title: Unbreakable DNA tension probes show that cell adhesion receptors detect the molecular force-extension curve of their ligands

abstract: Integrin receptors transduce the mechanical properties of the extracellular matrix. Past studies using DNA probes showed that integrins sense the magnitude of ligand forces with pN resolution. An open question is whether integrin receptors also sense the force-extension trajectory of their ligands. The challenge in addressing this question pertains to the lack of molecular probes that can control force-extension trajectories independently of force magnitude. To address this limitation, we synthesized two reversible DNA probes that fold with identical self-complementary domains but with different topologies. Thus, these probes unfold at the same steady-state force magnitude but following different kinetic pathways to reach the fully extended ssDNA state. Hairpin-like probes unzip with a low barrier of 14 pN while the pseudo-knot-like probes shear at 59 pN. Confirming that we had created probes with different barriers of unfolding, we quantified platelet integrin forces and measured 50-fold more tension signal with the unzipping probes over the shearing probes. In contrast, fibroblasts opened both probes to similar levels indicating more static forces. Surprisingly, fibroblast mechanotransduction markers, such as YAP levels, fibronectin production, actin organization, and integrin activation were significantly elevated on unzipping probes. This demonstrates that integrin receptors within focal adhesions sense the molecular force-extension profile of their ligands and not only the magnitude of equilibrium mechanical resistance.


title: Multistate structures of the MLL1-WRAD complex bound to H2B-ubiquitinated nucleosome

abstract:
The human Mixed Lineage Leukemia-1 (MLL1) complex orchestrates methylation of histone H3K4 to promote transcription and is stimulated by monoubiquitination of histone H2B. Recent structures of the MLL1-WRAD core complex, which comprises the MLL1 methyltransferase, WDR5, RbBp5, Ash2L, and DPY-30, have revealed variation in the docking of MLL1-WRAD on nucleosomes and left ambiguous portions of Ash2L and the position of DPY30. We used an integrated approach combining cryo-electron microscopy and mass spectrometry-crosslinking to determine structures of the MLL1-WRAD complex bound to ubiquitinated nucleosomes containing the Ash2L intrinsically disordered region (IDR), SPRY insertion region, Sdc1-DPY30 interacting region (SDI-motif), and the DPY30 dimer. We resolved three additional states of MLL1-WRAD lacking one or more subunits, which may reflect different steps in the assembly of MLL1-WRAD. The subunits in all four states are positioned on the nucleosome in manner that is similar to a previous structure of MLL1-WRAD bound to ubiquitinated nucleosome, but that differs from structures with unmodified nucleosomes, suggesting that H2B-ubiquitin favors assembly of the active complex. Our results provide a more complete picture of MLL1-WRAD and the role of ubiquitin in promoting formation of the active methyltransferase complex.

SignificanceThe Mixed Lineage Leukemia-1 (MLL1) complex plays a role in activating transcription by methylating lysine 4 in histone H3, a reaction that is stimulated by the presence of ubiquitin conjugated to histone H2B. Recent structures of the core MLL1 complex, termed MLL1-WRAD, have revealed the existence of multiple docking states and have also left ambiguous portions of the structure. Here we combine mass spectrometry-cross linking with cryo-EM to model additional regions of the MLL1-WRAD complex and identify a series of states that light on complex assembly and the role that ubiquitin plays in orienting MLL1-WRAD on nucleosomes.

category: biophysics


title: Older circuits get richer: neurogenesis timeline shapes hubness and neurogenetic profiles of the adult human brain

abstract: Understanding the architectural principle that shapes the topology of the human connectome at its multiple spatial scales is a major challenge for systems neuroscience. This would provide key fundamental principles and a theory for browsing brains networks, to ultimately generate hypothesis and approach to which extent key structures might impact different brain pathologies. In this work, we propose the hypothesis that the centrality of the different brain nodes in the human connectome is a product of their embryogenic age, and accordingly, early-born nodes should display higher hubness, and viceversa for late-born nodes. We tested our hypothesis by identifying and segmenting eighteen macroregions with a well-known embryogenic age, over which we calculated nodes centrality in the structural and functional networks at different spatial resolutions. First, nodes structural centrality correlated with their embryogenic age, fully confirming our working hypothesis. However, at the functional level, distinct trends were found at different resolutions. Secondly, the difference in embryonic age between nodes inversely correlated with the probability of existence and the weights of the links. This indicated the presence of a temporal developmental gradient that shapes connectivity and where nodes connect more to nodes with a similar age. Finally, brain transcriptomic analysis revealed high association between embryonic age, structural-functional centrality and the expression of genes related to nervous system development, synapse regulation, and human neurological diseases. Overall, these results support the hypothesis that the embryogenic age of brain regions shapes the topology of adult brain networks. Our results show two key principles, "preferential age attachment" and "older gets richer" on the wiring of the human brain, thus shedding new light on the relationship between brain development, transcriptomics, node centrality, and neurological diseases.

category: neuroscience


title: CCR4 and CCR7 differentially regulate thymocyte subset localization with distinct outcomes for central tolerance

abstract: Developing thymocytes must efficiently enter the medulla to test if they are autoreactive to diverse self-peptides displayed by thymic antigen presenting cells (APCs). While CCR7 is known to promote medullary entry and negative selection, we previously found that CCR4 contributes to these processes, raising the question of whether CCR4 and CCR7 play redundant roles in central tolerance. Here, synchronized positive selection assays and 2-photon microscopy reveal that CCR4 and CCR7 promote medullary accumulation and negative selection of distinct post-positive selection thymocyte subsets. CCR4 is upregulated within hours of positive selection and promotes medullary entry and clonal deletion of immature post-positive selection thymocytes. In contrast, CCR7 promotes medullary entry and negative selection of mature thymocytes. Our data uncouple CCR7 expression from robust medullary accumulation. Furthermore, CCR4 ligands are expressed by activated thymic APCs, and CCR4 promotes tolerance to self-antigens expressed by these cells. These data support a model in which CCR4 and CCR7 contribute to early and late phases of negative selection, respectively.

category: immunology


title: Identification and functional characterization of two heme uptake systems required for virulence of the fish pathogen Flavobacterium psychrophilum

abstract: Bacterial pathogens have a critical impact on aquaculture, a sector that accounts for half of the human fish consumption. Flavobacterium psychrophilum (phylum Bacteroidetes) is responsible for bacterial cold-water disease in salmonids worldwide. The molecular factors involved in host invasion, colonization and hemorrhagic septicaemia are mostly unknown. In this study, we identified two new TonB-dependent receptors, HfpR and BfpR, that are required for adaptation to iron conditions encountered during infection and for virulence in rainbow trout. Transcriptional analyses revealed that their expression is tightly controlled and upregulated under specific iron sources and concentrations. Characterization of deletion mutants showed that they act without redundancy: BfpR is required for optimal growth in the presence of high hemoglobin level, while HfpR confers the capacity to acquire nutrient iron from heme or hemoglobin under iron scarcity. The gene hfpY, co-transcribed with hfpR, encodes a protein related to the HmuY family. We demonstrated that HfpY binds heme and contributes significantly to host colonization and disease severity. Overall, these results are consistent with a model in which both BfpR and Hfp systems promote heme uptake and respond to distinct signals to adapt iron acquisition to the different stages of pathogenesis. Our findings give insight into the molecular basis of pathogenicity of a serious pathogen belonging to the understudied family Flavobacteriaceae and point to the newly identified heme receptors as promising targets for antibacterial development.

category: microbiology


title: Two functionally distinct heme/iron transport systems are virulence determinants of the fish pathogen Flavobacterium psychrophilum


title: Modulation of the gut microbiome by novel synthetic glycans for the production of propionate and the reduction of cardiometabolic risk factors

abstract:
BackgroundIncreasing evidence indicates that an altered gut microbiome participates in the development of cardiometabolic syndrome and associated risk factors, such as insulin resistance, dyslipidemia, and obesity, and that targeting the gut microbiome is a promising strategy to lower the risk for cardiometabolic diseases. Part of this reduction is mediated by specific metabolites generated by the gut microbiome. Propionate, a short-chain fatty acid (SCFA) produced from dietary glycans by certain gut microbes is known to exert multiple beneficial metabolic effects. Here, we identify KB39, a novel gut microbiome-targeting synthetic glycan selected for its strong propionate-producing capacity, and demonstrate its effects in vivo to reduce cardiometabolic disease using western diet-fed LDL receptor knock-out mice.

MethodsEx vivo fermentation screening of a large library of synthetic glycan ensembles was performed using gut microbiome communities from healthy subjects and overweight patients with type 2 diabetes. A synthetic glycan identified for its high propionate-producing capacity (KB39) was then tested in vivo for effects on systemic, blood and cecal metabolic parameters in Ldlr-/- mice fed a western diet.

ResultsEx vivo screening of ~600 synthetic glycans using human gut microbiota from healthy subjects and patients with type 2 diabetes identified a novel glycan (KB39) with high propionate-producing capacity that increased propionate contribution to total SCFA and propionate-producing bacterial taxa compared to negative control. In western diet-fed Ldlr-/- mice, KB39 treatment resulted in an enrichment in propiogenic bacteria and propionate biosynthetic genes in vivo and an increase in absolute and relative amounts of propionate in the cecum. This also resulted in significant decreases in serum total cholesterol, LDL-cholesterol, and insulin levels, as well as reduced hepatic triglycerides and cholesterol content compared to non-treated animals. Importantly, KB39 treatment significantly reduced atherosclerosis, liver steatosis and inflammation, upregulated hepatic expression of genes involved in fatty acid oxidation and downregulated transcriptional markers of inflammation, fibrosis and insulin resistance with only a mild lowering of body weight gain.

ConclusionsOur data show that KB39, a novel synthetic glycan supporting a high propionate-producing microbiome, can reduce cardiometabolic risk factors and disease in mice and suggest this approach could be of benefit for the prevention or treatment of cardiometabolic diseases in humans.

CLINICAL PERSPECTIVEWhat is new?

O_LIA novel synthetic glycan, KB39, was selected from a library of compounds for its high propionate-producing capacity and beneficial effects on the human gut microbiome composition
C_LIO_LIKB39 modulates the gut microbiome for high propionate production and significantly reduces cardiometabolic risk factors and disease in a murine model of cardiometabolic diseases
C_LI

What are the clinical implications?

O_LIKB39, delivered orally, could be of benefit for the prevention or treatment of cardiometabolic diseases in humans
C_LIO_LIThe efficacy of KB39 in mice compared to the clinical drug fenofibrate justifies further study in humans
C_LI


title: Moxifloxacin-mediated killing of Mycobacterium tuberculosis involves respiratory downshift, reductive stress, and ROS accumulation

abstract:
Moxifloxacin is central to treatment of multidrug-resistant tuberculosis. Effects of moxifloxacin on Mycobacterium tuberculosis redox state were explored to identify strategies for increasing lethality and reducing the prevalence of extensively resistant tuberculosis. A non-invasive redox biosensor and an ROS-sensitive dye revealed that moxifloxacin induces oxidative stress correlated with M. tuberculosis death. Moxifloxacin lethality was mitigated by supplementing bacterial cultures with an ROS scavenger (thiourea), an iron chelator (bipyridyl), and, after drug removal, an antioxidant enzyme (catalase). Lethality was also reduced by hypoxia and nutrient starvation. Moxifloxacin increased the expression of genes involved in the oxidative stress response, iron-sulfur cluster biogenesis, and DNA repair. Surprisingly, and in contrast with Escherichia coli studies, moxifloxacin decreased expression of genes involved in respiration, suppressed oxygen consumption, increased the NADH/NAD+ ratio, and increased the labile iron pool in M. tuberculosis. Lowering the NADH/NAD+ ratio in M. tuberculosis revealed that NADH-reductive stress facilitates an iron-mediated ROS surge and moxifloxacin lethality. Treatment with N-acetyl cysteine (NAC) accelerated respiration and ROS production, increased moxifloxacin lethality, and lowered the mutant prevention concentration. Moxifloxacin induced redox stress in M. tuberculosis inside macrophages, and co-treatment with NAC potentiated the anti-mycobacterial efficacy of moxifloxacin during nutrient starvation, inside macrophages, and in mice where NAC restricted the emergence of resistance. Thus, oxidative stress, generated in a novel way, contributes to moxifloxacin-mediated killing of M. tuberculosis. The results open a way to make fluoroquinolones more effective anti-tuberculosis agents and provide a mechanistic basis for NAC-mediated enhancement of fluoroquinolone lethality in vitro and in vivo.

Author Summary

A new paradigm was revealed for stress-mediated bacterial death in which moxifloxacin treatment of M. tuberculosis decreases respiration rate (respiration increases in E. coli). Although moxifloxacin-induced, ROS-mediated bacterial death was observed, it derived from elevated levels of NADH and iron, a phenomenon not seen with antibiotic-treated E. coli. Nevertheless, stimulation of respiration and ROS by N-acetyl cysteine (NAC) enhanced moxifloxacin-mediated killing of M. tuberculosis, thereby reinforcing involvement of ROS in killing. NAC stimulation of moxifloxacin-mediated killing of M. tuberculosis and restriction of the emergence of resistance in a murine model of infection emphasize the importance of lethal action against pathogens. The work, plus published benefits of NAC to TB patients, encourage studies of NAC-based enhancement of fluoroquinolones.


title: Anthropogenic disturbance impacts gut microbiome homeostasis in a Malagasy primate

abstract: Increasing anthropogenic disturbances in Madagascar are exerting constrains on endemic Malagasy lemurs and their habitats, with possible effects on their health and survival. An important component of health is the gut microbiome, which might be disrupted by various stressors associated with environmental change. We have studied the gut microbiome of grey-brown mouse lemurs (Microcebus griseorufus), one of the smallest Malagasy primates and an important model of the convergent evolution of diseases. We sampled two sites: one situated in a national park and the other consisting of a more disturbed site around human settlement. We found that more intense anthropogenic disturbances indeed disrupted the gut microbiome of this lemur species with a marked reduction in its diversity and led to a shift in microbial community composition. Interestingly, we noted a decrease in beneficial bacteria together with a slight increase in disease-associated bacteria and alterations in microbial metabolic functions. Because of the crucial services provided by the microbiome to host health and in disease, such negative alterations in the gut microbiome of the lemur from anthropogenically disturbed habitat might impact the health of mouse lemurs, rendering them susceptible to diseases and ultimately affecting their survival in the shrinking biodiversity seen in Madagascar. Gut microbiome analyses might thus serve as an early warning signal for pending threats to lemur populations.


title: Evidence for a widespread third system for bacterial polysaccharide export across the outer membrane comprising a composite OPX/β-barrel translocon

abstract: In Gram-negative bacteria, secreted polysaccharides have multiple critical functions. In Wzx/Wzy- and ABC transporter-dependent pathways, an outer membrane (OM) polysaccharide export (OPX) type translocon exports the polysaccharide across the OM. The paradigm OPX protein WzaE. coli is an octamer, in which the eight C-terminal domains form an -helical OM pore, and the eight copies of the three N-terminal domains (D1-D3) a periplasmic cavity. In synthase-dependent pathways, the OM translocon is a 16- to 18- stranded {beta}-barrel protein. In Myxococcus xanthus, the secreted polysaccharide EPS is synthesized in a Wzx/Wzy-dependent pathway. Here, using experiments and computational structural biology, we characterize EpsX as an OM 18-stranded {beta}-barrel protein important for EPS synthesis and identify AlgE, a {beta}-barrel translocon of a synthase-dependent pathway, as its closest structural homolog. We also find that EpsY, the OPX protein of the EPS pathway, only consists of the periplasmic D1 and D2 domains and lacks the domain for spanning the OM (henceforth D1D2OPX protein). In vivo, EpsX and EpsY mutually stabilize each other, supporting their direct interaction. Based on these observations, we propose a model whereby EpsY and EpsX make up a novel type of translocon for polysaccharide export across the OM. Specifically, in this composite translocon, EpsX functions as the OM-spanning translocon together with the periplasmic D1D2OPX protein EpsY. Based on computational genomics, similar composite systems are present widespread in Gram-negative bacteria. This model provides a framework for these proteins future experimental characterization.


title: Re-engineering of CUP1 promoter and Cup2/Ace1 transactivator to convert Saccharomyces cerevisiae into a whole-cell eukaryotic biosensor capable of detecting 10 nM of bioavailable copper

abstract:
While copper is an essential micronutrient and a technologically indispensable heavy metal, it is toxic at high concentrations, harming the environment and human health. Currently, copper is monitored with costly and low-throughput analytical techniques that do not evaluate bioavailability, a crucial parameter which can be measured only with living cells. We overcame these limitations by building upon yeast S. cerevisiaes native copper response and constructed a promising next-generation eukaryotic whole-cell copper biosensor. We combined a dual-reporter fluorescent system with an engineered CUP1 promoter and overexpressed Cup2 transactivator, constructing through four iterations a total of 16 variants of the biosensor, with the best one exhibiting a linear range of 10-8 to 10-3 M of bioavailable copper. Moreover, this variant distinguishes itself by superior specificity, detection limit, and linear range, compared to other currently reported eukaryotic and prokaryotic whole-cell copper biosensors. By re-engineering the transactivator, we altered the systems sensitivity and growth rate, while assessing the performance of Cup2 with heterologous activation domains. Thus, in addition to presenting the next-generation whole-cell copper biosensor, this work urges for an iterative design of eukaryotic biosensors and paves the way toward higher sensitivity through transactivator engineering.

Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

category: molecular biology


title: Nrf2 prevents diabetic cardiomyopathy via antioxidation and normalization of glucose and lipid metabolism in the heart

abstract:
ObjectiveMetabolic disorders and oxidative stress are the main causes of diabetic cardiomyopathy. Activation of nuclear factor erythroid 2-related factor 2 (Nrf2) exerts a powerful antioxidant effect and prevents the progression of diabetic cardiomyopathy. However, the mechanism of its cardiac protection and direct action on cardiomyocytes are not well understood.

MethodsIn this study, cardiomyocyte-restricted Nrf2 transgenic mice (Nrf2-TG) were used to directly observe whether cardiomyocyte-specific overexpression of Nrf2 can prevent diabetic cardiomyopathy and correct glucose and lipid metabolism disorders in the heart.

ResultsCompared to wild-type (WT) mice, Nrf2-TG mice showed resistance to diabetic cardiomyopathy in a streptozotocin (STZ)-induced type 1 diabetes mouse model. This was primarily manifested as improved echocardiography results as well as reduced myocardial fibrosis, cardiac inflammation, and oxidative stress.

ConclusionThese results showed that Nrf2 can directly act on cardiomyocytes to play a cardioprotective role. Mechanistically, the cardioprotective effects of Nrf2 depend on its antioxidation activity, partially through improving glucose and lipid metabolism by targeting the metabolic pathways of Akt/GSK-3 {beta}/HK-II and AMPK/Sirt1/PGC-1.


title: Overactive mitochondrial DNA replisome causes neonatal heart failure via ferroptosis

abstract: Increasing mitochondrial DNA (mtDNA) replication and amount have been proposed as therapeutic approaches for mitochondrial dysfunction, but also as a mechanism of premature aging. We addressed this fascinating paradox by enhancing mtDNA replication via two mechanisms: increasing both mtDNA replication licensing and processivity. We crossed mice overexpressing Twinkle helicase (boosting mtDNA replication initiation) with mtDNA mutator mice (exonuclease-deficient mtDNA replicase, increasing mtDNA mutagenesis and replication processivity). The former model is asymptomatic by two years of age, whereas the latter manifests with progeroid symptoms at six months. Surprisingly, the double transgenics demonstrate postnatally halted growth and devastating cardiomyopathy, fatal within weeks. The mice show high mtDNA replication preventing cardiac maturation and the postnatal shift to oxidative metabolism, causing ferroptotic cardiomyocyte death. Our findings emphasize the critical importance of mtDNA replisome regulation for perinatal cardiac maturation. Furthermore, the data implicate ferroptosis as a cell death mechanism for neonatal mitochondrial cardiomyopathies.


title: Centriolar satellites expedite mother centriole remodeling to promote ciliogenesis.

abstract: Centrosomes are orbited by centriolar satellites, dynamic multiprotein assemblies nucleated by PCM1. To study the requirement for centriolar satellites, we generated mice lacking PCM1. Pcm1-/- mice display partially penetrant perinatal lethality with survivors exhibiting hydrocephalus, oligospermia and cerebellar hypoplasia, as well as variable expressivity of other ciliopathy features including cystic kidneys. Pcm1-/- multiciliated ependymal cells and PCM1-/- retinal pigmented epithelial 1 (RPE1) cells showed reduced ciliogenesis. PCM1-/- RPE1 cells displayed reduced docking of the mother centriole to the ciliary vesicle and removal of CP110 and CEP97 from the distal mother centriole, indicating compromized early ciliogenesis. We show these molecular cascades are maintained in vivo, and we suggest that the cellular threshold to trigger ciliogenesis varies between cell types. We propose that PCM1 and centriolar satellites facilitate efficient trafficking of proteins to and from centrioles, inducing the departure of CP110 and CEP97 to initiate ciliogenesis.

category: cell biology


title: Rheology of growing axons

abstract: The growth of axons is a key process in neural system development, which relies upon a subtle balance between external mechanical forces and remodeling of cellular constituents. A key problem in the biophysics of axons is therefore to understand the overall response of the axon under stretch, which is often modeled phenomenologically using morphoelastic or viscoelastic models. Here, we develop a microscopic mixture model of growth and remodeling based on protein turnover and damage to obtain the macroscopic rheology of axonal shafts. First, we provide an estimate for the instantaneous elastic response of axons. Second, we demonstrate that under moderate traction velocities, axons behave like a viscoelastic Maxwell material. Third, for larger velocities, we show that failure takes place due to extensive damage.

category: developmental biology


title: Western spotted skunks provide important food web linkages in forest of the Pacific Northwest

abstract: There are increasing concerns about the decreasing population trends of small mammalian carnivores around the world. With limited knowledge about their ecology and natural history, small mammal conservation and management remains difficult. To address one of these deficiencies for western spotted skunks (Spilogale gracilis), we investigated their diet in the Oregon Cascades of the Pacific Northwest during 2017 - 2019. We collected 130 spotted skunk scats opportunistically and with detection dog teams and identified prey items using DNA metabarcoding and mechanical sorting. Western spotted skunk diet consisted of invertebrates such as wasps, millipedes, and gastropods, vertebrates such as small mammals, amphibians, and birds, and plants such as Gaultheria, Rubus, and Vaccinium. Diet also consisted of items such as black-tailed deer that were likely scavenged. Comparison in diet by season revealed that spotted skunks consumed more insects during the dry season (June - August) and marginally more mammals during the wet season (September - May). We observed similar diet in areas with no record of human disturbance and areas with a history of logging. Western spotted skunks provide important food web linkages between aquatic, terrestrial, and arboreal systems by facilitating energy and nutrient transfer, and serve important functional roles of seed dispersal and scavenging. Through prey-switching, western spotted skunks may dampen the effects of irruptions of prey, such as wasps during dry springs and summers, which could then provide ecosystem resilience to environmental change.

category: ecology


title: hemispheR: an R package for fisheye canopy image analysis

abstract:
Hemispherical photography is a relevant tool to estimate canopy attributes such as leaf area index (LAI). Advancements in digital photography and image processing tools have supported long-lasting use of digital hemispherical photography (DHP). While some open-source tools have been made available for DHP, very few solutions have been made available in R programming packages, and none of these allows a full processing workflow to retrieve LAI and other canopy attributes from fisheye images.

To fill this gap, we developed an R package (hemispheR) to support the whole processing of DHP images in an automated, fast, and reproducible way. The package functions, which are designed for step-by-step single-image analysis, can be performed sequentially in a pipeline, while allowing inspecting the quality of each image processing step. The package allows to analyze both circular and fullframe fisheye images, collected either with upward facing (forest canopies) or downward facing (short canopies and crops) camera orientation. In addition, the package allows to implement two consolidated LAI methods (LAI-2000/2200 and 57{degrees} method).

A case study is presented to demonstrate the reliability of canopy attributes derived from hemispheR in temperate deciduous forests with variable canopy density and structure. Canopy attributes were validated against either results obtained from a reference proprietary software, either by benchmarking measurements obtained from terrestrial laser scanning. Results indicated hemispheR provide reliable openness and leaf area index values in forest canopies as compared with reference values.

By providing a simple, transparent, and flexible image processing procedure, hemispheR supported the use of DHP for routine measurements and monitoring of forest canopy attributes. Hosting the package in a Git repository will further support development of the package, through either collaborative coding or forking projects.


title: Genomic legacies of ancient adaptation illuminate the GC-content evolution in bacterial genomes

abstract: In conventional views, the bacterial adaptation is characterized by strong purifying selection as well as rapid evolution in changing environments. However, the genomic GC content varies greatly but has some degree of phylogenetic stability. Using 11,083 representative genomes, we report a phylogenetically constrained bimodal distribution of the genomic GC. Results suggest that such divergence of the genomic GC can be well explained by the DNA replication and repair (DRR) system, in which multiple pathways are observed correlated to the genomic GC. The biased conservations of various stress-related genes especially the DRR-related ones imply distinct adaptive evolution of the ancestral lineages of high or low GC clades which may be induced by major environmental changes in early evolution. Furthermore, our findings support that the mutational biases resulted from these legacies of adaptation have changed the course of adaptive evolution in bacteria thus causing great variation in the genomic GC. This study demonstrates the importance of indirect effects from natural selection which may be easily misinterpreted as neutral processes.

category: evolutionary biology


title: Comparing evolvability and pleiotropy across environments

abstract: The environment can play an important role in determining evolutionary outcomes (Reboud and Bell 1997; Stanton et al. 2000; Gresham et al. 2008; Cooper and Lenski 2010; Becks and Agrawal 2012; Bailey et al. 2015). Populations may increase in fitness more after evolution in one environment than in another (Hegreness et al. 2008). This may be because the distance between the ancestral genotype and the top of the locally accessible fitness peak may be greater in one environment or the other. Additionally, the rate at which mutations occur and populations move up the peaks could differ. For this reason, comparing evolvability across environments presents an interesting problem. Here, we show an environment impacts evolvability in two ways, directly by impacting the rate of adaptative evolution, and indirectly by limiting the range of fitness outcomes that are possible. We show that correlated responses are often highly idiosyncratic, due to variation in the range of possible fitness outcomes in the environment and differences in pleiotropic effects across environments but can also be predicted from the ancestral growth rate regardless of the environment in which populations evolve. Interestingly, we also show a negative correlation between increase in an environment X following evolution in environment Y and the increase in environment Y following evolution in environment X. These results highlight the necessity to measure fitness in both environments when comparing the evolvability or repeatability of evolution across environments.


title: Mechanism-based crosslinking probes capture E. coli ketosynthase FabB in conformationally-distinct catalytic states.

abstract:
Ketosynthases (KS) catalyse essential carbon-carbon bond forming reactions in fatty acid biosynthesis using a two-step, ping-pong reaction mechanism. In E. coli, there are two homodimeric elongating KSs, FabB and FabF, both of which possess overlapping substrate selectivity. However, FabB is essential for the biosynthesis of unsaturated fatty acids (UFAs) required for cell survival in the absence of exogenous UFAs. Additionally, FabB has reduced activity towards substrates longer than 14 carbons, whereas FabF efficiently catalyses the elongation of saturated C14 and unsaturated C16:1 acyl-acyl carrier protein (ACP) complexes. In this study, we solved two crosslinked crystal structures of FabB in complex with ACPs functionalized with long-chain fatty acid crosslinking probes that approximate catalytic steps. Both homodimeric structures possess asymmetric substrate binding pockets, suggestive of cooperative relationships between the two FabB monomers when engaged with C14 and C16 acyl chains. In addition, these structures capture an unusual rotamer of the active site gating residue, F392, potentially representative of the catalytic state prior to substrate release. These structures demonstrate the utility of mechanism-based crosslinking methods to capture and elucidate at near atomic resolution conformational transitions accompanying KS-mediated catalysis.

SynopsisCrystal structures of KS-ACP crosslinked complex elucidate chain length preference and substrate processing mechanism of E. coli FabB.

category: biochemistry


title: Nuclease proteins CifA and CifB promote spermatid DNA damage associated with symbiont-induced cytoplasmic incompatibility

abstract: The worldwide endosymbiosis between arthropods and Wolbachia bacteria is an archetype for reproductive parasitism. This parasitic strategy rapidly increases the proportion of symbiont-transmitting mothers, and the most common form, cytoplasmic incompatibility (CI), impacts insect evolution and arboviral control strategies. During CI, sperms from symbiotic males kill embryos of aposymbiotic females via two nuclear-targeting proteins, CifA and CifB, that alter sperm chromatin organization in Drosophila melanogaster. Here we hypothesize that Cif proteins metabolize nucleic acids of developing sperm to initiate genome integrity changes. Using in vitro and in situ transgenic, mutant, enzymatic, and cytochemical assays, we show that CifA is a previously-unrecognized DNase and RNase, and CifB is a DNase. Notably, in vitro nuclease activity translates to in situ spermatid DNA damage at the canoe stage of spermiogenesis. Evolution-guided mutations ablate Cif enzymatic activity. Nucleic acid metabolism by Cif enzymes expands a fundamental understanding of the mechanism of symbiont-mediated reproductive parasitism.


title: Structural and Preliminary Biochemical Characterization of MppQ, a PLP-Dependent Aminotransferase from 	Streptomyces hygroscopicus

abstract: MppQ is an enzyme of unknown function from Streptomyces hygroscopicus that is involved in the biosynthesis of the nonproteinogenic amino acid L-enduracididine (L-End). Since L-End is a component of several peptides showing high activity against methicillin-resistant Staphylococcus aureus (MRSA), a complete understanding of its biosynthetic pathway is of utmost importance for developing chemoenzymatic routes for syntheses of novel antibiotics. In this work, we report high-resolution X-ray crystal structures of MppQ complexed with pyridoxal-5-phosphate (PLP) and pyridoxamine-5-phosphate (PMP). The structure of MppQ shares a fold with known Type I PLP-dependent aminotransferases, consisting of an N-terminal extension, large domain, and a small domain. We also report the first functional characterization of MppQ, which we incubated with enzymatically produced 2-ketoenduracidine and observed conversion to L-End via mass spectroscopy. Additionally, we have observed that MppQ has a relatively high affinity for 2-ketoarginine, a shunt product in the L-End biosynthetic pathway, indicating a possible role of MppQ in increasing efficiency of L-End biosynthesis by converting 2-ketoarginine back to the starting material, L-arginine.


title: Engineering functional membrane-membrane interface by InterSpy

abstract: Engineering synthetic interfaces between membranes has potential applications in designing non-native cellular communication pathways and creating synthetic tissues. Here, InterSpy is introduced as a synthetic biology tool consisting of a heterodimeric protein engineered to form and maintain membrane-membrane interfaces between apposing synthetic as well as cell membranes through SpyTag/SpyCatcher interaction. Inclusion of split fluorescent protein fragments in the designed InterSpy toolkit allows tracking the formation of membrane-membrane interface and reconstitution of functional fluorescent protein in the space between apposing membranes. We first demonstrate InterSpy by testing split protein designs using a mammalian cell-free expression system. By utilizing co-translational helix insertion, cell-free synthesized InterSpy fragments are incorporated into the membrane of liposomes and supported lipid bilayers with a desired topology. Functional reconstitution of split fluorescent protein between the membranes is strictly dependent on SpyTag/SpyCatcher. Finally, since InterSpy is fully genetically encoded, the engineered system is adapted to cells and showcased. InterSpy demonstrates the power of cell-free expression systems in functional reconstitution of synthetic membrane interfaces via proximity-inducing proteins. This technology may also prove useful for synthetic biology where cell-cell contacts and communication are recreated in a controlled manner using minimal components.

category: synthetic biology


title: Screening candidate genes related to psoas muscle traits in Debao and Landrace pigs based on transcriptome analysis

abstract: To identify the important genes that affect the phenotypic differences between the Landrace and Debao pigs, especially the differences in metabolism and muscle growth. Differentially expressed genes of psoas major were detected by mRNA transcriptome sequencing in Landrace and Debao pigs. By extracting the total RNA of the psoas major muscle of the Landrace pig and the Debao pig, purifying the mRNA, constructing the cDNA library, conducting transcriptome sequencing, and then through the sequencing quality evaluation, we know that the sequencing quality of this study is relatively high. A total of 17,943 genes were detected in all samples, including 17,870 known genes and 73 new genes. Defined genes with |log2FC| greater than 2 and Q-value less than 0.001, and screened them as significantly differentially expressed genes. A total of 1661 differentially expressed genes were screened from the samples of Landrace pigs and Debao pigs, among which 1255 genes were differentially up-regulated and 406 genes were differentially down-regulated. Through differential gene analysis, it is concluded that these genes are mainly involved in metabolic regulation, muscle and fat development and other processes, especially some important functional genes such as MAPK14, FOS, SIRT1, KRAS, EGR1, CDNNB1, etc. To sum up, this study used transcriptome sequencing method, and then selected differentially expressed genes between Landrace pigs and Debao pigs through data analysis, and finally screened out important genes affecting phenotypic differences, which provided genetic support for breeding better breeds in the future.

category: genomics


title: Lumbar time-varying muscle synergies in trunk flexion and bending movements at different velocities

abstract:
PurposeAlthough the extent of which the central nervous system uses muscle synergies as a movement control strategy remains an open area of research, it is widely agreed that synergies facilitate the robustness of the neuromuscular system, allowing for effective postural control and flexible movement. This work aimed to investigate the muscle activation patterns of the trunk and time-varying muscle synergies using a novel 18-muscle 3-DOF, 3-D musculoskeletal model of the lumbar spine developed by the authors.

Methods24 different biaxial trunk movements were simulated via the optimization of kinetic and kinematic measures towards obtaining the corresponding muscle activation patterns at 3 different velocities. These patterns were subsequently used to extract the principal (phasic and tonic) spatiotemporal synergies associated with the observed muscle activation patterns in the range of simulated movements.

ResultsFour dominant synergies were able to explain a considerable percent (about 75%) of the variance of the simulated muscle activities. The extracted synergies were spatially tuned in the direction of the main simulated movements (flexion/extension and right/left lateral bending). The temporal patterns demonstrated gradual monotonic shifts in tonic synergies and biphasic modulatory components in phasic synergies with spatially tuned time-delays. The increase in velocity resulted in an elevated amplitude coefficient and accelerated activation of phasic synergies.

ConclusionOur results suggest the plausibility of a time-varying synergies strategy in the dynamic control of trunk movement. Further work is needed to explore leveraging these concepts in various applications, such as rehabilitation and musculoskeletal biomechanics, towards providing more insight into the mechanisms underlying trunk stability and flexibility.

category: bioengineering


title: MAECI: A Pipeline For Generating Consensus Sequence With Nanopore Sequencing Long-read Assembly and Error Correction

abstract: Nanopore sequencing produces long reads and offers unique advantages over next-generation sequencing, especially for the assembly of draft bacterial genomes with improved completeness. However, assembly errors can occur due to data characteristics and assembly algorithms. To address these issues, we developed MAECI, a pipeline for generating consensus sequences from multiple assemblies of the same nanopore sequencing data and error correction. Systematic evaluation showed that MAECI is an efficient and effective pipeline to improve the accuracy and completeness of bacterial genome assemblies. The available codes and implementation are at https://github.com/langjidong/MAECI.

category: bioinformatics


title: Collagen remodeling dictates pancreatic cancer bioenergetics and outcome through DDR1 activation or degradation

abstract: Pancreatic ductal adenocarcinoma (PDAC) is a highly desmoplastic, aggressive cancer that frequently progresses by liver metastasis1. Cancer-associated fibroblasts (CAF), extracellular matrix (ECM), and type I collagen (Col I) support2-5 or restrain PDAC progression and may impede blood supply and nutrient availability6-8. The dichotomous role of the stroma in PDAC, and the mechanisms through which it influences patient survival and enables desmoplastic cancers escape nutrient limitation remain poorly understood. Here we show that matrix metalloprotease (MMP)-cleaved or intact Col I (cCol I and iCol I, respectively) exert opposing effects on PDAC bioenergetics, macropinocytosis (MP), tumor growth and liver metastasis. While cCol I activates DDR1 (discoidin domain receptor-1)-NF-{kappa}B-p62-NRF2 signaling to promote PDAC growth, iCol I triggers DDR1 degradation and restrains PDAC growth. Patients whose tumors are enriched in iCol I and low in DDR1 and NRF2 have improved median survival compared to those enriched in cCol I, DDR1 and NRF2. Inhibition of DDR1-stimulated NF-{kappa}B or mitochondrial biogenesis blocked tumorigenesis in wildtype mice but not in mice expressing MMP-resistant Col I. In summary, the diverse effects of tumor stroma on PDAC growth, metastasis, and patient survival are mediated through the Col I-DDR1-NF-{kappa}B-NRF2-mitochondrial biogenesis pathway, presenting multiple new opportunities for PDAC therapy.

category: cancer biology


title: Biological responses to terahertz radiation with different power density in primary hippocampal neurons

abstract:
BackgroundTerahertz (THz) radiation is a valuable imaging and sensing tool which is widely used in industry and medicine. However, it biological effects including genotoxicity and cytotoxicity are not clear, particularly on the nervous system. In this study, we investigated how THz radiation with different frequency and intensity would affect the morphology, cell growth and function of rat hippocampal neurons in vitro.

MethodsPrimary hippocampal neuronal cells were isolated from newborn Wistar rat pups and were exposed to THz radiation with the frequencies of 0.12 THz and 0.141 THz, at an power intensities of 10 mW/cm2 and 30 mW/cm2 respectively. The cellular activities were evaluated by Cell Counting Kit-8 after exposure for 10 min and 30 min. The cellular apoptosis was examined by Annexin V staining using flow cytometry and the ultrastructure was detected by electron microscopy. Moreover, the amount of amino acid in the cultured neurons was measured by high performance liquid chromatography (HPLC).

ResultsNo obvious morphological changes, such as axon length and axon number, could be detected in primary neurons after exposure to 10mW/cm2 or 50 mW/cm2 THz for 30 min. However, the overall cellular activities were differentially affected by 10mW/cm2 and 50 mW/cm2 THz. Obvious cellular apoptosis was induced by both 10mW/cm2 and 50 mW/cm2 exposure. We also found that the amounts of most amino acids did not change significantly after exposure to these two types of THz radiation.

ConclusionsThe effects of THz radiation on hippocampal neurons depend on frequency, power, exposure time as well as the measurement time after the exposure. These results will provide important information for further investigation of the effect of THz radiation on the nervous system.


title: Sub-type selective muscarinic acetylcholine receptor modulation for the treatment of parkinsonian tremor

abstract: Parkinsons Disease is characterized by hallmark motor symptoms including resting tremor, akinesia, rigidity, and postural instability. In patient surveys of Parkinsons Disease symptoms and quality of life, tremor consistently ranks among the top concerns of patients with disease. However, the gold standard of treatment, levodopa, has inconsistent or incomplete anti-tremor effects in patients, necessitating new therapeutic strategies to help relieve this burden. Non-selective anti-muscarinic acetylcholine receptor therapeutic agents which target each of the 5 muscarinic receptor subtypes have been used as an adjunct therapy in this disease, as well as other movement disorders, and have been shown to have anti-tremor efficacy. Despite this, anti-muscarinic therapy is poorly tolerated due to adverse effects. Recent pharmacological advances have led to the discovery of muscarinic subtype selective antagonists that may keep the anti-tremor efficacy of non-selective compounds, while reducing or eliminating adverse effects. Here, we directly test this hypothesis using pharmacological models of parkinsonian tremor combined with recently discovered selective positive allosteric modulators and antagonists of the predominant brain expressed muscarinic receptors M1, M4, and M5. Surprisingly, we find that selective modulation of M1, M4, or M5 does not reduce tremor in these pre-clinical models, suggesting that central or peripheral M2 or M3 receptors may be responsible for the anti-tremor efficacy of non-selective anti-muscarinic therapies currently used in the clinic.


title: Molecular differences in APOBEC3 counteraction by primate herpesviruses reveal an evolutionary link between the genesis of APOBEC3B and novel viral ribonucleotide reductase function

abstract: Viruses have evolved diverse mechanisms to antagonize host immunity. A recently discovered example is the direct inhibition and relocalization of cellular APOBEC3B (A3B) by the Epstein-Barr virus (EBV) ribonucleotide reductase (RNR) subunit BORF2. Recent reports also indicate interactions between the RNRs of other human herpesviruses and A3B, as well as interactions with the homologous APOBEC3A (A3A) enzyme. Here, we investigate the conservation of this APOBEC3 (A3) counteraction mechanism and reveal novel insights into its evolutionary origin. First, we found that herpesvirus RNRs bind to A3s via partially distinct surfaces. Second, we show that RNR-mediated relocalization and enzymatic inhibition of cellular A3s depend on binding to different regions of the A3 catalytic domain and that the inhibition is specific to A3B. Finally, we show that the ability of viral RNRs to counteract A3s is conserved in herpesviruses that infect humans and Old World monkeys, but absent in viruses infecting New World monkeys that naturally lack A3B. These results combine to indicate that the genesis of A3B at a critical branchpoint in primate evolution may have been a driving force in selecting herpesviruses with an expanded RNR functionality through counteraction of this antiviral enzyme.


title: Surface detection of SARS-CoV-2 by lateral flow LAMP

abstract: Slowing the transmission of SARS-CoV-2 requires rapid and accurate diagnostic testing. Toward this end, loop-mediated isothermal amplification (LAMP), an isothermal genomic detection method, offers great promise but the readout tends to be difficult because it does not generate linear DNA products. Rapid antigen tests are coupled to lateral flow strips, with one (negative) or two (positive) bands providing simple rapid readout, but are less sensitive than genomic amplification methods. To address the need for a genomic amplification method that can be visualized on a lateral flow strip, we developed a novel strand-displacement probe. In this work we validate this pipeline for purified RNA, intact virus, and even virus deposited onto a surface. We demonstrate robust sensitivity (100 genomic copies) and and we demonstrate the utility of our assay as a surveillance system, with the capability to detect viral particles from surfaces, even after a week of complete dry-down. Our innovation couples the diagnostic advantages of a nucleic acid amplification test (NAAT) with the simplicity of lateral-flow readouts.

category: genetics


title: Melanoma cells release DEL-1 via small extracellular vesicles

abstract: DEL-1 (developmental endothelial locus-1) induces integrin signaling and recognizes phosphatidylserine exposed on apoptotic cells. We show that DEL-1, which is thought to be a secreted molecule, is not found in melanoma cell culture supernatants but is exported by an endosomal pathway and released via small extracellular vesicles (sEV). Proteomics of DEL-1 positive sEV, but not of DEL-1 negative sEV contain proteins associated with poor survival in cancer. To determine whether DEL-1 is suitable to predict treatment responses, we isolated sEV from plasma of melanoma patients before and 90 days of treatment with checkpoint inhibitors. Although we could not detect DEL-1 in plasma sEV even in patients with progressive disease (most likely due to the very low protein yield from sEV isolated from 1ml plasma), the principal component analysis allowed a clear differentiation between controls and patients as well as between patients before and after treatment. Interestingly, in one patient with complete regression, in the post treatment sample, the protein expression profile remained in the pre-treatment cluster. The low protein yield of patient sEV and the low patient number are clear limitations of this study, but results demonstrate that this method could have the potential to predict treatment responses.


title: Gut Enterochromaffin Cells are Critical Drivers of Visceral Pain and Anxiety

abstract: Gastrointestinal (GI) discomfort is a hallmark of most gut disorders and represents a significant component of chronic visceral pain 1. For the growing population afflicted by irritable bowel syndrome (IBS), GI hypersensitivity and pain persist long after signs of tissue injury have resolved 2. IBS also exhibits a strong sex bias afflicting women three-fold more than men 1. Identifying the molecules, cells, and circuits that mediate both the acute and persistent phases of visceral pain is a critical first step in understanding how environmental and endogenous factors produce long-term changes in the nervous system or associated tissues to engender chronic pain syndromes 3,4. Enterochromaffin (EC) cells within the gut epithelium are exceedingly rare sensory neuroendocrine cells that detect and transduce noxious stimuli to nearby nerve endings via serotonin. Here, we manipulate murine EC cell activity using genetic strategies to ascertain their contributions to visceral pain. We show that acute EC cell activation is sufficient to elicit hypersensitivity to gut distension and necessary for the sensitizing actions of isovalerate, a bacterially derived short-chain fatty acid irritant associated with inflammatory GI disorders. Remarkably, prolonged EC cell activation by itself is sufficient to produce persistent visceral hypersensitivity, even in the absence of an instigating inflammatory episode. Perturbing the activity of these rare EC cells led to a marked increase in anxiety-like behaviors that normalized after blocking serotonergic signaling. Sex differences were also observed accross a range of assays indicating that females have a higher baseline visceral sensitivity. Our findings validate a critical role for EC cell-mucosal afferent signaling in acute and persistent GI pain while highlighting mechanistically defined genetic models for studying visceral hypersensitivity, sex differences, and associated behaviors.

category: physiology


title: Cholinergic stimulation modulates the functional composition of CA3 cell types in the hippocampus

abstract: The functional heterogeneity of hippocampal CA3 pyramidal neurons has emerged as a key aspect of circuit function. Here, we explored the effects of long-term cholinergic activity on the functional heterogeneity of CA3 pyramidal neurons in organotypic slices. Application of agonists to either acetylcholine receptors (AChRs) generally, or muscarinic AChRs (mAChRs) specifically, induced robust increases in network activity in the low-gamma range. Prolonged AChR stimulation for 48 hrs uncovered a population of hyperadapting CA3 pyramidal neurons that typically fired a single, early action potential in response to current injection. Although these neurons were present in control networks, their proportions were dramatically increased following long-term cholinergic activity. Characterised by the presence of a strong M-current, the hyperadaptation phenotype was abolished by acute application of either M-channel antagonists or the re-application of AChR agonists. We conclude that long-term mAChR activation modulates the intrinsic excitability of a subset of CA3 pyramidal cells, uncovering a highly plastic cohort of neurons that are sensitive to chronic ACh modulation. Our findings provide evidence for the activity-dependent plasticity of functional heterogeneity in the hippocampus.


title: Transcriptional Heterogeneity and Cell Cycle Regulation as Central Determinants of Primitive Endoderm Priming.

abstract: During embryonic development cells acquire identity at the same time as they are proliferating, implying that an intrinsic facet of cell fate choice requires coupling lineage decisions to rates of cell division. How is the cell cycle regulated to promote or suppress heterogeneity and differentiation? We explore this question combining time lapse imaging with single cell RNA-seq in the contexts of self-renewal, priming and differentiation of embryonic stem cells (ESCs) towards the Primitive Endoderm lineage (PrE). Since ESCs are derived from the Inner Cell Mass of the mammalian blastocyst, ESCs in standard culture conditions are transcriptionally heterogeneous containing subfractions that are primed for either of the two ICM lineages, Epiblast and PrE. These subfractions represent dynamic states that can readily interconvert in culture, and the PrE subfraction is functionally primed for endoderm differentiation. Here we find that differential regulation of cell cycle can tip the balance between these primed populations, such that naive ESC culture conditions promote Epiblast-like expansion and PrE differentiation stimulates the selective proliferation of PrE-primed cells. In endoderm differentiation, we find that this change is accompanied by a counter-intuitive increase in G1 length that also appears replicated in vivo. While FGF/ERK signalling is a known key regulator of ESCs and PrE differentiation, we find it is not just responsible for ESCs heterogeneity, but also cell cycle synchronisation, required for the inheritance of similar cell cycles between sisters and cousins. Taken together, our results point to a tight relationship between transcriptional heterogeneity and cell cycle regulation in the context of lineage priming, with primed cell populations providing a pool of flexible cell types that can be expanded in a lineage-specific fashion while allowing plasticity during early determination.


title: The angular gyrus is engaged by autobiographical recall not object-semantics, or event-semantics: Evidence from contrastive propositional speech production

abstract: A long history of neuropsychology and neuroimaging has implicated the angular gyrus (AG) with a myriad of cognitive functions. We investigated the functional engagement of AG by three forms of memory: 1) episodic/autobiographical memory 2) object semantic-memory, and 3) event-semantic processing using the previously under-examined naturalistic task of propositional speech production. We conducted an ALE meta-analysis of imaging studies of propositional speech, followed by an fMRI study in which we conducted direct cross-domain comparisons. The results from the meta-analysis suggest that the AG is only engaged as part of the propositional speech network when the task carries an autobiographical component. This conclusion was strongly supported by the results from the fMRI study. The key fMRI findings were: 1) AG was positively engaged during autobiographical memory retrieval. 2) The AG was strongly deactivated for definitions of object semantics and non-propositional speech. 3) The level of AG activation increased with the degree to which the event descriptions relied on input from the episodic memory system. 4) Critically, the AG showed a very different pattern to that of known semantic representation regions (the anterior temporal lobe; ATL) - whilst AG activation increased with the autobiographical nature of the task, the ATL was equally responsive to all conditions. These results provide clear evidence that the AG is not acting as a semantic hub. In contrast, the AG activation profile directly mirrored that found in the wider autobiographical retrieval network. We propose that information stored elsewhere in the episodic system is temporally buffered online in the AG during autobiographical retrieval/memory construction.


title: Molecular dissection of condensin II-mediated chromosome assembly using in vitro assays

abstract: In vertebrates, condensin I and condensin II cooperate to assemble rod-shaped chromosomes during mitosis. Despite recent progress in our understanding of the mechanism of action of condensin I, how condensin II might work has remained largely unexplored. By introducing recombinant condensin II complexes into Xenopus egg extracts, we dissect the roles of its individual subunits in chromosome assembly. We show that two HEAT subunits, CAP-D3 and CAP-G2, play positive and negative roles, respectively, in condensin II-mediated chromosome assembly. The SMC ATPase and the basic amino acid clusters of the kleisin subunit are essential for this process. Deletion of the C-terminal tail of CAP-D3 increases the ability of condensin II to assemble chromosomes and further exposes a hidden function of CAP-G2 in the lateral compaction of chromosomes. Taken together, our results uncover a multilayered regulatory mechanism unique to condensin II, and provide profound implications in the evolution of condensin II.


title: Chromatin regulates genome-wide transcription factor binding affinities

abstract: Transcription factor binding across the genome is regulated by DNA sequence and chromatin features. However, it is not yet possible to quantify the impact of chromatin context on genome-wide transcription factor binding affinities. Here we report the establishment of a method to determine genome-wide absolute apparent binding affinities of transcription factors to native, chromatinized DNA. Our experiments revealed that DNA accessibility is the main determinant of transcription factor binding in the genome, which largely restricts nanomolar affinity binding of YY1, SP1 and MYC/MAX to promoters, while FOXA1 also interacts with non-promoter elements with high affinity. Furthermore, whereas consensus DNA binding motifs for transcription factors are important to establish very high-affinity binding sites, these motifs are not always strictly required to generate nanomolar affinity interactions in the genome. Finally, we uncovered transcription factor concentration dependent binding to specific gene classes, suggesting transcription factor concentration dependent effects on gene expression and cell fate. Importantly, our method adds a quantitative dimension to transcription factor biology which enables stratification of genomic targets based on transcription factor concentration and prediction of transcription factor binding sites under non-physiological conditions, such as disease associated overexpression of (onco)genes.


title: Socially learned arbitrary call use in a wild primate

abstract: Human speech is an unusually flexible form of primate communication. A limited repertoire of phonemes is assembled into morphemes whose combinations are linked to referents by social learning in culturally specific ways. Non-human primate vocal utterances, in contrast, are drastically less flexible at every level, i.e., acoustic diversity, combinatoriality and referential function. Here, we were interested in whether primate vocal behaviour, despite its inflexible nature, is still amenable to social learning, a well-documented, powerful mechanism in many other aspects of primate behaviour. We provided wild vervet monkeys with social learning opportunities to operate a food dispensing apparatus by uttering specific vocalisations,  move-grunts, that were contextually inappropriate. In two separate experiments, subjects witnessed how another group member produced move-grunts whilst operating a food dispenser. In experiment 1, subjects only heard playbacks of the calls, followed by food release; in experiment 2, they witnessed a complete demonstration video of conspecific including the calls, followed by food release. None of the subjects learned the task in experiment 1, but 1 of 39 subjects, a juvenile female, succeeded in experiment 2 by producing move-grunts at the food dispenser to obtain food rewards. We discuss our results and behavioural observations in relation to the current theory and its eventual implications for reconstructing the evolutionary pathway to human speech.

category: animal behavior and cognition


title: One-trial social learning of vervet monkey alarm calling

abstract: How do non-human primates learn to use their alarm calls? Social learning is a promising candidate, but its role in the acquisition of meaning and call usage has not been studied systematically, neither during ontogeny nor in adulthood. To investigate the role of social learning in alarm call comprehension and use, we exposed groups of wild vervet monkeys to two unfamiliar animal models in the presence or absence of conspecific alarm calls. To assess the learning outcome of these experiences, we then presented the models for a second time to the same monkeys, but now without additional alarm call information. In subjects previously exposed in conjunction with alarm calls, we found heightened predator inspection compared to control subjects exposed without alarm calls, indicating one-trial social learning of  meaning. Moreover, some juveniles (but not adults) produced the same alarm calls they heard during the initial exposure whereas the authenticity of the models had an additional effect. We concluded that, in non-human primates, call meaning can be acquired by one-trail social learning but that subject age and core knowledge about predators additionally moderate the acquisition of novel call-referent associations.


title: Genetically encoded sensors for analysing neurotransmission among synaptically-connected neurons

abstract: Anatomical connectome mapping in small areas of the nervous system as well as large-scale detection of neuronal activity patterns have been respectively achieved; however, it is still challenging to evaluate the functional connections among anatomically-connected neurons in a large-scale nervous system. We have developed a novel method to visualize neurotransmission named Split Protein HEmispheres for REconstitution (Sphere). By splitting a sensor into two fragments and expressing them in pre- and postsynaptic neurons separately, functional neurotransmitter sensors can be reconstituted only at the synapses between those neurons. We developed a Sphere-SF-iGluSnFR to measure glutamate levels, and further demonstrated that this system is functional in cultured cells, worms, and mouse brains. Moreover, this system is applicable to sensors other than glutamate, and colour variants have also been developed. This could allow for brain-wide imaging of functional synaptic transmission among particular neurons and identification of important neuronal circuits in the nervous system.

category: neuroscience


title: Emergence of distinct neural subspaces in motor cortical dynamics during volitional adjustments of ongoing locomotion

abstract:
3The brain is capable of simultaneously carrying out multiple functions, such as making different types of movements at the same time. One example is how we are able to both carry out stereotyped walking or running movements, while concurrently performing precise, target-directed movements such as kicking a ball in a soccer match. Recently, research has shown that different computations within the same population of neurons can be carried out without disrupting each other by confining the processes into separate subspaces. Whether this strategy is used to precisely control our limbs while maintaining locomotion is still an open question. Here, we recorded the activity of primary motor cortex in nonhuman primates during obstacle avoidance on a treadmill. We found that the same neural population was active during both basic unobstructed locomotion and volitional obstacle avoidance movements. Additionally, we identified the neural modes spanning the subspace of the low-dimensional dynamics in M1 using both supervised and unsupervised techniques. We found that motor cortex employs a subspace that consistently maintains the same cyclic activity throughout obstacle stepping, despite large changes in the movement itself. All the variance corresponding to the large change in movement during the obstacle avoidance is confined to its own distinct subspace. Our findings suggest that M1 utilizes different activity subspaces to coordinate the maintenance of ongoing locomotor-related neural dynamics and fast volitional gait adjustments during complex locomotion.

4 Significance StatementOur ability to modulate our ongoing walking gait with precise, voluntary adjustments is what allows us to navigate complex terrains. Locomotion and precise, goal-directed movements, such as reaching are two distinct movement modalities and have been shown to have differing requirements of motor cortical input. It is unknown how these two movements are represented in M1 low dimensional dynamics when both are carried out at the same time, such as during obstacle avoidance. We developed a novel obstacle avoidance paradigm in freely-moving non-human primates and discovered that the strategy employed by motor cortex is to confine the rhythmic locomotion-related dynamics and the voluntary, gait-adjustment movement into separate subspaces.


title: Human statistical learning dynamically shapes the hippocampal processing of temporal associations

abstract: Recent research suggests that the human hippocampus becomes sensitive to temporal associative patterns during statistical learning (SL). And yet, how such sensitivity develops throughout the course of learning remains unclear. We investigated this question in human adults performing a visual SL task, in which the transition probability among stimuli varied systematically across learning sessions. Hippocampal activity acutely reflected this cross-session change and was predictive of behavioral performance. Using a hidden Markov model to decode stimulus patterns from hippocampal activity, we observed a "mere-exposure" effect, such that brief experience with the temporally structured stimuli was sufficient to alter the hippocampal processing. As learning unfolded, feedback from the repeated exposure shaped the hippocampal processing to better accommodate the environmental statistics. Our findings provide direct evidence of how experience with associative patterns sculpts hippocampal activity in a nonlinear fashion, bringing new insight to its active role in SL.


title: A deep hierarchy of predictions enables assignment of semantic roles in real-time speech comprehension

abstract: Understanding speech requires mapping fleeting and often ambiguous soundwaves to meaning. Humans are known to exploit their capacity to contextualize to facilitate this process, but how internal knowledge is used and deployed in real time remains an open question. Existing models of speech processing focus on either word recognition irrespective of meaning or interactions among abstract linguistic representations without time constraints, providing only partial insights into the dynamics of speech comprehension. Here, we present a model that incrementally extracts multiple levels of information from continuous speech signals in real time, based on the inversion of a generative model that represents the listeners internal knowledge of linguistic and non-linguistic processing levels in a nested temporal hierarchy. In each hierarchy, the model periodically incorporates bottom-up incoming evidence to update its internal representations and generate new top-down predictions. We show that a context level, beyond linguistic representations, can provide the model with semantic predictions informed by sensory inputs, crucial for the disambiguation among multiple meanings of the same word. We also show that hierarchical predictions can reduce peripheral processing effort via minimizing uncertainty and prediction error, especially when sensory precisions become degraded. With this proof-of-concept model we demonstrate that the deployment of hierarchical predictions is a possible strategy for the brain to utilize structured knowledge dynamically for speech comprehension.


title: Comprehensive Structural Variant Detection: From Mosaic to Population-Level

abstract: Long-read Structural Variation (SV) calling remains a challenging but highly accurate way to identify complex genomic rearrangements. Here, we present Sniffles2, which is faster and more accurate than state-of-the-art SV caller across different coverages, sequencing technologies, and SV types. Furthermore, Sniffles2 solves the problem of family to population-level SV calling to produce fully genotyped VCF files by introducing a gVCF file concept. Across 31 Mendelian samples, we accurately identified causative SVs around MECP2, including highly complex alleles with three overlapping SVs. Sniffles2 also enables the detection of mosaic SVs in bulk long-read data. This way, we were able to identify multiple mosaic SVs across a multiple system atrophy patient brain. The identified SV showed a remarkable diversity within the cingulate cortex, impacting both genes involved in neuron function and repetitive elements. In summary, we demonstrate the utility and versatility of Sniffles2 to identify SVs from the mosaic to population levels.


title: Immediate predation risk alters the relationship between potential and realised selection on male traits in the Trinidad guppy Poecilia reticulata

abstract: Predation risk perception can alter mating behaviours in males and females, but the consequences for sexual selection remain underexplored. We have previously shown that in experimental populations of Trinidadian guppies Poecilia reticulata the opportunity for sexual selection (i.e. the variance in male reproductive fitness) was higher following exposure to a simulated risk of predation than in a no-risk condition. We build upon this result by exploring whether imminent predation risk affects: 1) the relationship between the opportunity for sexual selection and the actual strength of selection on male traits and 2) the traits contributing to male fitness, and he shape of selection on these traits. While predation risk increased the variance in male fitness, realised selection on traits remained unaffected. Pre- and postcopulatory traits follow complex patterns of nonlinear and correlational selection in both treatments. Differences in selection gradients deviate from predictions based on evolutionary responses to predation, the most notable being stronger selection on courtship rate under predation risk. Our results demonstrate that the operation of sexual selection can be altered by perception of an imminent predation risk and reinforce the notion that both trait-based and variance-based metrics should be employed for an informative quantification.


title: A playback paradox? Nathusius' bats, Pipistrellus nathusii, bypass mating and fueling opportunities on migratory transit flights

abstract: In late summer, migratory bats of the temperate zone face the challenge of accomplishing two energy-demanding tasks almost at the same time: migration and mating. Both require information and involve search efforts, such as localizing prey or finding potential mates. In non-migrating bat species, playback studies showed that listening to vocalizations of other bats, both con-and heterospecifics, may help a recipient bat to find foraging patches and mating sites. However, we are still unaware of the degree to which migrating bats depend on con- or heterospecific vocalizations for identifying potential feeding or mating opportunities during nightly transit flights. Here, we investigated the vocal responses of Nathusius pipistrelle bats, Pipistrellus nathusii, to simulated feeding and courtship aggregations at a coastal migration corridor. We presented migrating bats either feeding buzzes or courtship calls of their own or a heterospecific migratory species, the common noctule, Nyctalus noctula. We expected that during migratory transit flights, simulated feeding opportunities would be particularly attractive to bats, as well as simulated mating opportunities which at the same time indicate suitable roosts for a stopover. However, we found that the echolocation call activity of P. nathusii decreased during the playback of conspecific feeding buzzes and courtship calls, yet the call activity remained unaffected when heterospecific call types were broadcast. Our results therefore suggest that while on migratory transits, P. nathusii circumnavigate conspecific feeding and mating aggregations, possibly to save time or to reduce the risks associated with social interactions. This avoidance behavior could be a result of optimization strategies by P. nathusii when performing long-distance migratory flights.


title: Species-typical chemical signals mediate reproductive isolation in a teleost fish

abstract: Pheromones play an important role in conspecific mate preference across taxa, but the mechanisms underlying the pheromonal basis of reproductive isolation in vertebrates are poorly understood. In swordtail fish (genus Xiphophorus), conspecific mate preference depends on female perception of male urine-borne pheromone signals. We focused on interspecific differences between the sympatric X. birchmanni and X. malinche, which form natural hybrid zones as a consequence of changes in water chemistry. Here we show that pheromones are localized to the testis and use solid phase extraction followed by high performance liquid chromatography / mass spectrometry (HPLC/MS) to characterize pheromone chemical composition. Analyzing HPLC/MS readouts for pure peaks with high relative intensity identified two distinct chemical components present in X. birchmanni but absent in X. malinche. Experimental manipulation of signal composition showed that the presence of these components is critical to conspecific mate preference by X. birchmanni. Characterizing chemical signals allows for powerful tests of how they interact with the environment and receiver perception, thereby contributing to both the maintenance and breakdown of reproductive isolation.


title: GoFish: A low-cost, open-source platform for closed-loop behavioural experiments on fish

abstract: Fish are the most species-rich vertebrate group, displaying vast ecological, anatomical and behavioural diversity, and therefore are of major interest for the study of behaviour and its evolution. Despite this, with respect to other vertebrates, fish are relatively underrepresented in behavioural research. This is partly due to the difficulty of implementing stimuli, manipulanda, and data recording underwater, meaning that this is frequently done with gates to control subjects, physical displays as stimuli, and visual annotation of videos to record data. To overcome these restrictions we developed GoFish, a fully-automated platform for behavioural experiments. GoFish includes real-time video tracking of subjects, presentation of stimuli in a computer screen, an automatic feeder device, and closed-loop control of task contingencies and data acquisition. The design and software components of the platform are freely available, while the hardware is widely available and relatively inexpensive. The control software, Bonsai, is user-friendly and supported by a growing community of users. As an illustration and test of its use, we present the results of 2 experiments on discrimination learning, reversal, and choice in goldfish (Carassius auratus). GoFish enables the relatively easy implementation of high-throughput tasks and the acquisition of rich behavioural data. Our platform has the potential to become a widely used tool that facilitates complex behavioural experiments in aquatic species.


title: Delayed dynamics of migratory response to CTLA-4 blockade reveals a mechanistic view on potential T cells reinvigoration following immune checkpoint blockade

abstract:
Cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) and programmed cell death protein 1 (PD-1) receptors, two clinically relevant targets for immunotherapy of cancer, are negative regulators of in immune cell activation and migration. However, optimizing therapeutic outcomes still requires fundamental research to reach a comprehensive insight into the coherent function of immune regulators.

Here, we investigated the statistical dynamics of T cells migration as a measure of the functional response to these pathways in an experimental setup of immune checkpoint blockade. For this purpose, we used a previously developed 3-dimensional organotypic culture of patient-derived tumor spheroids.

Experiment-based dynamical modeling remarked distinct characteristics of the receptors regulation followed through with the modification of their proportions in the immune modulation. We demonstrated that time-delayed kinetics of PD-1 activation just overrides its relatively more efficient cell-level function which potentially makes an operative contribution to the functional dominance of CTLA-4 in the tumor microenvironment. Simulation results showed good agreement with data for tumor cells reduction and active immune cells count observed in each experiment.

These analyses propose a new mechanistic view on relative immunogenicity of PD-1 and CTLA-4 inhibitors manifested in literature and point the possible inherent obstacles in checkpoint inhibition-based immunotherapy of cancer to address in the future.

SignificanceEx vivo monitoring of temporal response to PD-1 and CTLA-4 in the closure of T cell movement dynamics and elucidating their feasible commitment to the kinetic constraints at cell-level resolution. Delayed dynamics of migratory response to CTLA-4 inhibition revealed a mechanistic view on potential T cell reinvigoration following immune checkpoint blockade.


title: In Utero Activation of NK Cells in Congenital CMV Infection

abstract:
BackgroundCongenital cytomegalovirus (CMV) infection is the most common infectious cause of birth defects and neurological damage in newborns. Despite a well-established role for NK cells in control of CMV infection in older children and adults, it remains unknown whether fetal NK cells can sense and respond to CMV infection acquired in utero.

MethodsHere, we investigate the impact of congenital CMV infection on the neonatal NK cell repertoire by assessing the frequency, phenotype, and functional profile of NK cells in cord blood samples from newborns with congenital CMV and from uninfected controls enrolled in a birth cohort of Ugandan mothers and infants.

ResultsWe find that neonatal NK cells from congenitally CMV infected newborns show increased expression of cytotoxic mediators, signs of maturation and activation, and an expansion of mature CD56-negative NK cells, an NK cell subset associated with chronic viral infections in adults. Activation was particularly prominent in NK cell subsets expressing the Fc{gamma} receptor CD16, indicating a role for antibody-mediated immunity against CMV in utero.

ConclusionThese findings demonstrate that NK cells can be activated in utero and suggest that NK cells may be an important component of the fetal and infant immune response against CMV.

category: immunology


title: Decision heuristics in contexts exploiting intrinsic skill

abstract: Heuristics can inform human decision making in complex environments through a reduction of computational requirements and a robustness to overparameterisation. However, tasks capturing the efficiency of reduced decision dimensionality typically ignore action proficiency in determining rewards. The value of movement parameterisation in sensorimotor control questions whether heuristics preserve efficiency when actions are non-trivial. We developed a novel selection-execution task requiring joint optimisation of action selection and spatio-temporal skill. Optimal choices could be determined by either a spatio-temporal forward simulation or a simpler spatial heuristic. Sequential-sampling models of action-selection response times parsimoniously distinguished human participants who adopted either strategy. Heuristics preserved broad decisional advantages over forward simulations. In addition, heuristics aligned with greater action proficiency, though predominantly through the core feature (spatial) shaping their decision policy. We accordingly reveal evidence that the dimensionality of information guiding action selection might be yoked to the granularity of plasticity in the motor system.


title: Bmal1-knockout mice exhibit reduced cocaine-seeking behaviour and cognitive impairments

abstract: Brain and Muscle Arnt-like Protein 1 (BMAL1) is an essential component of the molecular clock underlying circadian rhythmicity. Recently, its function has also been associated with alterations in mood, and reward processing. We investigated the behavioural and neurobiological impact of Bmal1 gene deletion in mice, as well as how these alterations affect rewarding effects of cocaine. Additionally, key clock genes and components of the dopamine system were assessed in several brain areas. Our results evidence behavioural alterations in Bmal1-KO mice including changes in locomotor activity with impaired habituation to environments as well as short term memory and social recognition impairments. In addition, Bmal1-KO mice experienced reduced cocaine-induced sensitization and rewarding effects of cocaine as well as reduced cocaine-seeking behaviour. Furthermore, Bmal1 deletion influenced the expression of other clock-related genes in the mPFC and striatum as well as alterations in the expression of dopaminergic elements. Overall, the present article offers a novel and extensive characterization of Bmal1-KO animals. We suggest that reduced cocaines rewarding effects in these mutant mice might be related to Bmal1 role as an expression regulator of MAO and TH, two essential enzymes involved in dopamine metabolism.


title: Factors determining distributions of rainforest Drosophila shift from interspecific competition to high temperature with decreasing elevation

abstract: Species turnover with elevation is a widespread phenomenon and provides valuable information on why and how ecological communities might reorganize as the climate warms. Tropical mountains typically have pronounced thermal gradients and intense species interactions, providing a testing ground for investigating the relationship between thermal tolerances and biotic interactions as the proximate factors determining species distributions. We investigated thermal tolerances and interspecific competition as causes of species turnover in the nine most abundant species of Drosophila along elevational gradients in the Australian Wet Tropics. Upper thermal limits varied less among species than lower thermal limits; nonetheless, at low elevation sites, these small differences determined the composition of communities through environmental sorting. In contrast, community composition at cool, high elevations was driven by interspecific competition rather than tolerance to low temperatures. These results run counter to common assumptions about the role of abiotic and biotic factors in structuring communities along thermal gradients, and indicate that tropical insects may be vulnerable to future warming wherever their distributions lie along elevational and other temperature gradients.


title: Newfoundland and Labrador: A mosaic founder population of an Irish and British diaspora from 300 years ago.

abstract: The founder population of Newfoundland and Labrador (NL) is a unique genetic resource, in part due to geographic and cultural isolation, where historical records describe a migration of European settlers to NL in the 18th and 19th centuries. However, the fine-scale genetic structure and ancestry of the population has not been well described. Here, we leverage dense genome-wide SNP data on 1,807 NL individuals to reveal fine-scale genetic structure in NL that is clustered around coastal communities and correlated with Christian denomination. We show that the majority of NL European ancestry can be traced back to the south-east and south-west of Ireland and England. We date a substantial population size bottleneck approximately 10-15 generations ago in NL, associated with increased haplotype sharing and autozygosity. Our results elucidate novel insights into the population history of NL and demonstrate evidence of a population conducive to further genetic studies and biomarker discovery.


title: Estimates of in vivo turnover numbers by simultaneously considering data from multiple conditions improve metabolic predictions

abstract:
Turnover numbers characterize a key property of enzymes, and their usage in constraint-based metabolic modeling is expected to increase prediction accuracy of diverse cellular phenotypes. In vivo turnover numbers can be obtained by ranking of estimates obtained by integrating reaction rate and enzyme abundance measurements from individual experiments; yet, their contribution to improving predictions of condition-specific cellular phenotypes remains elusive. Here we show that available in vitro and in vivo turnover numbers lead to poor prediction of condition-specific growth rates with protein-constrained models of Escherichia coli and Saccharomyces cerevisiae, particularly in the ultimate test scenario when protein abundances are integrated in the model. We then demonstrate that in vitro turnover numbers can be corrected via a constraint-based approach that simultaneously leverages heterogeneous physiological data from multiple experiments. We find that the resulting estimates of in vivo turnover numbers lead to improved prediction of condition-specific growth rates, particularly when protein abundances are used as constraints, and are more precise than the available in vitro turnover numbers. Therefore, our approach provides the means to decrease the bias of in vivo turnover numbers and paves the way towards cataloguing in vivo kcatomes of other organisms.

Significance StatementIntegration of turnover numbers in protein-constrained metabolic models has provided insights in enzyme allocation and can improve prediction of metabolic phenotypes. While in vivo turnover numbers are estimated by ranking the outcomes from the integration of condition-specific proteomics data and reaction rates, simultaneous consideration of physiological read-outs over multiple conditions has not been considered, yet. Here we designed and tested a constraint-based approach that leverages heterogeneous data to improve estimates of in vivo turnover numbers in model unicellular organisms. We showed that the resulting turnover numbers are more precise than in vitro estimates from public databases and lead to improved prediction of condition-specific growth. The approach is readily applicable to non-model organisms and paves the way to documenting their kcatomes.


title: Reproducible stool metagenomic biomarkers linked to the melanoma immunotherapy positive outcome

abstract: The human gut microbiome plays an important role both in humans health and disease. Recent studies have shown the undeniable influence of gut microbiota composition on cancer immunotherapy efficacy. However, these researches show a lack of consensus in defining reproducible metagenomic markers for a positive immunotherapy outcome. Accordingly, extended published data re-analysis may help reveal clearer associations between the composition of the gut microbiota and treatment response. In this study, we analyzed 358 stool metagenomes from 5 studies published earlier: 210 metagenomes from melanoma patients with positive immunotherapy outcome, 148 metagenomes from melanoma patients with negative immunotherapy outcome. The biomarkers were selected by the group comparison of patients stool samples with different treatment responses (47 responders vs 55 non-responders, 102 metagenomes). Selected biomarkers were verified using the available data describing the influence of the fecal microbiota transplantation on melanoma immunotherapy outcomes (9 donors, 6 responders, 19 non-responders, 256 metagenomes). According to our analysis, the resulting cross-study reproducible taxonomic biomarkers correspond to 12 Firmicutes, 4 Bacteroidetes, and 3 Actinobacteria. 140 gene groups were identified as reproducible functional biomarkers, including those potentially involved in production of immune-stimulating molecules and metabolites. In addition, we ranked taxonomic biomarkers by the number of functional biomarkers found in their metagenomic context. In other words, we predicted a list of the potential "most beneficial" bacteria for a positive response to melanoma immunotherapy. The obtained results can be used to make recommendations for the gut microbiota correction in cancer immunotherapy, and the resulting list of biomarkers can be considered for potential diagnostic ways for predicting melanoma immunotherapy outcome. Another important point is the functional biomarkers of positive immunotherapy outcome are distributed in different bacterial species that can explain the lack of consensus of defining melanoma immunotherapy beneficial species between different studies.


title: Age at cancer diagnosis by breed, weight, sex, and cancer type in a cohort of over 3,000 dogs: determining the optimal age to initiate cancer screening in canine patients

abstract: The goal of cancer screening is to detect disease at an early stage when treatment may be more effective. Until recently, cancer screening in dogs has relied upon annual physical examinations and routine laboratory tests, which are largely inadequate for detecting preclinical disease. With the introduction of non-invasive "liquid biopsy" cancer detection methods, the discussion is shifting from "How to screen dogs for cancer" to "When to screen dogs for cancer". To address this question, data from 3,452 cancer-diagnosed subjects were analyzed to determine the age at which dogs of certain breeds and weights are typically diagnosed with cancer. In the study population, the median age at cancer diagnosis was 8.8 years, with males diagnosed at younger ages than females, and spayed/neutered dogs diagnosed at significantly later ages than intact dogs. Overall, weight was inversely correlated with age at cancer diagnosis, and purebred dogs were diagnosed at significantly younger ages than mixed-breed dogs. For breeds with 10 or more subjects, a breed-based median age at diagnosis was calculated. A weight-based linear regression model was developed to predict the median age at diagnosis for breeds represented by fewer than 10 subjects and for mixed-breed dogs. The study findings support a general recommendation to start cancer screening for all dogs at the age of 7, and as early as 4 years of age for breeds with a lower median age at cancer diagnosis, in order to increase the chances of early detection and treatment.


title: Functional autapses form in striatal parvalbumin interneurons but not medium spiny neurons

abstract:
Autapses (or self-synapses) selectively form in specific cell types in many brain regions including the neocortex and the hippocampus, where they provide feedback control over self-spiking activities. Previous morphological studies also found putative autapses in medium spiny neurons (MSNs) of the striatum. However, it remains unclear whether striatal neurons indeed form physiologically functional autapses. We performed whole-cell recordings from striatal neurons in acute mouse brain slices, and identify autaptic neurons by the occurrence of prolonged asynchronous release (AR) of neurotransmitter after high-frequency burst of action potentials (APs) in the same cell. To our surprise, we found no autaptic release in all recorded MSNs after the AP burst, even in the presence of Sr2+ that should desynchronize and thus prolong synaptic vesicle release. In sharp contrast, we observed robust autaptic AR events in half of the recorded parvalbumin (PV)-positive neurons. Autaptic responses in PV cells were mediated by GABAA receptors, and the AR strength was dependent on the frequency and the number of APs during the burst. Further simulation results show that autapses regulate burst spiking in PV cells by providing self-inhibition and thus shape network oscillation at certain frequencies. Together, we reveal that, distinct from MSNs, striatal PV neurons form functional autapses, activation of which would regulate self-activities in PV cells, and thereby shape MSN firing and network oscillations.

Author summarySynapses, which usually occur between two neurons, are key structures for signal communication in the nervous system. However, some types of neurons form autapses, where a neuron synapses onto itself. Autaptic transmission provides feedback signal regulating self-spiking activities. Neuronal and network activities in the striatum play critical roles in motor control and other brain functions. Previous studies suggest formation of autapses in striatal principal MSNs, but it remains unclear whether striatal neurons form functional autapses. We performed direct recordings from striatal neurons and examined the occurrence of autaptic transmission in acute brain slices. Surprisingly, we did not detect any autaptic responses in MSNs. A large proportion of striatal PV neurons, however, produced robust autaptic GABA release upon high-frequency stimulation, indicating selective formation of autapses in striatal PV cells. Our computation simulations suggest that autapses provide self-inhibition in PV cells and thereby shape activities in MSNs and striatal network, particularly when PV cells discharge at high frequencies corresponding to a high dopamine state. Together, our findings indicate that PV cells, but not MSNs, in the striatum form physiologically functional autapses. Autapses in PV cells could be essential circuit elements in the striatum and contribute to striatal functions, such as motor control.


title: Trait anxiety is associated with hidden state inference during aversive reversal learning

abstract: Updating beliefs in changing environments can be achieved either by gradually adapting expectations or by identifying a hidden structure composed of separate states, and inferring which state fits one s observations best. Previous studies have found that a state inference mechanism might be associated with relapse phenomena, such as return of fear, that commonly represent a major obstacle in clinical treatment of anxiety disorders. Here, we tested whether variability in trait anxiety among healthy individuals is associated with a tendency towards inferring a hidden structure of an aversive environment, as opposed to learning gradually from observations. In a Pavlovian probabilistic aversive learning paradigm, participants had to follow changes in cue-associated shock contingencies by providing probability ratings on each trial. In three sessions, the contingencies switched between high and a low levels of shock probability (60/40%, 75/25% or 90/10%). High trait anxiety was associated with steeper behavioral switches after contingency reversals, and more accurate probability ratings overall. To elucidate the computational mechanisms behind these behavioral patterns, we compared a "1-state" model, which reflects gradual updating, with a novel state-inference model ("n-state"). High trait anxiety was associated with improved fit of the state inference model (n-state) compared to the gradual model (1-state) in the session characterized by the largest shock contingency changes (90/10). This finding provides evidence that trait anxiety variations among health adults are associated with tendency to infer hidden causes that generate the observed aversive outcomes. This was particularly evident in environments with larger contingency changes and less outcome uncertainty. This association may contribute to relapse phenomena observed among high trait anxious individuals.