection by SARS-CoV-2 or HIV, and reveal common features of autoreactive immune receptor repertoires, indicating that machine learning on immune repertoires can yield new immunological knowledge.

category: immunology


title: Lhx9 haploinsufficiency alters transcription in the adult mouse ovary, causing subfertility and abnormal epithelium

abstract:
Understanding the molecular pathways that underpin ovarian development and function is vital for improving the research approaches to investigating fertility. Despite a significant improvement in our knowledge of molecular activity in the ovary, many questions remain unanswered in the quest to understand factors influencing fertility and ovarian pathologies such as cancer. Here we present an investigation into the expression and function of developmental transcription factor LIM Homeobox 9 (LHX9) in the adult mouse ovary. We have characterised Lhx9 expression in several cell types of the mature ovary across follicle stages. To elucidate the function of this expression, we carried out an investigation of ovarian anatomy and transcription in a Lhx9+/- knockout mouse model displaying subfertility. Despite a lack of gross anatomical differences between genotypes, RNA-sequencing found that 90 genes were differentially expressed between Lhx9+/- and Lhx9+/+ mice. Gene ontology analyses revealed a downregulation of genes with major roles in ovarian steroidogenesis and an upregulation of genes with implications for ovarian cancer. Analysis of the ovarian epithelium revealed Lhx9+/- mice have a disorganised epithelial phenotype and a significant increase in epithelial marker gene expression. These results provide an analysis of Lhx9 in the adult mouse ovary and a new candidate for fertility research and ovarian epithelial cancer.

Summary sentenceLhx9 haploinsufficient mice are subfertile with altered expression of steroid genes in the adult ovary and abnormal ovarian surface epithelium.

category: developmental biology


title: ALFF Response Interaction With Learning During Feedback in Individuals With Multiple Sclerosis

abstract: Amplitude of low-frequency fluctuations (ALFF) is defined as changes of BOLD signal during resting state (RS) brain activity. Previous studies identified differences in RS activation between healthy and multiple sclerosis (MS) participants. However, no research has investigated the relationship between ALFF and learning in MS. We thus examine this here. Twenty-five MS and 19 healthy participants performed a paired-associate word learning task where participants were presented with extrinsic or intrinsic performance feedback. Compared to healthy participants, MS participants showed higher local brain activation in the right thalamus. We also observed a positive correlation in the MS group between ALFF and extrinsic feedback within the left inferior frontal gyrus, and within the left superior temporal gyrus in association with intrinsic feedback. Healthy participants showed a positive correlation in the right fusiform gyrus between ALFF and extrinsic feedback. Findings suggest that while MS participants do not show a feedback learning impairment compared to the healthy participants, ALFF differences might suggest a general maladaptive pattern of task unrelated thalamic activation and adaptive activation in frontal and temporal regions. Results indicate that ALFF can be successfully used at capturing pathophysiological changes in local brain activation in MS in association with learning through feedback.


title: tDCS induced GABA change is associated with the simulated electric field in M1, an effect mediated by grey matter volume in the MRS voxel

abstract:
Background and ObjectiveTranscranial direct current stimulation (tDCS) has wide ranging applications in neuro-behavioural and physiological research, and in neurological rehabilitation. However, it is currently limited by substantial inter-subject variability in responses, which may be explained, at least in part, by anatomical differences that lead to variability in the electric field (E-field) induced in the cortex. Here, we tested whether the variability in the E-field in the stimulated cortex during tDCS, estimated using computational simulations, explains the variability in tDCS induced changes in GABA, a neurophysiological marker of stimulation effect.

MethodsData from five previously conducted MRS studies were combined. The anode was placed over the left primary motor cortex (M1, 3 studies, N = 24) or right temporal cortex (2 studies, N = 32), with the cathode over the contralateral supraorbital ridge. Single voxel spectroscopy was performed in a 2x2x2cm voxel under the anode in all cases. MRS data were acquired before and either during or after 1mA tDCS using either a sLASER sequence (7T) or a MEGA-PRESS sequence (3T). sLASER MRS data were analysed using LCModel, and MEGA-PRESS using FID-A and Gannet. E-fields were simulated in a finite element model of the head, based on individual MPRAGE images, using SimNIBS. Separate linear mixed effects models were run for each E-field variable (mean and 95th percentile; magnitude, and components normal and tangential to grey matter surface, within the MRS voxel). The model included effects of time (pre or post tDCS), E-field, grey matter volume in the MRS voxel, and a 3-way interaction between time, E-field and grey matter volume. Additionally, we ran a permutation analysis using PALM to determine whether E-field anywhere in the brain, not just in the MRS voxel, correlated with GABA change.

ResultsIn M1, higher mean E-field magnitude was associated with greater tDCS-induced decreases in GABA (t(24) = 3.24, p = 0.003). Further, the association between mean E-field magnitude and GABA change was moderated by the grey matter volume in the MRS voxel (t(24) = -3.55, p =0.002). These relationships were consistent across all E-field variables except the mean of the normal component. No significant relationship was found between tDCS-induced GABA decrease and E-field in the temporal voxel. No significant clusters were found in the whole brain analysis.

ConclusionsOur data suggest that the electric field induced by tDCS within the brain is variable, and is significantly related to tDCS-induced decrease in GABA, a key neurophysiological marker of stimulation. These findings strongly support individualised dosing of tDCS, at least in M1. Further studies examining E-fields in relation to other outcome measures, including behaviour, will help determine the optimal E-fields required for any desired effects.

HighlightsO_LIWe study the link between individually simulated electric field dose and tDCS-induced change in GABA in the cortex.
C_LIO_LIThe electric field strength in the brain correlates with a decrease in GABA in the motor cortex.
C_LIO_LIThe correlation between the electric field and GABA change is modulated by the amount of grey matter in the MRS voxel.
C_LIO_LIWe find no association between the electric field and GABA in the temporal cortex.
C_LI


title: Dapagliflozin improves endothelial integrity and hemodynamics in endotoxin treated mice through an apolipoprotein M dependent pathway

abstract:
RationaleSodium-glucose co-transporter inhibitors (SGLT2i) are under active clinical investigation in patients with acute inflammatory conditions, based on their clinical cardio-and nephroprotective effects, and a pre-clinical study that demonstrated SGLT2i improve renal outcomes and survival in a lipopolysaccharide (LPS) model. However, a unified mechanism that explains how SGLT2i could prevent hemodynamic consequences of inflammatory conditions has not been described. Apolipoprotein M (ApoM) is inversely associated with mortality in inflammatory conditions and improves cardiac function in endotoxin-treated mice via sphingosine-1-phosphate (S1P) signaling.

ObjectiveTest the hypothesis that pre-treatment with SGLT2i dapagliflozin (Dapa) improves hemodynamics in endotoxin-treated mice via the ApoM/S1P pathway.

Methods and ResultsMice with diet-induced obesity were gavaged with vehicle or Dapa for 4 days prior to LPS (10 mg/kg, IP). We found that mice receiving Dapa restored circulating ApoM levels, likely by increasing expression of the multi-ligand protein receptor megalin in the proximal tubules. Dapa attenuated LPS-induced reductions in cardiac dysfunction including reductions in ejection fraction, cardiac index, and coronary sinus area as well as vascular permeability as ascertained by intravital microscopy. Using both ApoM transgenic and knockout mice and S1P receptor inhibitors, we show that the ApoM/S1P pathway is important for the beneficial effects of Dapa in the LPS model.

ConclusionsIn the setting of acute inflammation, our data suggest that SGLT2i maintains levels of megalin, leading to preservation of ApoM, which in turn promotes endothelial barrier integrity and improves hemodynamics. Our studies suggest a novel mechanism by which SGLT2i can preserve intravascular volume in the acute inflammatory setting.


title: Diverse states and stimuli tune olfactory receptor expression levels to modulate food-seeking behavior

abstract: Animals must weigh competing needs and states to generate adaptive behavioral responses to the environment. Sensorimotor circuits are thus tasked with integrating diverse external and internal cues relevant to these needs to generate context-appropriate behaviors. However, the mechanisms that underlie this integration are largely unknown. Here, we show that a wide range of states and stimuli converge upon a single C. elegans olfactory neuron to modulate food-seeking behavior. Using an unbiased ribotagging approach, we find that the expression of olfactory receptor genes in the AWA olfactory neuron is influenced by a wide array of states and stimuli, including feeding state, physiological stress, and recent sensory cues. We identify odorants that activate these state-dependent olfactory receptors and show that altered expression of these receptors influences food-seeking and foraging. Further, we dissect the molecular and neural circuit pathways through which these diverse cues are integrated by AWA. This reveals a modular organization in which state-related signals arising from different cell types in the body converge on AWA and independently control chemoreceptor expression. The synthesis of these signals by AWA allows animals to generate sensorimotor responses that reflect the animals overall state. Our findings suggest a general model in which state-dependent transcriptional changes at the sensory periphery modulate animals sensorimotor responses to meet their ongoing needs and states.


title: SARM1 activation and its downstream pathways are distinct in neuronal compartments

abstract: Sterile alpha and TIR motif containing 1 (SARM1) is a critical regulator of axon degeneration that acts through hydrolysis of NAD+ following injury. Recent work has defined the structure and catalytic activity of SARM1, yet the specific pathways controlled by SARM1 following diverse neuronal insults are not well defined. Here we show that the mechanisms of SARM1 activation and downstream signaling pathways are both context and localization dependent. Endogenous SARM1 is present in axons, dendrites and cell bodies and is primarily localized to the outer mitochondrial membrane. In cultured hippocampal neurons, knockout of SARM1 protects axons from degeneration by treatment with mitochondrial toxins but does not affect the degeneration of other cell compartments. In contrast, direct activation of SARM1 leads to degeneration of axons, dendrites, and cell bodies, indicating that SARM1 is present and can be activated in these neuronal compartments. Inhibition of dual leucine zipper kinase (DLK, MAP3K12) and caspases were not protective in these paradigms, though both SARM1-dependent and independent dendrite degeneration required calpain activity. Together, these results demonstrate that SARM1 activation is not specific to axons though distinct pathways regulate degeneration in axons and dendrites.


title: The single-cell atlas of the murine reproductive tissues during preterm labor

abstract: Preterm birth, the leading cause of perinatal morbidity and mortality worldwide, frequently results from the multi-etiological syndrome of preterm labor. Intra-amniotic infection is the best-established causal link to preterm labor and birth, and involves the premature activation of the parturition cascade in the reproductive tissues (uterus, decidua, and cervix). However, the simultaneous investigation of the cellular landscape and interaction networks in such tissues at single-cell resolution had not been undertaken. Herein, we utilized scRNA-seq to reveal that preterm labor affects the cellularity of the uterus, decidua, and cervix through immune cell infiltration and altered transcriptomic profiles of non-immune cell types. Preterm labor-affected cells showed shared and distinct immune processes, the latter indicating tissue-specific cellular functions. Cell-cell communication analysis implicated immune and non-immune cell types in preterm labor-associated inflammatory pathways across tissues. Next, correlation between uterine transcriptomic data from mice with local or systemic inflammation-induced preterm labor indicated significant overlap in labor-associated gene expression. Last, comparison of cell-cell communication in the murine and human uterus revealed shared signaling pathways implicated in preterm and term parturition. Together, these findings provide novel insight into the cellular landscape, dynamics, and communications in the reproductive tissues during preterm labor leading to premature birth.


title: Two enhancer binding proteins activate σ54 -dependent transcription of a quorum regulatory RNA in a bacterial symbiont

abstract: To colonize a host, bacteria depend on an ensemble of signaling systems to convert information about the various environments encountered within the host into specific cellular activities. How these signaling systems coordinate transitions between cellular states in vivo remains poorly understood. To address this knowledge gap, we investigated how the bacterial symbiont Vibrio fischeri initially colonizes the light organ of the Hawaiian bobtail squid Euprymna scolopes. Previous work has shown that the small RNA Qrr1, which is a regulatory component of the quorum-sensing system in V. fischeri, promotes host colonization. Here, we report that transcriptional activation of Qrr1 is repressed by the regulator BinK, which inhibits cellular aggregation by V. fischeri prior to light organ entry. We show that Qrr1 expression depends on the alternative sigma factor {sigma}54 and the transcription factors LuxO and SypG, which function together as an OR logic gate, thereby ensuring Qrr1 is expressed during colonization. Finally, we provide evidence that this regulatory mechanism is widespread throughout the Vibrionaceae family. Together, our work reveals how coordination between the signaling pathways underlying aggregation and quorum sensing promotes host colonization, which provides insight into how integration among signaling systems facilitates complex processes in bacteria.

category: microbiology


title: A toolbox for systematic discovery of stable and transient protein interactors in baker's yeast

abstract: Identification of both stable and transient interactions is essential for understanding protein function and regulation. While assessing stable interactions is more straightforward, capturing transient ones is challenging. In recent years, sophisticated tools have emerged to improve transient interactor discovery, with many harnessing the power of evolved biotin ligases for proximity labelling. However, biotinylation-based methods have lagged behind in the model eukaryote, Saccharomyces cerevisiae, possibly due to the presence of several abundant, endogenously biotinylated proteins. In this study, we optimised robust biotin- ligation methodologies in yeast and increased their sensitivity by creating a bespoke technique for downregulating endogenous biotinylation which we term ABOLISH (Auxin- induced BiOtin LIgase diminiSHing). We used the endoplasmic reticulum insertase complex (EMC) to demonstrate our approaches and uncover new substrates. To make these tools available for systematic probing of both stable and transient interactions, we generated five full-genome collections of strains in which every yeast protein is tagged with each of the tested biotinylation machineries; some on the background of the ABOLISH system. This comprehensive toolkit enables functional interactomics of the entire yeast proteome.

category: molecular biology


title: Parkinson mutations in DNAJC6 cause lipid defects and neurodegeneration that are rescued by Synj1

abstract: Recent evidence links dysfunctional lipid metabolism to the pathogenesis of Parkinsons disease, but the mechanisms are not resolved. Here, we created a new Drosophila knock-in model of DNAJC6/Auxilin and find that the pathogenic mutation causes synaptic dysfunction, neurological defects and neurodegeneration, as well as specific lipid metabolism alterations. In these mutants membrane lipids containing long-chain polyunsaturated fatty acids, including phosphatidylinositol lipid species that are key for synaptic vesicle recycling and organelle function are reduced. Overexpression of another protein mutated in Parkinsons disease, Synaptojanin-1, known to bind and synthesize specific phosphoinositides, strongly rescues the DNAJC6/Auxilin neuronal defects and neurodegeneration. Our work reveals a functional relation between two proteins mutated in Parkinsons disease and implicates deregulated phosphoinositide metabolism in the maintenance of neuronal integrity and neuronal survival in Parkinsonism.

category: cell biology


title: Competition between severing and tubulin induced repair of microtubules

abstract:
Cytoskeletal structures aid in cell polarization, motility, and intracellular transport. Their functions are predicated on the rapid turnover of cytoskeleton proteins, which is achieved by the coordinated effort of multiple regulatory proteins whose dynamics is not well understood. Recent in-vitro experiments have investigated mixtures of regulatory proteins and cytoskeletal filaments and have reported novel synergistic effects. For example, free tubulin can repair nanoscale damages of microtubules created by proteins that sever microtubules and, in doing so, affect their disassembly. Based on this observation, we propose a model for microtubule severing as a competition between the processes of damage spreading and tubulin-induced repair. Using theory and simulations, we demonstrate that this model is in quantitative agreement with in vitro experiments. We predict the existence of a critical tubulin concentration above which severing becomes rare but fast. Additionally, we find that microtubule length control via severing with tubulin-induced repair is hypersensitive to changes in the concentration of free tubulin and leads to a dramatically increased dynamic range of filament lengths in steady state. Our work describes how the concerted action of multiple microtubule associated proteins produces novel dynamical properties of these cytoskeletal filaments.

Significance StatementHow cytoskeleton-associated proteins coordinate rapid turnover of cytoskeletal structures is not well understood. Recent experiments suggest that microtubule severing is the result of two competing processes of damage spreading and tubulin-induced repair. We find that this process predicts the existence of a critical tubulin concentration above which severing becomes rare but fast, and additionally, a dramatically increased dynamic range of microtubule filament lengths at steady state. This work describes how combining multiple microtubule associated proteins can produce novel dynamical properties which are not seen when these proteins are studied in isolation.


title: Induction of Sis1 promotes fitness but not feedback in the heat shock response

abstract: Previously, we combined modeling and experiments to demonstrate that the heat shock response (HSR) functions as a negative feedback loop in which undefined chaperone clients activate the HSR by sequestering Hsp70, and subsequent induction of Hsp70 deactivates the response (Zheng et al., 2016; Krakowiak et al., 2018). Here, we formally define newly synthesized proteins (NSPs) as a major class of HSR activators and determine the role of Sis1, a co-chaperone of Hsp70, in HSR regulation. We develop and experimentally validate a new mathematical model that incorporates NSPs and Sis1. Unexpectedly, genetic decoupling and pulse-labeling experiments reveal that Sis1 induction promotes fitness during prolonged stress rather than providing negative feedback to the HSR. These results support an overall model in which NSPs signal the HSR by sequestering Sis1 and Hsp70, while induction of Hsp70 - but not Sis1 - attenuates the response.


title: Targeting Apollo-NADP+ to image NADPH generation in pancreatic beta-cell organelles

abstract: NADPH/NADP+ redox state supports numerous reactions related to cell growth and survival; yet the full impact is difficult to appreciate due to organelle compartmentalization of NADPH and NADP+. Our goal here was to target the Apollo-NADP+ sensor to measure glucose-stimulated NADPH production in pancreatic beta-cell organelles. To select the most pH stable version, we swapped various fluorescent proteins into the single colour sensor. We subsequently targeted the mTurquoise2 version of Apollo-NADP+ and confirmed activity in the cytoplasm, mitochondrial matrix, nucleus, and peroxisome. We subsequently measured the glucose- and glutamine-stimulated NADPH responses by single and dual colour imaging of the targeted sensors. Our data show NADPH production in the mitochondria prior to the cytoplasm, nucleus, and peroxisome. The glucose-stimulated responses were also blocked by pyruvate transport inhibitor consistent with mitochondrial metabolism determining NADPH generation in the other organelles. Overall, we developed multiple organelle-targeted Apollo-NADP+ sensors and revealed the dominance of beta-cell mitochondria in controlling NADPH/NADP+ redox state across organelles.


title: ATP6AP2-to-MMP14, a key pathway for osteoblast to osteocyte transition

abstract: Osteocytes, derived from osteoblast (OB)-lineage, occupy lacunae within bone matrix and exhibit unique morphology with dendrite-like projections to form an inter-connected network. Such a network is essential for osteocytes to monitor and orchestrate bone homeostasis. Thus, it is of considerable interest to investigate how osteocytes are formed and how they built the network. Here we provide evidence for ATP6AP2, an accessory subunit of V-ATPase, in OB-lineage cells to be critical for OB-to-osteocyte transition, and osteocyte distribution, maturation, and morphogenesis. Mice (ATP6AP2Ocn-Cre) that selectively deplete ATP6AP2 in OB-lineage cells results in altered osteocyte distribution and morphology, impaired osteocyte maturation and dendrite-like processes, increased osteocyte cell death and cortical woven bone formation. Further mechanistic studies identify MMP14 (matrix metalloproteinase-14) as a critical downstream of ATP6AP2 for osteocyte differentiation. ATP6AP2 interacts with MMP14 and promotes MMP14 surface distribution largely in immature- or osteoid-osteocytes, where this pathway regulates bone matrix remodeling and osteocyte differentiation. Expression of MMP14 into ATP6AP2 knock out OB-lineage cells in the mouse cortical bone could diminish the deficits in osteocyte maturation, dendrite-like process formation, and survival. These results thus demonstrate an un-recognized function of ATP6AP2 in promoting OB-to-osteocyte transition and uncover a pathway from ATP6AP2-to-MMP14 in osteocyte differentiation.


title: The climatic constrains of the historical global spread of mungbean

abstract: While the domestication process has been investigated in many crops, the detailed route of cultivar range expansion and factors governing this process received relatively little attention. Here using mungbean (Vigna radiata var. radiata) as a test case, we investigated the genomes of more than one thousand accessions to illustrate climatic adaptations role in dictating the unique cultivar spread routes in the face of continual human intervention. Despite the geographical proximity between South and Central Asia, mungbean cultivars first spread from South Asia to Southeast, East, and finally reached Central Asia. Combining evidence from demographic inference, climatic niche modeling, plant morphology, and records from ancient Chinese sources, we showed that the specific spread route was shaped by the unique combinations of climatic constraints and farmer practices across Asia, which imposed divergent selection favoring higher yield in the south but short-season and more drought-tolerant varieties in the north. Our results suggest that mungbean did not radiate from the domestication center as expected purely under human activity, but instead the spread route is highly constrained by climatic adaptation, echoing the idea that human commensals are more difficult to spread through the south-north axis of continents.


title: Chronic nicotine exposure alters sperm small RNA content in a C57BL/6J mouse model: Implications for multigenerational inheritance

abstract: Multigenerational inheritance is a non-genomic form of heritability characterized by altered phenotypes in the first generation born from the exposed parent. Multigenerational factors may account for inconsistencies and gaps in heritable nicotine addiction vulnerability. Our lab previously found that F1 offspring of male C57BL/6J mice chronically exposed to nicotine exhibited altered hippocampus functioning and related learning, nicotine-seeking, nicotine metabolism, and basal stress hormones. In an effort to identify germline mechanisms underlying these multigenerational phenotypes, the current study sequenced small RNA extracted from sperm of males chronically administered nicotine using our previously established exposure model. We identified 16 miRNAs whose expression in sperm was dysregulated by nicotine exposure. A literature review of previous research on these transcripts suggested an enrichment for regulation of psychological stress and learning. mRNAs predicted to be regulated by differentially expressed sperm small RNAs were further analyzed using biological enrichment analysis, which also supported enrichment of gene expression pathways involved in hippocampus-dependent learning. Our findings point to links between nicotine-exposed F0 sperm miRNA and altered F1 phenotypes in this multigenerational inheritance model. Specifically, differentially expressed F0 sperm miRNAs may regulate the previously observed changes in F1 learning and stress. These findings provide a valuable foundation for future functional validation of these hypotheses and characterization of mechanisms underlying male-line multigenerational inheritance.


title: Integrated Genomics Approaches Identify Mediators of Transcriptional and Epigenetic Responses to Afghan Desert Particulate Matter in Human Small Airway Epithelial Cells

abstract: New respiratory diseases in personnel deployed to Southwest Asia after September 11th raise major concerns about the impacts of airborne particulate matter. Although regulations exist, the knowledge about how particulates influence disease states is limited, precluding the appropriate recognition, prevention and treatment of deployment-related lung diseases. We applied two genomics assays, Precision Run-on sequencing (PRO-seq) and the assay for transposase accessible chromatin with sequencing (ATAC-seq), to characterize the small airway epithelial cell response to Afghan desert particulate matter (APM). In addition to cellular pathways that regulate susceptibility to further insults, such as cytochrome P450 or P53, we identified novel pathways associated with APM exposure, such as GRHL2 loss or TEAD3 activation. We further demonstrated that TEAD3 activation was unique to APM exposure despite similar inflammatory responses when compared with wood smoke particle (WSP) exposure. Our results establish the utility of an integrated genomics approach in characterizing responses to exposures and identify cellular and genomic targets for advanced investigation of the pathogenesis of deployment- related respiratory disease.


title: From defaults to databases: parameter and database choice dramatically impact the performance of metagenomic taxonomic classification tools

abstract: In metagenomic analyses of microbiomes, one of the first steps is usually the taxonomic classification of reads by comparison to a database of previously taxonomically classified genomes. While different studies comparing metagenomic taxonomic classification methods have determined that different tools are "best", there are two tools that have been used the most to-date: Kraken (k-mer based classification against a user-constructed database) and MetaPhlAn (classification by alignment to clade-specific marker genes), the latest versions of which are Kraken2 and MetaPhlAn 3, respectively. We found large discrepancies in both the proportion of reads that were classified as well as the number of species that were identified when we used both Kraken2 and MetaPhlAn 3 to classify reads within metagenomes from human-associated or environmental datasets. We then investigated which of these tools would give classifications closest to the real composition of metagenomic samples using a range of simulated and mock samples and examined the combined impact of tool-parameter-database choice on the taxonomic classifications given. This revealed that there may not be a one-size-fits-all "best" choice. While Kraken2 can achieve better overall performance, with higher precision, recall and F1 scores, as well as alpha- and beta-diversity measures closer to the known composition than MetaPhlAn 3, the computational resources required for this may be prohibitive for many researchers, and the default database and parameters should not be used. We therefore conclude that the best tool-parameter-database choice for a particular application depends on the scientific question of interest, which performance metric is most important for this question and the limit of available computational resources.


title: A reverse vaccinology approach identifies putative vaccination targets in the zoonotic nematode Ascaris.

abstract: Ascariasis is the most prevalent helminthic disease affecting both humans and pigs and is caused by the roundworms Ascaris lumbricoides and Ascaris suum. While preventive chemotherapy continues to be the most common control method, recent reports of anthelminthic resistance highlight the need for development of a vaccine against ascariasis. The aim of this study was to use a reverse vaccinology approach to identify potential vaccine candidates for Ascaris. Three Ascaris proteomes predicted from whole-genome sequences were analysed. Candidate proteins were identified using open-access bioinformatic tools (e.g. Vacceed, VaxiJen, Bepipred 2.0) which test for different characteristics such as sub-cellular location, T-cell and B-cell molecular binding, antigenicity, allergenicity and phylogenetic relationship with other nematode proteins. From over 100,000 protein sequences analysed, four transmembrane proteins were predicted to be non-allergen antigens and potential vaccine candidates. The four proteins are a Piezo protein, two voltage-dependent calcium channels and a protocadherin-like protein, are all expressed in either the muscle or ovaries of both Ascaris species, and all contained high affinity epitopes for T-cells and B-cells. The use of a reverse vaccinology approach allowed the prediction of four new potential vaccination targets against ascariasis in humans and pigs. These targets can now be further tested in in vitro and in vivo assays to prove efficacy in both pigs and humans.


title: Conformational fingerprinting of allosteric modulators in metabotropic glutamate receptor 2

abstract: Activation of G protein-coupled receptors (GPCRs) is an allosteric process. It involves conformational coupling between the orthosteric ligand binding site and the G protein binding site. Factors that bind at sites other than the orthosteric ligand binding site and alter this allosteric activation process are allosteric modulators and are important class of therapeutics. For many receptors, how modulation of signaling is represented at the structural level is unclear. Here, we developed FRET sensors to quantify receptor modulation at each of the three structural domains of metabotropic glutamate receptor 2 (mGluR2). We identified the conformational fingerprint for several allosteric modulators in live cells. This approach enabled us to derive a receptor-centric representation of allosteric modulation and to correlate structural modulation to the standard signaling modulation metrics. Single-molecule FRET analysis revealed that a NAM increases the occupancy of one of the intermediate states while a PAM increases the occupancy of the active state. Moreover, we found that the effect of allosteric modulators on the receptor dynamics is complex and depend on the orthosteric ligand. Collectively, our findings provide a structural mechanism of allosteric modulation in mGluR2 and suggest possible strategies for design of future modulators.


title: Metformin decreases Cyp26a1 to prevent hepatocarcinogenesis through down- regulating CD8+ T cells

abstract: Hepatocellular carcinoma (HCC) is a highly heterogeneous cancer, which limits the selectivity of prevention and treatment. Preclinical and clinical studies suggested that in patients with diabetes, prolonged use of metformin, the AMPK activator, was associated with a reduction of HCC incidence. This association promotes us to investigate the possible functions and mechanisms of metformin in HCC without diabetes backgrounds. Here, we found that several unique pathways that changed during chronic liver injury of Fah-/- mice, including glucose metabolic process and retinol metabolism. Further, metformin suppressed the tumor formation in chronic liver injury of Fah-/- mice. RNA sequencing, in vivo and in vitro experiments showed that metformin suppressed Cyp26a1 gene expression of hepatocyte. Moreover, the down-regulation of Cyp26a1 leads to the increased level of all-trans-retinoic acid (atRA), which could suppress the tumor formation in our model. On the other hand, flow multicolor analysis showed that the cell number and proportion of cancer promoting (pro-tumor) CD8+ T cells increased significantly during chronic liver injury in Fah-/- mice, and both metformin and atRA treatment could reduce the number and proportion of pro-tumor CD8+ T cells. We also found metformin decreased the Cyp26a1 expression through the AMPK/JNK/c-Jun pathway. In short, the association between the metformin and atRA may explain the commonness of their anti-tumor activities. Our findings highlight the importance of targeting the precancerous microenvironment for the prognosis, prevention and treatment of HCC.

category: cancer biology


title: Network analysis reveals a major role for 14q32 cluster miRNAs in determining transcriptional differences between IGHV-mutated and unmutated CLL

abstract: Tumour cells from patients with chronic lymphocytic leukaemia (CLL) can express unmutated (U-CLL) or mutated (M-CLL) immunoglobulin heavy chain (IGHV) genes with differing clinical behaviours, variable B cell receptor (BCR) signalling capacity and distinct transcriptional profiles. As it remains unclear to what extent these differences reflect the tumour cells innate pre/post germinal centre origin or their BCR signalling competence, we applied RNA sequencing, small RNA sequencing and DNA methylation array analysis to 38 CLL cases categorised into three groups by IGHV mutational status and BCR signalling capacity. We identified 492 mRNAs and 38 miRNAs differentially expressed between U-CLL and M-CLL, but only 9 mRNAs and 0 miRNAs associated with BCR competence within M-CLL. A significant proportion of the IGHV-associated miRNAs derived from chr14q32 clusters (14/38 (37%)), where all miRNAs were co-expressed with the MEG3 lncRNA, as part of the DLK1-DIO3 genomic imprinted region, a locus of known importance in the pathogenesis of other human tumours. Integrative in silico analysis of miRNA/mRNA data revealed pronounced regulatory potential for the 14q32 miRNAs, potentially accounting for up to 25% of the IGHV-related transcriptome signature. GAB1, a positive regulator of BCR signalling, was predicted to be regulated by five 14q32 miRNAs and we confirmed that two of these (miR-409-3p and miR-411-3p) significantly repressed activity of the GAB1 3UTR. Our analysis demonstrates a potential key role of the 14q32 miRNA locus in the regulation of CLL-related gene regulation.


title: Receptor-like kinases FER and SRK mediate Lotus japonicus freezing tolerance and climate adaptation

abstract: Many plant species have succeeded in colonizing a wide range of diverse climates through local adaptation, but the underlying molecular genetics remain obscure. We previously found that winter survival was a direct target of selection during colonization of Japan by the perennial legume Lotus japonicus and identified associated candidate genes. Here, we show that two of these, the FERONIA-receptor like kinase (LjFER) and a novel S-receptor-like kinase (LjSRK) are required for non-acclimated freezing tolerance and show haplotype-dependent cold-responsive expression. Our work demonstrates that recruiting a conserved growth regulator, FER, and a novel receptor-like kinase, SRK, into the set of cold-responsive genes has contributed to freezing tolerance and local climate adaptation in L. japonicus, offering new functional genetic insight into perennial herb evolution.

category: plant biology


title: miRNA408 and its encoded peptide, miPEP408, regulate arsenic stress response in Arabidopsis

abstract:
MicroRNAs (miRNAs) are small non-coding RNAs that play a central role in regulating various developmental and biological processes. The expression of miRNAs is differentially modulated in response to various stresses. Based on the recent findings, it has been shown that some of the pri-miRNAs encode small regulatory peptides, microRNA-encoded peptides (miPEP). miPEPs are reported to regulate the growth and development of plants by modulating corresponding miRNA expression; however, the role of these peptides in different stresses has not been explored yet. Here, we reported that pri-miR408 encodes a small peptide, miPEP408, that regulates the expression of miR408, its targets, and associated phenotype in Arabidopsis. Plants overexpressing miR408 showed severe sensitivity under low sulphur (LS), Arsenite As(III) and LS+As(III) stress, while miR408 mutant developed through the CRISPR/Cas9 approach showed tolerance. Transgenic lines showed phenotypic alteration and modulation in the expression of genes involved in the sulphur reduction pathway and affect sulphate and glutathione accumulation. Similar to miR408 overexpressing lines, the exogenous application of synthetic miPEP408 or miPEP408 overexpression led to sensitivity in plants under LS, As(III) and combined LS+As(III) stress compared to control. This study suggests the involvement of miR408 and miPEP408 in heavy metal and nutrient deficiency responses.

One-sentence summarymiR408 and peptide encoded by miR408, miPEP408, regulate arsenic stress and low sulphur responses in Arabidopsis.


title: Differential genetic variation underlying Ammonium and Nitrate responses in Arabidopsis thaliana

abstract:
Nitrogen is an essential element required for plant growth and productivity. Understanding the mechanisms and natural genetic variation underlying nitrogen use in plants will facilitate engineering plant nitrogen use to maximize crop productivity while minimizing environmental costs. To understand the scope of natural variation that may influence nitrogen use, we grew 1135 Arabidopsis thaliana natural genotypes on two nitrogen sources, nitrate and ammonium, and measured both developmental and defense metabolite traits. By using different environments and focused on multiple traits, we identified a wide array of different nitrogen responses. These responses are associated with a large number of genes, most of them not previously associated with nitrogen responses. Only a small portion of these genes appear to be shared between environments or traits while most of the detected genes are predominantly specific to a developmental or defense trait under a specific nitrogen source. Finally, by using a large population we were able to identify unique nitrogen responses, like preferring ammonium or nitrate, that appear to be generated by combinations of loci rather than a few large effect loci. This suggests that it may be possible to obtain novel phenotypes in complex nitrogen responses by manipulating sets of genes with small effects rather than solely focusing on large effect single gene manipulations.

One Sentence SummaryUsing a large collection of natural genotypes, and studying both developmental and metabolic responses, we found a large number of genes that are involved in the plants nitrogen response.


title: The localization of PHRAGMOPLAST ORIENTING KINESIN1 at the division site depends on two microtubule binding proteins TANGLED1 and AUXIN-INDUCED-IN-ROOT-CULTURES9 in Arabidopsis.

abstract:
Proper plant growth and development requires spatial coordination of cell divisions. Two unrelated microtubule-binding proteins, TANGLED1 (TAN1) and AUXIN-INDUCED-IN-ROOT-CULTURES9 (AIR9), are together required for normal growth and division-plane orientation in Arabidopsis. tan1 air9 double mutants have synthetic growth and division-plane orientation defects while single mutants lack obvious defects. Here we show that the division-site localized protein, PHRAGMOPLAST-ORIENTING-KINESIN1 (POK1), was aberrantly lost from the division site during metaphase and telophase in tan1 air9 mutants. Since TAN1 and POK1 interact via the first 132 amino acids of TAN1 (TAN11-132), we assessed its localization and function in the tan1 air9 double mutant. TAN11-132 rescued tan1 air9 mutant phenotypes and localized to the division site in telophase. However, replacing six amino-acid residues within TAN11-132 that disrupts POK1-TAN1 interaction in the yeast-two-hybrid system caused loss of both rescue and division-site localization of TAN11-132 in tan1 air9 mutants. Full-length TAN1 with the same alanine substitutions had defects in phragmoplast guidance and reduced TAN1 and POK1 localization at the division site but rescued most tan1 air9 mutant phenotypes. Together, these data suggest that TAN1 and AIR9 are required for POK1 localization, and yet unknown proteins may stabilize TAN1-POK1 interactions.

One sentence summarySpecific amino acids within TAN1 are required for its correct localization and function partially through interaction with POK1; both TAN1 and AIR9 mediate POK1 division site localization.


title: How do Protein Contaminants Influence DDA and DIA Proteomics

abstract: Mass spectrometry-based proteomics is constantly challenged by the presence of contaminant background signals. In particular, protein contaminants from reagents and sample handling are often abundant and impossible to avoid. For data-dependent acquisition (DDA) proteomics, exclusion list can be used to reduce the influence of protein contaminants. However, protein contamination has not been evaluated and is rarely addressed in data-independent acquisition (DIA). How protein contaminants influence proteomics data is also unclear. In this study, we established the protein contaminant FASTA and spectral libraries that are applicable to all proteomic workflows and evaluated the impact of protein contaminants on both DDA and DIA proteomics. We demonstrated that including our contaminant libraries can reduce false discoveries and increase protein identifications, without influencing the quantification accuracy in various proteomic software platforms. With the pressing need to standardize proteomic workflow in the research community, we highly recommend including our contaminant FASTA and spectral libraries in all bottom-up proteomics workflow. Our contaminant libraries and a step-by-step tutorial to incorporate these libraries in different DDA and DIA data analysis platforms can be a valuable resource for proteomics researchers, which are freely accessible at https://github.com/HaoGroup-ProtContLib.

category: systems biology


title: Cortical magnification underlies differences across but not around the visual field

abstract: Performance changes dramatically both across (eccentricity) and around (polar angle) the visual field. All neurophysiological and virtually all behavioral studies of cortical magnification investigate eccentricity effects without considering polar angle. But, does cortical magnification underlie differences around the visual field? We magnified stimulus size according to eccentricity and polar angle to equate their cortical representation. We found that magnifying the stimulus eliminates differences across -but not around- the visual field.


title: Equilibrative Nucleoside Transporter 3 is an IFN-stimulated Gene that Facilitates Viral Genome Release

abstract: An increasing body of evidence emphasizes the role of metabolic reprogramming in immune cells to fight off infections. However, little is known about the regulation of metabolite transporters that facilitate and support metabolic demands. In this study, we found that equilibrative nucleoside transporter 3 (ENT3) expression is part of the innate immune response, and is rapidly upregulated upon bacterial and viral infection. The transcription of ENT3 is directly under the regulation of IFN-induced signaling, positioning this metabolite transporter as an Interferon-stimulated gene (ISG). Moreover, we unveil that several viruses, including SARS-CoV2, require ENT3 to facilitate their entry into the cytoplasm. The removal or suppression of ENT3 expression is sufficient to significantly decrease viral replication in vitro and in vivo.


title: Light 'em up: efficient screening of gold grids in cryo-EM

abstract: Transmission electron cryo-microscopy (cryo-EM) allows for obtaining 3D structural information by imaging macromolecules embedded in thin layers of amorphous ice. To obtain high-resolution structural information, samples need to be thin to minimize inelastic scattering which blurs images. During data collection sessions, time spent on finding areas on the cryo-EM grid with optimal ice thickness should be minimized as imaging time on high-end Transmission Electron Microscope TEM systems is costly. Recently, grids covered with thin gold films have become popular due to their stability and reduced beam-induced motion of the sample. Gold foil grids have substantially different densities between the gold foil and ice, effectively resulting in the loss of dynamic range between thin and thick regions of ice, making it challenging to find areas with suitable ice thickness efficiently during grid screening and thus increase expensive imaging time. Here, an energy filter-based plasmon imaging is presented as a fast and easy method for grid screening of the gold grids.


title: Rac rather than Rho drives the early transcriptional response to extracellular matrix stiffness and mediates repression of ATF3

abstract: The Rho family GTPases, Rac and Rho, play critical roles in transmitting mechanical information contained within the extracellular matrix (ECM) to the cell. Rac and Rho have well described roles in regulating stiffness-dependent actin remodeling, proliferation and motility. However, much less is known about the relative roles of these GTPases in stiffness-dependent transcription, particularly at the genome-wide level. Here, we selectively inhibited Rac and Rho in mouse embryonic fibroblasts cultured on deformable substrata and used RNA sequencing to elucidate and compare the contribution of these GTPases to the early transcriptional response to ECM stiffness. Surprisingly, we found that Rac activation is almost exclusively responsible for the initial transcriptional response to ECM stiffness. We also identified Activating Transcription Factor 3 (ATF3) as a major target of stiffness/Rac signaling and showed that ATF3 repression by ECM stiffness connects the stiffness-dependent activation of Rac to the induction of cyclin D1.


title: EGFR inhibition stabilizes cardiomyocyte cohesion in a murine arrhythmogenic cardiomyopathy model

abstract:
The epidermal growth factor receptor (EGFR) is a ubiquitously expressed receptor tyrosine kinase, which is dysregulated in several kinds of cancer and heart diseases. Arrhythmogenic cardiomyopathy (AC) is a familial heart disease in part caused by impaired desmosome turnover. Thus, stabilization of desmosome integrity, may provide potential new treatment options. Desmosomes, apart from cellular cohesion, provide the structural framework of a signaling hub. Here, we investigated the role of EGFR inhibition on cardiomyocyte cohesion under physiological and pathophysiological conditions using the murine plakoglobin knockout AC model, in which EGFR was upregulated. EGFR inhibition led to enhanced cardiomyocyte cohesion. Immunoprecipitation showed an interaction of EGFR and desmoglein 2 (DSG2). Immunostaining and AFM revealed enhanced DSG2 localization and binding at cell borders upon EGFR inhibition. Enhanced area composita length and desmosome assembly were observed upon EGFR inhibition, confirmed by enhanced DSG2 and desmoplakin (DP) recruitment to cell borders. Thus, inhibiting EGFR, thereby stabilizing desmosome integrity, might provide new treatment options for AC.

SummaryShoykhet et al. show EGFR inhibition-enhanced cardiomyocyte cohesion. This positive adhesiotropy is achieved via an increase in desmosomal assembly and DP/DSG2 localization at cell borders and is the first step towards a novel therapeutic approach for arrhythmogenic cardiomyopathy.


title: EGFR inhibition leads to enhanced desmosome assembly and cohesion in cardiomyocytes


title: Purification of mitochondria from skeletal muscle tissue for transcriptomic analyses reveals localisation of nuclear-encoded non-coding RNAs

abstract: Mitochondria are central to cellular function, particularly in metabolically active tissues such as skeletal muscle. Non-coding RNAs (ncRNAs) typically localise within the nucleus and cytosol but may also translocate to subcellular compartments such as mitochondria. We aimed to investigate the nuclear-encoded ncRNAs that localise within the mitochondria of skeletal muscle cells and tissue. Intact mitochondria were isolated via immunoprecipitation and an enzymatic digestion approach was optimised to remove transcripts located exterior to mitochondria, making it amenable for high-throughput transcriptomic sequencing. Small-RNA sequencing libraries were successfully constructed from as little as 1.8ng mitochondrial RNA input. Small-RNA and whole transcriptome sequencing of mitochondria reveals the enrichment of over 200 miRNAs and 200 lncRNAs that have not previously been observed within skeletal muscle mitochondria. In summary, we describe a novel, powerful sequencing approach applicable to animal and human tissues and cells that reveals the unexpected diversity of nuclear-encoded ncRNA transcripts localised within skeletal muscle mitochondria.


title: Wastewater microorganisms impact microbial diversity and important ecological functions of stream periphyton

abstract:
Wastewater treatment plant effluents can impact microbial communities in receiving streams. However, little is known about the role of microorganisms in wastewater as opposed to other wastewater constituents, such as nutrients and micropollutants. We aimed therefore at determining the impact of wastewater microorganisms on the microbial diversity and function of periphyton, key microbial communities in streams. Periphyton was grown in flow-through channels that were continuously alimented with a mixture of stream water and unfiltered or ultra-filtered wastewater. Impacts were assessed on periphyton biomass, activities and tolerance to micropollutants, as well as on microbial diversity. Our results showed that wastewater microorganisms colonized periphyton and modified its community composition, resulting for instance in an increased abundance of Chloroflexi and a decreased abundance of diatoms and green algae. This led to shifts towards heterotrophy, as suggested by the changes in nutrient stoichiometry and the increased mineralization potential of carbon substrates. An increased tolerance towards micropollutants was only found for periphyton exposed to unfiltered wastewater but not to ultra-filtered wastewater, suggesting that wastewater microorganisms were responsible for this increased tolerance. Overall, our results highlight the need to consider the role of wastewater microorganisms when studying potential impacts of wastewater on the receiving water body.

Environmental implicationThe present study investigates the impact of wastewater microorganisms on periphyton, i.e. communities forming the microbial skin of streambeds. We were able to disentangle specific effects of wastewater microorganisms in the context of the complex wastewater matrix. Indeed, wastewater microorganisms induced strong changes in periphyton community composition and function, suggesting the need to consider wastewater microbial communities as a stressor per se, similarly to, e.g., nutrients and micropollutants. Moreover, since periphyton is at the basis of the food web in streams, these changes may have consequences for higher trophic levels.


title: Defining the age-dependent and tissue-specific circadian transcriptome in male mice

abstract:
Cellular circadian clocks direct a daily transcriptional program that supports homeostasis and resilience. Emerging evidence supports age-associated changes in circadian functions. To define age-dependent changes at the systems level, we profiled the circadian transcriptome in the hypothalamus, lung, heart, kidney, skeletal muscle, and adrenal gland in 3 age groups. We found age-dependent and tissue-specific clock output changes. Aging reduced the number of rhythmically expressed genes (REGs), indicative of weakened circadian control. Many genes gained rhythmicity in old tissues, reflecting an adaptive response. REGs were enriched for the hallmarks of aging, adding a new dimension to our understanding of aging. Differential gene expression analysis found that there were temporally distinct clusters of genes in tissue-specific manner. Increased daily gene expression variability is a common feature of aged tissues. This novel analysis extends the landscape of the understanding of aging and highlights the impact of aging on circadian clock function and temporal changes in gene expression.

HIGHLIGHTS- Rhythmically expressed genes (REGs) in Young, but not Old mice, are enriched for the aging hallmarks across all tissues.
- The numbers of REGs decline across all tissues with age implicating the circadian clock in altered homeostasis.
- Age- and tissue-specific differentially expressed genes (DEGs) cluster at specific times of the day.
- Increase in gene expression variability over a day is a common feature of aging tissues.


title: HSP90.6 is involved in grain filling via carbon and nitrogen metabolism in maize.

abstract:
Carbon and nitrogen are the two most abundant nutrients in all living things, and their metabolism maintains normal plant growth. However, the molecular mechanism underlying carbon and nitrogen metabolism remains largely unknown. Here, we found that HSP90.6 is involved in the metabolism of carbon and nitrogen. We performed gene cloning and functional characterization of a maize EMS mutant ehsp90.6, whose kernels were small. HSP90.6 encodes heat shock protein 90.6, which has a single-amino acid mutation within its HATPase_c domain. Transcriptome profiling showed that the expression of amino acid biosynthesis- and carbon metabolism-related genes was significantly downregulated in hsp90.6. HSP90.6 is involved in the 26S proteasome degradation pathway, which affects nitrogen recycling to regulate amino acid synthesis; this occurs by interactions between HSP90.6 and the 26S proteasome subunits RPN6 and PBD2 (PRC2). The loss of HSP90.6 significantly reduced the activity of the 26S proteasome, resulting in the accumulation of ubiquitinated proteins and defects in nitrogen recycling. Moreover, HSP90.6 interacted with the 14-3-3 protein GF14-6 to participate in carbon metabolism. Together, these findings revealed that HSP90.6 regulates nutrient metabolism in maize seeds by affecting 26S proteasome-mediated nitrogen recycling and GF14-6-mediated carbon metabolism.

One sentence summaryHSP90.6 is involved in nutrient metabolism via 26S proteasome-mediated protein degradation to promote nitrogen recycling and GF14-6 protein-mediated carbon metabolism.

The author responsible for the distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (https://academic.oup.com/plcell/pages/General-Instructions) is Weibin Song (songwb@cau.edu.cn).

HighlightsO_LIHATPase_c is necessary for HSP90.6 to regulate maize kernel development.
C_LIO_LIHSP90.6 is involved in nitrogen recycling via the 26S proteasome degradation pathway.
C_LIO_LIHSP90.6 interacts with the 14-3-3 protein GF14-6 to affect carbon metabolism.
C_LI

IN A NUTSHELLO_ST_ABSBackgroundC_ST_ABSSeeds are the main harvested organs of maize. Understanding the regulatory mechanism of grain filling is helpful to cultivate high-quality and high-yield maize. In the past few years, the regulatory network of grain filling has been explored through multiple means, including transcriptomic, proteomic and functional genomic techniques. Many genes that control grain filling through different mechanisms have been cloned, such as CTLP1 (Choline Transporter-like Protein 1), OS1 (Opaque Endosperm and Small Germ 1), and MN6 (Miniature Seed6). To identify new genes involved in maize grain filling, ethyl methanesulfonate (EMS) was used to induce mutations, and the ehsp90.6 mutant, which exhibited abnormal kernel development, was isolated by bulked segregant analysis RNA sequencing (BSR).

QuestionWhy does the single-amino acid mutation of HSP90.6 affect grain size, and how does the loss of HSP90.6 affect grain filling?

FindingsA single-amino acid mutant (ehsp90.6) and knockout mutant (hsp90.6) were obtained. We found that HSP90-6 was involved in the regulation of maize grain filling. A single-single amino acid mutation in the HATPase_c domain reduced the ATPase activity of HSP90.6, resulting in smaller grains. The functional loss of HSP90.6 resulted in the expression of amino acid biosynthesis- and carbon metabolism-related genes being significantly downregulated in hsp90.6. We indicated that HSP90.6 is involved in the 26S proteasome degradation pathway, which affects nitrogen recycling to regulate amino acid synthesis by interacting with the 26S proteasome subunits RPN6 and PBD2 (PRC2). Moreover, HSP90.6 was found to interact with the 14-3-3 protein GF14-6 to participate in carbon metabolism.

Next stepsTo further verify that the interaction between HSP90.6 and 26S proteasome subunits and GF14-6 affects grain filling, knockout validation of RPN6, PBD2 (PRC2) and GF14-6 will be performed. In addition, since GF14-6 interacts with the phosphorylated proteins, we will determine the phosphorylation site of HSP90.6. Due to the important role of HSP90 family proteins in plant development, there are other regulatory pathways that need to be explored.


title: Age-related changes of deep-brain neurophysiological activity

abstract: Cognitive decline with age is associated with brain atrophy and reduced brain activations, but the underlying neurophysiological mechanisms are unclear, especially in deeper brain structures primarily affected by healthy aging or neurodegenerative processes. Here, we characterize time-resolved, resting-state magnetoencephalography activity of the hippocampus and subcortical brain regions in a large cohort of healthy young and older volunteers from the Cam-CAN open repository. The data show age-related changes in both rhythmic and arrhythmic signal strength and temporal variability in multiple deeper brain regions, including the hippocampus, striatum, and thalamus. We observe a slowing of neural activity in deeper brain regions, which echoes previous reports of cortical slowing. We also report reduced occipito-parietal alpha peak associated with increased theta-band activity and signal variability in the hippocampus, an effect that may reflect compensatory processes as theta activity and variability were more strongly expressed when cognitive performances are preserved. Overall, this study advances the understanding of the biological nature of inter-individual variability in aging. The data provide new insight of how hippocampus and subcortical neurophysiological activity evolve with biological age, and highlight frequency-specific effects associated with cognitive decline vs. cognitive maintenance.


title: Robust cone-mediated signaling persists late into rod photoreceptor degeneration

abstract: Rod photoreceptor degeneration causes deterioration in the morphology and physiology of cone photoreceptors along with changes in retinal circuits. These changes could diminish visual signaling at cone-mediated light levels, thereby limiting the efficacy of treatments such as gene therapy for rescuing normal, cone-mediated vision. However, the impact of progressive rod death on cone-mediated signaling remains unclear. A mouse model of rod degeneration was used to investigate the fidelity of retinal ganglion cell (RGC) signaling throughout disease progression. Despite clear deterioration of cone morphology with rod death, cone-mediated signaling among RGCs remained surprisingly robust: spatiotemporal receptive fields changed little and the mutual information between stimuli and spiking responses was relatively constant. This relative stability held until nearly all rods had died and cones had completely lost well-formed outer segments. Interestingly, RGC information rates were higher and more stable for natural movies than checkerboard noise as degeneration progressed. The main change in RGC responses with photoreceptor degeneration was a decrease in response gain. These results suggest that gene therapies for rod degenerative diseases are likely to successfully prolong cone-mediated vision even if there are changes to cone morphology and density.


title: Induction of neutralizing antibodies against SARS-CoV-2 variants by a multivalent mRNA-lipid nanoparticle vaccine encoding SARS-CoV-2/SARS-CoV Spike protein receptor-binding domains

abstract: To address the need for multivalent vaccines against Coronaviridae that can be rapidly developed and manufactured, we compared antibody responses against SARS-CoV, SARS-CoV-2, and several variants of concern in mice immunized with mRNA-lipid nanoparticle vaccines encoding homodimers or heterodimers of SARS-CoV/SARS-CoV-2 receptor-binding domains. All vaccine constructs induced robust anti-viral antibody responses, and the heterodimeric vaccine elicited an IgG response capable of cross-neutralizing SARS-CoV, SARS-CoV-2 Wuhan-Hu-1, B.1.351 (beta), and B.1.617.2 (delta) variants.


title: Diet induced obesity and type 2 diabetes drives exacerbated sex-associated disease profiles in K18-hACE2-mice challenged with SARS-CoV-2

abstract: SARS-CoV-2 infection results in wide-ranging disease manifestation from asymptomatic to potentially lethal. Infection poses an increased threat of severity to at-risk populations including those with hypertension, diabetes, and obesity. Type 2 Diabetes (T2DM), is characterized, in part, by insulin insensitivity and impaired glucose regulation. T2DM patients have increased disease severity and poorer outcomes with COVID-19. We utilized the diet-induced obesity (DIO) model of Type 2 Diabetes in SARS-CoV-2-susceptible K18-hACE2 transgenic mice to better understand the obesity co-morbidity. Female DIO, but not male DIO mice challenged with SARS-CoV-2 were observed to have shortened time to morbidity compared to normal diet mice. Increase in susceptibility to SARS-CoV2 in female DIO was associated with increased total viral RNA burden compared to male mice. RNAseq analysis was performed on the lungs of non-challenged, challenged, females, males, of either normal diet or DIO cohorts to determine the disease specific transcriptional profiles. DIO female mice had more total activated genes than normal diet mice after challenge; however, male mice experienced a decrease. GO term analysis revealed the DIO condition increased interferon response signatures and interferon gamma production following challenge. Male challenged mice had robust expression of antibody-related genes suggesting antibody producing cell localization in the lung. DIO reduced antibody gene expression in challenged males. Collectively this study establishes a preclinical T2DM/obesity co-morbidity model of COVID-19 in mice where we observed sex and diet specific responses that begin to explain the effects of obesity and diabetes on COVID-19 disease.


title: Removal of Clinically Relevant SARS-CoV-2 Variants by An Affinity Resin Containing Galanthus nivalis Agglutinin

abstract: The Coronavirus-19 (COVID-19) pandemic due to the SARS-CoV-2 virus has now exceeded two years in duration. The pandemic has been characterized by the development of a succession of variants containing mutations in the spike protein affecting infectiousness, virulence and efficacy of vaccines and monoclonal antibodies. Resistance to vaccination and limitations in the current treatments available require the ongoing development of therapies especially for those with severe disease. The plant lectin Galanthus nivalis binds to mannose structures in the viral envelope. We hypothesized that viral binding should be unaffected by spike protein mutations. Known concentrations of seven clinically relevant SARS-CoV-2 variants were spiked in medium and passed three times over columns containing 1 gm of GNA affinity resin. Percent decrease in viral titer was compared with a control sample. Viral capture efficiency was found to range from 53 to 89% for all variants. Extrapolation indicated that an adult Aethlon Hemopurifier(R) would have more than sufficient binding capacity for viral loads observed in adult patients with severe COVID-19 infection.


title: Drosophila Keap1 oxidative/xenobiotic response factor interacts with B-type lamin to regulate nuclear lamina and heterochromatin

abstract: The essential function of the Keap1-Nrf2 pathway in mediating transcriptional response to xenobiotic and oxidative stimuli has been well established. However, the mechanisms whereby Keap1 and Nrf2 regulate developmental genes remains unclear. We hypothesized that Drosophila Keap1 (dKeap1) and Nrf2 (CncC) proteins regulate transcription through controlling high-order chromatin structure. Here, we describe evidence supporting that dKeap1 can regulate chromatin through interaction with lamin, the intermediate filament proteins that form nuclear lamina and organize the overall chromatin architecture. dKeap1 and lamin Dm0, the B-type lamin in Drosophila, interact with each other and form complexes in the nucleus. Overexpression of dKeap1 resulted in a redistribution of lamin Dm0 to the intra-nuclear area and consistently, caused a spreading of the heterochromatin marker H3K9me2 from the pericentromeric region to chromosome arms. Overexpression of dKeap1 fusion proteins in the dKeap1 null background significantly disrupted the nuclear lamina morphology, indicating that dKeap1 is required for the maintenance of a normal nuclear lamina. Knock down of dKeap1 partially rescued the lethality caused by lamin Dm0 overexpression, suggesting that dKeap1 and lamin Dm0 function in the same pathway during development. Taken together, these results support a model where dKeap1 regulates chromatin structure and developmental transcription through interaction with lamin proteins, revealing a novel epigenetic function of the Keap1 oxidative/xenobiotic response factor.


title: How to measure response diversity

abstract: Taxonomic diversity metrics are, at best, proxies for the inter- and intraspecific trait differences that mechanistically underlie the insurance effect of biodiversity--that diversity enhances and stabilises aggregate ecosystem properties. In theory, variation in functional (e.g., biomass) responses to environmental change can buffer and maintain ecosystem functioning. This variation, termed response diversity, is therefore a potentially critical determinant of ecological stability. However, response diversity has yet to be widely quantified, possibly due to difficulties in its measurement. Even when it has been measured, approaches have varied. Here, we review methods for measuring response diversity and propose a methodological framework for quantifying response diversity from experimental or observational data, which can be practically applied in lab and field settings across a range of taxa. Our approach, which is based on environment-dependent functional responses to any biotic or abiotic environmental variable, is conceptually simple and robust to any form of environmental response, including nonlinear responses. By capturing even subtle inter- or intraspecific variation in environmental responses, and environment-dependencies in response diversity, we hope this framework will motivate tests of the diversity-stability relationship from a new perspective, and provide an approach for mapping, monitoring, and protecting this critical element of biodiversity.


title: Regulated use of alternative Transcription Start Sites controls the production of cytosolic or mitochondrial forms of branched-chain aminotransferase in Kluyveromyces marxianus

abstract: Following a whole genome duplication (WGD) event approximately 100 million years ago, the yeast lineage from which the model Saccharomyces cerevisiae derives maintained two copies of genes where it was necessary to synthesise proteoforms with different sub-cellular localisation. In contrast, yeasts that did not undergo the WGD event have a single gene that must encode both proteoforms. We adopted an integrated in silico and experimental approach to study how this is achieved with BAT1, a gene that encodes mitochondrial and cytosolic forms of a branched chain aminotransferase (BCAT) in pre-WGD yeast such as Kluyveromyces marxianus. We determined that condition-specific regulation of alternative transcription sites gives rise to mRNA isoforms that differ at the 5end and that, when decoded, generate a mitochondrial or cytosolic proteoform. Furthermore, targeted mutants lacking specific transcription factors were generated to establish how this differentiation was regulated. As in S. cerevisiae, Gcn4 and Leu3 activated expression of the mRNA encoding the mitochondrial proteoform under conditions when branched chain amino acid synthesis was required. Unlike S. cerevisiae, however, K. marxianus lacked tight regulation of the mRNA encoding the cytosolic proteoform supporting the hypothesis that maintaining paralogous genes in post-WGD yeasts facilitated development of more sophisticated expression control mechanisms.


title: AnkG-Neurofascin complex structure reveals binding mechanisms required for integrity of the AIS

abstract: The axon initial segment (AIS) has characteristically clustering of voltage-gated sodium channels (Nav), cell adhesion molecule Neurofascin (Nfasc), and neuronal scaffold protein Ankyrin-G (AnkG) in neurons, which facilitate generation of action potential and maintenance of axonal polarity. However, the mechanisms underlying AIS assembly and maintenance remain poorly understood. Here we report the high-resolution crystal structure of the AnkG in complex with a fragment from Nfasc cytoplasmic tail that shows, in conjunction with binding affinity assays, the molecular basis of AnkG-Nfasc binding. We confirm AnkG interacts with the FIGQY motif in Nfasc, and identify another region required for their high affinity binding. Structural analysis revealed that ANK repeats form four hydrophobic or hydrophilic layers in the AnkG inner groove that coordinate interactions with Nfasc. Moreover, disruption of the AnkG-Nfasc complex abolishes Nfasc enrichment at the AIS in hippocampal neurons. Finally, structural and biochemical analysis indicated that L1 syndrome-associated mutations in L1CAM compromise binding with ankyrins. These results define the mechanisms underlying AnkG-Nfasc complex formation and show that AnkG-dependent clustering of Nfasc is required for AIS integrity.

category: biochemistry