Perigenual and Subgenual Anterior Cingulate Afferents Converge on Common Pyramidal Cells in Amygdala Subregions of the Macaque The subgenual (sgACC) and pregenual (pgACC) anterior cingulate are important afferents of the amygdala, with different cytoarchitecture, connectivity, and function. The sgACC is associated with arousal mechanisms linked to salient cues, while the pgACC is engaged in conflict decision-making, including in social contexts. After placing same-size, small volume tracer injections into sgACC and pgACC of the same hemisphere in male Macaques, we examined anterogradely labeled fiber distribution to understand how these different functional systems communicate in the main amygdala nuclei at both mesocopic and cellular levels. The sgACC has broad-based termination patterns. In contrast, the pgACC has a more restricted pattern which was always nested in sgACC terminals. Terminal overlap occurred in subregions of the accessory basal and basal nuclei, which we termed  hotspots. In triple-labeling confocal studies, the majority of randomly selected CAMKII (+) cells (putative amygdala glutamatergic neurons) in  hotspots received dual contacts from the sgACC and pgACC. The ratio of dual contacts occurred over a surprisingly narrow range, suggesting a consistent, tight balance of afferent contacts on postsynaptic neurons. Large boutons, which are associated with greater synaptic strength, were approximately 3 times more frequent on sgACC versus pgACC axon terminals in  hotspots, consistent with a fast  driver function. Together, the results reveal a nested interaction in which pgACC (conflict/social monitoring) terminals converge with the broader sgACC (salience) terminals at both the mesoscopic and cellular level. The pre-synaptic organization in  hotspots suggest that shifts in arousal states can rapidly, and flexibly influence decision-making functions in the amygdala.

Significance statementThe subgenual (sgACC) and perigenual cingulate (pgACC) have distinct structural and functional characteristics and are important afferent modulators of the amygdala. The sgACC is critical for arousal, while the pgACC mediates conflict-monitoring, including in social contexts. Using dual tracer injections in the same monkey, we found that sgACC inputs broadly project in the main amygdala nuclei, whereas pgACC inputs were more restricted and nested in zones containing sgACC terminals ( hotspots). The majority of CAMKII + (excitatory) amygdala neurons in  hotspots received converging contacts, which were tightly balanced. pgACC and sgACC afferent streams are therefore highly interdependent in these specific amygdala subregions, permitting  internal arousal states to rapidly shape responses of amygdala neurons involved in conflict and social monitoring networks.
Microglia and Fn14 regulate transcription and chromatin accessibility in developing neurons Cytokine signaling pathways that promote inflammation in peripheral tissues are repurposed to coordinate the refinement of synaptic connections in the developing brain. However, the downstream mechanisms through which these pathways mediate neural circuit maturation remain to be fully defined. Here, we demonstrate that Fn14, a cytokine receptor that promotes inflammation outside of the central nervous system, shapes the transcriptional profiles and chromatin landscapes of neurons in the developing brain. Single-nucleus RNA-sequencing revealed hundreds of misregulated genes in the thalamocortical neurons of the visual thalami of mice lacking either Fn14 or its microglial derived cytokine ligand TWEAK, including genes encoding proteins with critical roles in synaptic function, histone modification, and chromatin remodeling. Whole-genome analysis uncovered significant alterations in chromatin accessibility in the brains of mice lacking Fn14 or in wild-type mice following microglial depletion, and chromatin changes due to both manipulations were enriched near genes encoding regulators of synaptic function. Loss of microglia also led to the retention of excess synapses, suggesting that microglia may link modifications in neuronal chromatin to the functional refinement of neural circuits. Consistent with Fn14 shaping brain function beyond the visual system, Fn14 knockout mice displayed impairments in memory task proficiency as well as heightened sensitivity to pharmacologically induced seizures. Taken together, these results define a previously undescribed interaction between microglia, cytokine signaling, and the neuronal epigenome that is likely to contribute to neural circuit refinement and function in the brain.
Interpretable deep learning model reveals subsequences of various functions for long non-coding RNA identification Long non-coding RNAs (lncRNAs) play crucial roles in many biological processes and are implicated in several diseases. With the next-generation sequencing technologies, substantial un-annotated transcripts have been discovered. Classifying unannotated transcripts using biological experiments is more time-consuming and expensive than computational approaches. Several tools for identifying long non-coding RNAs are available. These tools, however, did not explain which features in their tools contributed to the prediction results. Here, we present Xlnc1DCNN, a tool for distinguishing long non-coding RNAs (lncRNAs) from protein-coding transcripts (PCTs) using a one-dimensional convolutional neural network with prediction explanations. The evaluation results of the human test set showed that Xlnc1DCNN outperformed other state-of-the-art tools in terms of accuracy and F1-score. The explanation results revealed that lncRNA transcripts were mainly identified as sequences with no conserved regions or with a region of transmembrane helix while protein-coding transcripts were mostly classified by conserved protein domains or families. The explanation results also conveyed the probably inconsistent annotations among the public databases, lncRNA transcripts which contain protein domains or families, as well as protein-coding transcripts which are nonsense-mediated decay or processed transcripts. Xlnc1DCNN is freely available at https://github.com/cucpbioinfo/Xlnc1DCNN. Host phenology can drive the evolution of intermediate virulence strategies in some obligate-killer parasites AO_SCPLOWBSTRACTC_SCPLOWThe traditional mechanistic trade-offs resulting in a negative correlation between transmission and virulence are the foundation of nearly all current theory on the evolution of parasite virulence. Several ecological factors have been shown to modulate the optimal virulence strategies predicted from mechanistic trade-off models, but these ecological factors have not yet been shown to be sufficient to explain the intermediate virulence strategies observed in any natural system. The timing of seasonal activity, or phenology, is a common factor that influences the types and impact of many ecological interactions but is difficult to incorporate into virulence evolution studies. We develop a mathematical model of a disease system with seasonal host activity to study the evolutionary consequences of host phenology on the virulence of obligate-killer parasite. Results from this model demonstrated that seasonal host activity is sufficient to drive the evolution of intermediate parasite virulence in some types of natural disease systems, even when a traditional mechanistic trade-off between transmission and virulence is not assumed in the modeling framework. The optimal virulence strategy in these systems can be determined by both the duration of the host activity period as well as the variation in the host emergence timing. Parasites with low virulence strategies are favored in environments with long host activity periods and in environments in which hosts emerge synchronously. The results demonstrate that host phenology can be sufficient to select for intermediate optimal virulence strategies, providing an alternative mechanism to account for virulence evolution in some natural systems. Heterogeneous interactions and polymer entropy decide organization and dynamics of chromatin domains Chromatin is known to be organized into multiple domains of varying sizes and compaction. While these domains are often imagined as static structures, they are highly dynamic and show cell-to-cell variability. Since processes such as gene regulation and DNA replication occur in the context of these domains, it is important to understand their organization, fluctuation and dynamics. To simulate chromatin domains, one requires knowledge of interaction strengths among chromatin segments. Here, we derive interaction strength parameters from experimentally known contact maps and use them to predict chromatin organization and dynamics. Taking two domains on the human chromosome as examples, we investigate its 3D organization, size/shape fluctuations, and dynamics of different segments within a domain, accounting for hydrodynamic effects. Considering different cell types, we quantify changes in interaction strengths and chromatin shape fluctuations in different epigenetic states. Perturbing the interaction strengths systematically, we further investigate how epigenetic-like changes can alter the spatio-temporal nature of the domains. Our results show that heterogeneous weak interactions are crucial in determining the organization of the domains. Computing effective stiffness and relaxation times, we investigate how perturbations in interactions affect the solid-like and liquid-like nature of chromatin domains. Quantifying dynamics of chromatin segments within a domain, we show how the competition between polymer entropy and interaction energy influence the timescales of loop formation and maintenance of stable loops. Analysis of science journalism reveals gender and regional disparities in coverage Science journalism is a critical way in which the public can remain informed and benefit from new scientific findings. Such journalism also shapes the publics view of the current state of scientific findings and legitimizes experts. Those covering science can only cite and quote a limited number of sources. Sources may be identified by the journalists research or by recommendations by other scientists. In both cases, biases may influence who is identified and ultimately included as an expert. To analyze possible biases in science journalism, we analyzed 22,001 non-research articles published by Nature. We chose to analyze Nature non-research articles since its research articles provide a natural comparator. Our analysis considered two possible sources of disparity: gender and name origin. To explore these sources of disparity, we extracted cited authors names as well as extracted names of quoted speakers. While citations and quotations within a piece do not reflect the entire information-gathering process, they can provide insight into the demographics of visible sources. We then used the extracted names to predict gender and name origin of the cited authors and speakers.

In order to appropriately quantify the level of difference, we must identify a suitable reference set for comparison. We chose first and last authors within primary research articles in Nature and a subset of Springer Nature articles in the same time period as our comparator. In our analysis, we found a skew towards male quotation in Nature science journalism-related articles. However, quotation is trending toward equal representation at a faster rate than first and last authorship in academic publishing. Interestingly, we found that the gender disparity in Nature quotes was column-dependent, with the "Career Features" column reaching gender parity. Our name origin analysis found a significant over-representation of names with predicted Celtic/English origin and under-representation of names with a predicted East Asian origin. This finding was observed both in extracted quotes and journal citations, but dampened in citations. Effect of urbanization and its environmental stressors on the intraspecific variation of flight functional traits in two bumblebee species The way urbanization shapes the intraspecific variation of pollinator functional traits is little understood. However, this topic is relevant for investigating ecosystem services and pollinator health. Here, we studied how urbanization affects the functional traits of workers in two bumblebee species (Bombus terrestris and B. pascuorum) sampled in 37 sites along a gradient of urbanization in North Italy (an area of 1800 km2 including the metropolitan context of Milan and other surrounding capital districts). Namely, we investigated the role played by land-use composition, configuration, temperature, flower resource abundance, and air pollutants on the variation of traits related to flight performance (i.e., body size, wing shape and size fluctuating asymmetry). These traits are relevant because they are commonly used as indicators of stress during insect development. The functional traits of the two bumblebees responded idiosyncratically to urbanization. Urban temperatures were associated with smaller wing sizes in B. pascuorum and with more accentuated fluctuating asymmetry of wing size in B. terrestris. Moreover, flower abundance correlated with bigger wings in B. terrestris and with less asymmetric wing sizes in B. pascuorum. Other traits did not vary significantly and other urban variables played minor effects. These patterns highlight that environmental stressors linked to urbanization negatively impact traits related to flight performance and development stability of these species with possible consequences on the pollination service they provide.

Overall, this study found species-specific variation patterns in syntopic taxa, expanding our understanding about the effects of anthropic disturbance in shaping relevant functional traits of pollinator model species. Phylogenetic analyses for orthogroup-based classification of GDSL-type esterase/lipase (GELP) family in angiosperm representative species GDSL-type esterase/lipase (GELP) enzymes have multiple functions in plants, spanning from developmental processes to the response to biotic and abiotic stresses. Genes encoding GELP belong to a large gene family with several tens to more than hundred members per angiosperm species.

Here, we applied iterative phylogenetic analyses to identify 10 main clusters subdivided into 44 expert-curated reference orthogroups (OGs) using three monocot and five dicot genomes. Our results show that some GELP OGs expanded while others were maintained as single copy genes.

This semi-automatic approach proves to be effective to characterize large gene families and provides a solid classification framework for the GELP members in angiosperms. The orthogroup-based reference will be useful to perform comparative studies, infer gene functions and better understand the evolutionary history of this gene family. Using Obviously-Related Instrumental Variables to Increase the Predictive Power of Polygenic Scores The conventional way of boosting the predictive power of polygenic scores is to increase the GWAS sample size by meta-analyzing GWAS results of multiple cohorts. In this paper, we challenge this convention. Through simulations, we show that Instrumental Variable (IV) regression using two polygenic scores constructed from independent GWAS summary statistics outperforms the typical Ordinary Least Squares (OLS) model employing a single meta-analysis based polygenic score in terms of bias, root mean squared error, and statistical power. We verify the empirical validity of the simulations by predicting educational attainment (EA) and height in a sample of siblings from the UK Biobank. We show that IV regression between-families approaches the SNP-based heritability, and improves the predictive power of polygenic scores by 12% (height) to 22% (EA). Furthermore, IV regression within-families provides the tightest lower bound for the direct genetic effect, increasing the lower bound for EA from 2.0% to 3.4%, and for height from 28.9% to 37.7%. Highly efficient synthetic CRISPR RNA/Cas9-based mutagenesis for rapid cardiovascular phenotypic screening in F0 zebrafish The zebrafish is a valuable vertebrate model to study cardiovascular formation and function due to the facile visualization and rapid development of the circulatory system in its externally growing embryos. Despite having distinct advantages, zebrafish have paralogs of many important genes, making reverse genetics approaches inefficient since generating animals bearing multiple gene mutations requires substantial efforts. Here, we present a simple and robust synthetic CRISPR RNA/Cas9-based mutagenesis approach for generating biallelic F0 zebrafish knockouts. Using a dual-guide synthetic CRISPR RNA/Cas9 ribonucleoprotein (dgRNP) system, we compared the efficiency of biallelic gene disruptions following the injections of one, two, and three dgRNPs per gene into the cytoplasm or yolk. We show that simultaneous cytoplasmic injections of three distinct dgRNPs per gene into one-cell stage embryos resulted in the most efficient and consistent biallelic gene disruptions. Importantly, this triple dgRNP approach enables efficient inactivation of cell autonomous and cell non-autonomous gene function, likely due to the low mosaicism of biallelic disruptions. In support of this finding, we provide evidence that the F0 animals generated by this method fully phenocopied the endothelial and peri-vascular defects observed in corresponding stable mutant homozygotes. Moreover, this approach faithfully recapitulated the trunk vessel phenotypes resulting from the genetic interaction between two vegfr2 zebrafish paralogs. Mechanistically, investigation of genome editing and mRNA decay indicates that the combined mutagenic actions of three dgRNPs per gene lead to an increased probability of frameshift mutations, enabling efficient biallelic gene disruptions. Therefore, our approach offers a highly robust genetic platform to quickly assess novel and redundant gene function in F0 zebrafish. Creatine transporter deficient rat model show motor dysfunction, cerebellar alterations and muscle creatine deficiency without muscle atrophy Creatine (Cr) is a nitrogenous organic acid and plays roles as fast phosphate energy buffer to replenish ATP, osmolyte, antioxidant, neuromodulator, and as a compound with anabolic and ergogenic properties in muscle. Cr is taken from the diet or endogenously synthetized by the enzymes AGAT and GAMT, and specifically taken up by the transporter SLC6A8. Loss-of-function mutations in the genes encoding for the enzymes or the transporter cause Cerebral Creatine Deficiency Syndromes (CCDS). CCDS are characterized by brain Cr deficiency, intellectual disability with severe speech delay, behavioral troubles, epilepsy and motor dysfunction. Among CCDS, the X-linked Cr transporter deficiency (CTD) is the most prevalent with no efficient treatment so far. Different animal models of CTD show reduced brain Cr levels, cognitive deficiencies and together they cover other traits similar to those of patients. However, motor function was poorly explored in CTD models and some controversies in the phenotype exist in comparison with CTD patients. Our recently described Slc6a8Y389C knock-in (KI) rat model of CTD showed mild impaired motor function, morphological alterations in cerebellum, reduced muscular mass, Cr deficiency and increased guanidinoacetate content in muscle, although no consistent signs of muscle atrophy. Our results indicate that such motor dysfunction co-occurred with both nervous and muscle dysfunction, suggesting that muscle strength and performance as well as neuronal connectivity might be affected by this Cr deficiency in muscle and brain. Molecular basis for gating of cardiac ryanodine receptor: underlying mechanisms for gain- and loss-of function mutations Cardiac ryanodine receptor (RyR2) is a large Ca2+ release channel in the sarcoplasmic reticulum and indispensable for excitation-contraction coupling in the heart. RyR2 is activated by Ca2+ and RyR2 mutations are implicated in severe arrhythmogenic diseases. Yet, the structural basis underlying channel opening and how mutations affect the channel remain unknown. Here, we addressed gating mechanism of RyR2 by combining high-resolution structures determined by cryo-electron microscopy with quantitative functional analysis of channels carrying various mutations in specific residues. We demonstrated two fundamental mechanisms for channel gating: interactions close to the channel pore stabilize the channel to prevent hyperactivity and a series of interactions in the surrounding regions is necessary for channel opening upon Ca2+ binding. Mutations at the residues involved in the former and the latter mechanisms cause gain-of-function and loss-of-function, respectively. Our results reveal gating mechanisms of the RyR2 channel and alterations by pathogenic mutations at the atomic level. Trypanosomal variant surface glycoprotein expression in human African trypanosomiasis patients Trypanosoma brucei gambiense, an extracellular protozoan parasite, is the primary causative agent of human African trypanosomiasis. T. b. gambiense is endemic to West and Central Africa, where it is transmitted by the bite of infected tsetse flies. In the bloodstream of an infected host, the parasite evades antibody recognition by altering the Variant Surface Glycoprotein (VSG) that forms a dense coat on its cell surface through a process known as antigenic variation. Each VSG has a variable N-terminal domain exposed to the host and a less variable C-terminal domain that is partially hidden from host antibodies. Our lab developed VSG-seq, a targeted RNA-seq method, to study VSG gene expression in T. brucei. Studies using VSG-seq to characterize antigenic variation in a mouse model have revealed marked diversity in VSG expression within parasite populations, but this finding has not yet been validated in a natural human infection. Here, we used VSG-seq to analyze VSGs expressed in the blood of 12 patients infected with T. b. gambiense. The number of VSGs identified per patient ranged from one to 14; notably, two VSGs were shared by more than one patient. Analysis of expressed VSG N-terminal domain types revealed that 82% of expressed VSGs encoded a type B N-terminus, a bias not seen in datasets from other T. brucei subspecies. C-terminal types in T. b. gambiense infection were also restricted. These results demonstrate a bias either in the underlying VSG repertoire of T. b. gambiense or in this subspecies expression of VSGs during infection. This work demonstrates the feasibility of using VSG-seq to study antigenic variation in human infections and highlights the importance of understanding VSG repertoires in the field.

Author SummaryHuman African Trypanosomiasis is a neglected tropical disease primarily caused by the extracellular parasite Trypanosoma brucei gambiense. To avoid elimination by the host, these parasites repeatedly replace their dense surface coat of Variant Surface Glycoprotein (VSG). Despite the important role of VSGs in prolonging infection, VSG expression during natural human infections is poorly understood. A better understanding of natural VSG gene expression dynamics can clarify the mechanisms that T. brucei uses to alter its VSG coat and improve how trypanosomiasis is diagnosed in humans. We analyzed the expressed VSGs detected in the blood of patients with trypanosomiasis. Our findings indicate that parasites in both natural and experimental infections express diverse sets of VSGs. Epidural Electrical Stimulation of the Cervical Dorsal Roots Restores Voluntary Upper Limb Control In Paralyzed Monkeys Recovering arm control is a top priority for people with paralysis. Unfortunately, the complexity of the neural mechanisms underlying arm control practically limited the effectiveness of neurotechnology approaches. Here, we exploited the neural function of surviving spinal circuits to restore voluntary arm and hand control in three monkeys with spinal cord injury using spinal cord stimulation. Our neural interface leverages the functional organization of the dorsal roots to convey artificial excitation via electrical stimulation to relevant spinal segments at appropriate movement phases. Stimulation bursts targeting specific spinal segments produced sustained arm movements enabling monkeys with arm paralysis to perform an unconstrained reach-and-grasp task. Stimulation specifically improved strength, task performances and movement quality. Electrophysiology suggested that residual descending inputs were necessary to produce coordinated movements. The efficacy and reliability of our approach hold realistic promises of clinical translation. Crowdsourced identification of multi-target kinase inhibitors for RET- and TAU- based disease: the Multi-Targeting Drug DREAM Challenge A continuing challenge in modern medicine is the identification of safer and more efficacious drugs. Precision therapeutics, which have one molecular target, have been long promised to be safer and more effective than traditional therapies. This approach has proven to be challenging for multiple reasons including lack of efficacy, rapidly acquired drug resistance, and narrow patient eligibility criteria. An alternative approach is the development of drugs that address the overall disease network by targeting multiple biological targets ( polypharmacology). Rational development of these molecules will require improved methods for predicting single chemical structures that target multiple drug targets. To address this need, we developed the Multi-Targeting Drug DREAM Challenge, in which we challenged participants to predict single chemical entities that target pro-targets but avoid anti-targets for two unrelated diseases: RET-based tumors and a common form of inherited Tauopathy. Here, we report the results of this DREAM Challenge and the development of two neural network-based machine learning approaches that were applied to the challenge of rational polypharmacology. Together, these platforms provide a potentially useful first step towards developing lead therapeutic compounds that address disease complexity through rational polypharmacology.

Author SummaryMany modern drugs are developed with the goal of modulating a single cellular pathway or target. However, many drugs are, in fact,  dirty; they target multiple cellular pathways or targets. This phenomenon is known as multi-targeting or polypharmacology. While some strive to develop  cleaner therapeutics that eliminate secondary targets, recent work has shown that multi-targeting therapeutics have key advantages for a variety of diseases. However, while multi-targeting drugs that affect a precisely-defined profile of targets may be more effective, it is difficult to computationally predict which molecules have desirable target profiles. Here, we report the results of a competitive crowdsourcing project (the Multi-Targeting Drug DREAM Challenge), where we challenged participants to predict chemicals that have desired target profiles for cancer and neurodegenerative disease. Frequency modulated timer regulates mammalian hibernation Mammalian hibernators decrease basal metabolism and body temperature (Tb) to minimize energy expenditure in harsh seasons. During hibernation, Tb drops to low temperature (<10 {degrees}C) and remains constant for days, known as deep torpor in small mammalian hibernators. Spontaneous interbout arousals interrupt torpor bouts, when Tb recovers to euthermic state ~37 {degrees}C. Torpor-interbout arousal event repeats during several months of hibernation. However, little is known about mechanisms governing Tb fluctuation across torpor-interbout arousal cycles during hibernation. Recent improvement in data-logging techniques enables us to monitor Tb for more than hundred days with high precision, opening up new avenues for quantitative analysis to address the principle governing Tb fluctuation. Here, we analyzed Tb fluctuation across torpor-interbout arousal cycle of Syrian hamster, which can hibernate in response to chronic cold and short photoperiod under a laboratory condition, using generalized harmonic analysis and discovered a model with frequency modulation quantitatively reproducing Tb fluctuation. This analysis also identified an unexpectedly longer period of 120-430 days as the period that modulates another period of several days, generating Tb fluctuation for Syrian hamster. We propose that concerted action of two endogenous periods governs torpor-interbout arousal cycles during hibernation. Muscle Activation Profile During Perturbed Walking is Modulated According to Body State During an unexpected loss of balance, avoiding a fall requires people to readjust their footing rapidly and effectively. We investigated the activation patterns of the ankle and knee muscles, and muscle fiber type recruitment resulting from unannounced, medio-lateral (i.e., right/left) horizontal-surface walking perturbations in twenty healthy adults (27.00{+/-}2.79 years, 10 females). Surface electromyography (sEMG) total spectral power for specific frequency bands (40-60Hz, 60-150Hz, 150-250Hz, 250-400Hz and 400-1000Hz), from tibialis anterior (TA) and vastus lateralis (VL) muscles were analyzed. Compared to non-perturbed walking, we found a significant increase in the total spectral power of lower-extremity muscles during the first 3 seconds after perturbation. When two feet were on the ground in time of perturbation we found a different muscle fiber type recruitment pattern between VL and TA muscles. This was not significant for perturbations implemented when one foot was on the ground. Our findings suggest that muscle operating frequency is modulated in real time to fit body state and somatosensory input in the context of functional goal requirements such as a rapid change of footing in response to unexpected loss of balance in single and double-support phases of gait.

New & NoteworthyTo study muscle spectral profiles in response to loss of balance, we investigated the dynamics of muscle spectral power changes, across different frequency bands after unannounced mechanical perturbations during walking. We showed increased activation of high-frequency motor units of the lower-limb muscles, subside 3 seconds after perturbation. Differences in power increase of specific frequency bands suggest that muscle activation is modulated in real time to fit body state in the context of functional goal requirements. In vitro pectate lyase activity and carbon uptake assays and whole genome sequencing of Bacillus amyloliquefaciens subsp. plantarum strains for a pectin defective pathway The pectin lyase activity of 59 Bacillus amyloliquefaciens subsp. plantarum (Bap) strains was tested in vitro on Pectate Agar (PA) and Tris-Spizizen Salts (TSS) medium. Bap strains were cultured on TSA medium and washed three times with sterile water before the inoculation on PA media. Higher and lower pectate lyase activity were observed in six (AP193, AP203, AP299, AP80, AP102, and AP52) and four (AP 194, AP214, AP215, and AP305) Bap strains compared to other Bap strains. A total of 12 Bap strains (AP67, AP71, AP77, AP78, AP85, AP102, AP108, AP135, AP143, AP189, AP192, and AP193) grew vigorously on TSS medium. A total of six Bap strains (AP194, AP204, AP214, AP216, AP219, and HD73) had lower growth compared to other Bap strains. Pectin (1%) were used for in vitro PA and TSS medium. Pectate lyase and utilization activity were not found in Bacillus thuringiensis subsp. kurstaki strain HD73 compared to Bap strains. A draft genome sequence for strains AP194 and AP214 that were negative for pectin utilization were generated using an Illumina MiSeq. RAST analysis revealed that the pectin-associated gene altronate hydrolase (uxaA) absent in AP 214 strain. Multiple amino acid alignments of exuT and uxuB gene sequence showed dissimilarities among AP194, AP214, and reference Bap strains. Correlation between speed and turning naturally arises for sparsely sampled cell movements Mechanisms regulating cell movement are not fully understood. One feature of cell movement that determines how far cells displace from an initial position is persistence, the ability to perform movements in a direction similar to the previous movement direction. Persistence is thus determined by turning angles between two sequential displacements and be characterized by an average turning angle or persistence time. Recent studies found that a cells average speed and turning are negatively correlated, suggesting a fundamental cell-intrinsic program whereby cells with a lower turning ability (i.e., larger persistence time) are intrinsically faster (or faster cells turn less). By simulating correlated or persistent random walks (PRWs) using two different frameworks (one based on von Mises-Fisher (vMF) distribution and another based on Ornstein-Uhlenbeck (OU) process) we show that the negative correlation between speed and turning naturally arises when cell trajectories are sub-sampled, i.e., when the frequency of sampling is lower than frequency at which cells make movements. This effect is strongest when the sampling frequency is on the order of magnitude with the typical cell persistence time and when cells vary in persistence time. Both conditions are observed for datasets of T cell movements in vivo that we have analyzed. In simulations the correlation arises due to randomness of cell movements resulting in highly variable persistence times for individual cells that with sub-sampling leads to large variability of average cell speeds. Interestingly, previously suggested methodology of calculating displacement of cell cohorts with different speeds resulted in similar results whether or not there is a cell-intrinsic correlation between cell speed and persistence. For both vMF- and OU-based simulations of PRWs we could find parameter values (distribution of persistence times, speeds, and sampling frequency) that matched experimentally measured correlations between speed and turning for two datasets of T cell movement in vivo suggesting that such simple correlations are not fully informative on the intrinsic link between speed and persistence. Our results thus suggest that sub-sampling may contribute to (and perhaps fully explains) the observed correlation between speed and turning at least for some cell trajectory data and emphasize the role of sampling frequency in inference of critical cellular parameters of cell motility such as speeds.

Secondary AbstractMeasurement of cell movements often results in a negative correlation between average speed and average turning angle suggesting an existence of a universal, cell-intrinsic movement program. We show that such a negative correlation may arise if cells in the population differ in their ability for persistent movement when the movement data are sub-sampled. We show that sub-sampling of cell trajectories generated using two different frameworks of persistent random walk can match the experimentally observed correlation between speed and turning for T cell movements in vivo. Flexible control of Pavlovian-instrumental transfer based on expected reward value The Pavlovian-instrumental transfer (PIT) paradigm is widely used to assay the motivational influence of reward-predictive cues, reflected by their ability to invigorate instrumental behavior. Leading theories of incentive learning assume that cues are assigned motivational properties based on the value of the rewards they predict. We outline an alternative account of PIT which recognizes that reward-predictive cues may suppress rather than motivate instrumental behavior under certain conditions. We posit that cues capable of triggering a strong reward expectancy will tend to inhibit instrumental behavior, which is exploratory by nature, in order to facilitate retrieval of the expected reward before it is lost or stolen. According to this view, the motivation to engage in instrumental behavior during a cue should be inversely related to the value of the predicted reward, since there is more to lose by failing to secure a high-value reward than a low-value reward. We tested this hypothesis in rats using a high-conflict PIT protocol which elicits competing instrumental and food-port entry responses. In Experiment 1, cues signaling different reward magnitudes (1, 3, or 9 pellets) differentially influenced these behaviors. Whereas the 1-pellet cue increased instrumental behavior, cues signaling 3 or 9 pellets suppressed instrumental behavior and elicited high levels of food-port activity. Experiment 2 found that reward-predictive cues suppressed instrumental behavior and increased food-port activity in a flexible manner that was disrupted by post-training reward devaluation. We discuss how the PIT task provides a useful tool for studying cognitive control over cue-motivated behavior in rodents. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. Sequence-dependent model of genes with dual σ factor preference Escherichia coli uses {sigma} factors to quickly control large gene cohorts during stress conditions. While most of its genes respond to a single {sigma} factor, approximately 5% of them have dual {sigma} factor preference. The most common are those responsive to both {sigma}70, which controls housekeeping genes, and {sigma}38, which activates genes during stationary growth and stresses. Using RNA-seq and flow-cytometry measurements, we show that  {sigma}70+38 genes are nearly as upregulated in stationary growth as  {sigma}38 genes. Moreover, we find a clear quantitative relationship between their promoter sequence and their response strength to changes in {sigma}38 levels. We then propose and validate a sequence dependent model of {sigma}70+38 genes, with dual sensitivity to {sigma}38 and {sigma}70, that is applicable in the exponential and stationary growth phases, as well in the transient period in between. We further propose a general model, applicable to other stresses and {sigma} factor combinations. Given this, promoters controlling {sigma}70+38 genes (and variants) could become important building blocks of synthetic circuits with predictable, sequence-dependent sensitivity to transitions between the exponential and stationary growth phases. Pericyte-to-endothelial cell signaling via vitronectin-integrin regulates blood-CNS barrier Endothelial cells of blood vessels of the central nervous system (CNS) constitute blood-CNS barriers. Barrier properties are not intrinsic to these cells; rather they are induced and maintained by CNS microenvironment. Notably, the abluminal surface of CNS capillaries are ensheathed by pericytes and astrocytes. However, extrinsic factors from these perivascular cells that regulate barrier integrity are largely unknown. Here, we establish vitronectin, an extracellular-matrix protein secreted by CNS pericytes, as a regulator of blood-CNS barrier function via interactions with its integrin receptor, 5 in endothelial cells. Genetic ablation of vitronectin or mutating vitronectin to prevent integrin binding as well as endothelial-specific deletion of integrin 5 causes barrier leakage. Furthermore, vitronectin-integrin 5 signaling maintains barrier integrity by actively inhibiting transcytosis in endothelial cells. These results demonstrate that signaling from perivascular cells to endothelial cells via ligand-receptor interactions is a key mechanism to regulate barrier permeability. A humanized nanobody phage display library yields potent binders of SARS CoV-2 spike Neutralizing antibodies targeting the SARS-CoV-2 spike protein have shown a great preventative/therapeutic potential. Here, we report a rapid and efficient strategy for the development and design of SARS-CoV-2 neutralizing humanized nanobody constructs with sub-nanomolar affinities and nanomolar potencies. CryoEM-based structural analysis of the nanobodies in complex with spike revealed two distinct binding modes. The most potent nanobody, RBD-1-2G(NCATS-BL8125), tolerates the N501Y RBD mutation and remains capable of neutralizing the B.1.1.7 (Alpha) variant. Molecular dynamics simulations provide a structural basis for understanding the neutralization process of nanobodies exclusively focused on the spike-ACE2 interface with and without the N501Y mutation on RBD. A primary human airway air-lung interface (ALI) ex vivo model showed that RBD-1-2G-Fc antibody treatment was effective at reducing viral burden following WA1 and B.1.1.7 SARS-CoV-2 infections. Therefore, this presented strategy will serve as a tool to mitigate the threat of emerging SARS-CoV-2 variants. CRISPR-based kinome-screening revealed MINK1 as a druggable player to rewire 5FU-resistance in OSCC through AKT/MDM2/p53 axis Cisplatin, 5FU and docetaxel (TPF) are the most common chemotherapy regimen used for advanced OSCC. However, many cancer patients experience relapse, continued tumor growth, and spread due to drug resistance, which leads to treatment failure and metastatic disease. Here, using a CRISPR/Cas9 based kinome knockout screening, Misshapen-like kinase 1 (MINK1) is identified as an important mediator of 5FU resistance in OSCC. Analysis of clinical samples demonstrated significantly higher MINK1 expression in the tumor tissues of chemotherapy non-responder as compared to chemotherapy responders. The nude mice and zebrafish xenograft experiments indicate that knocking out MINK1 restores 5FU mediated cell death in chemoresistant OSCC. An antibody based phosphorylation array screen revealed MINK1 as a negative regulator of p53. Mechanistically, MINK1 modulates AKT phosphorylation at Ser473, which enables p-MDM2 (Ser 166) mediated degradation of p53. We also identified lestaurtinib as a potent inhibitor of MINK1 kinase activity. The patient derived TPF resistant cell based xenograft data suggest that lestaurtinib restores 5FU sensitivity and facilitates a significant reduction of tumor burden. Overall, our study suggests that MINK1 is a major driver of 5FU resistance in OSCC. The novel combination of MINK1 inhibitor lestaurtinib and 5FU needs further clinical investigation in advanced OSCC. Catchet-MS identifies IKZF1-targeting Thalidomide analogues as novel HIV-1 latency reversal agents A major pharmacological strategy toward HIV cure aims to reverse latency in infected cells as a first step leading to their elimination. While the unbiased identification of molecular targets physically associated with the latent HIV-1 provirus would be highly valuable to unravel the molecular determinants of HIV-1 transcriptional repression and latency reversal, due to technical limitations, this has not been possible. Here we use a dCas9 targeted chromatin and histone enrichment strategy coupled to mass spectrometry (Catchet-MS) to describe the protein composition of the latent and activated HIV-1 5LTR. Catchet-MS identified known and novel latent 5LTR-associated host factors. Among these, IKZF1 is a novel HIV-1 transcriptional repressor, required for Polycomb Repressive Complex 2 recruitment to the LTR. We find the clinically advanced thalidomide analogue iberdomide, and the FDA approved analogues lenalidomide and pomalidomide, to be novel LRAs that, by targeting IKZF1 for degradation, reverse HIV-1 latency in CD4+T-cells isolated from virally suppressed people living with HIV-1.

One Sentence SummarydCas9 targeted chromatin and histone enrichment for mass spectrometry (Catchet-MS) led to the identification of IKZF1-targeting thalidomide analogues as novel HIV-1 latency reversal agents