title: Co-clinical FDG-PET Radiomic Signature in Predicting Response to Neoadjuvant Chemotherapy in Triple Negative Breast Cancer

abstract:
PurposeWe sought to exploit the heterogeneity afforded by patient-derived tumor xenografts (PDX) to optimize robust radiomic features associated with response to therapy in the context of a co-clinical trial and implement PDX-optimized image features in the corresponding clinical study to predict and assess response to therapy using machine-learning (ML) algorithms.

MethodsTNBC patients and subtype-matched PDX were recruited into a co-clinical FDG-PET imaging study to predict response to therapy. One hundred thirty-one imaging features were extracted from PDX and human segmented tumors. Robust image features were identified based on reproducibility, cross-correlation, and volume independence. A rank importance of predictors using ReliefF was used to identify predictive radiomic features in the preclinical PDX trial in conjunction with ML algorithms: classification and regression tree (CART), Naive Bayes (NB), and support vector machines (SVM). The top four PDX-optimized image features, defined as radiomic signatures (RadSig), from each task were then used to predict or assess response to therapy. Performance of RadSig in predicting/assessing response was compared to SUVmean, SUVmax, and lean body mass normalized SULpeak measures.

ResultsSixty-four out of 131 preclinical imaging features were identified as robust. NB-RadSig performed highest in predicting and assessing response to therapy in the preclinical PDX trial. In the clinical study, the performance of SVM-RadSig and NB-RadSig to predict and assess response was practically identical and superior to SUVmean, SUVmax, and SULpeak, measures.

ConclusionsWe optimized robust FDG-PET radiomic signatures (RadSig) to predict and assess response to therapy in a context of a co-clinical imaging trial.

DECLARATIONSO_ST_ABSFundingC_ST_ABSThis work was supported by NCI grants U24CA209837, U24CA253531, and U54CA224083; U2CCA233303, and K12CA167540; Siteman Cancer Center (SCC) Support Grant P30CA091842; and Internal funds provided by Mallinckrodt Institute of Radiology.

Conflicts of interest/Competing interests.None.

Availability of data and materialAll the co-clinical data will be available for download through the Washington University School of Medicine Co-Clinical Imaging Research Resource web portal at https://c2ir2.wustl.edu/, co-clinical database (CCDB).

Code availabilityNot applicable.

Authors contributionsConceptualization: SR, FOA, KIS; Methodology: SR, TDW, SL, KIS; Formal analysis and investigation: SR, KIS; Writing - original draft preparation: SR; Writing - review and editing: RLW, FD, KIS; Funding acquisition: RWL, FOA, SL, KIS; Resources: SL; Supervision: FD, KIS. All authors read and approved the final manuscript.

Ethics approvalAll studies were performed with approval from the Washington University Humans subjects research committee and animal studies committee.

Consent to participateInformed consent to participate in the study was obtained from all participants.

Consent for publicationNot applicable.

category: bioinformatics


title: Alpha-ketoglutarate augments prolyl hydroxylase-2 mediated inactivation of phosphorylated-Akt to inhibit induced-thrombosis and inflammation

abstract: Phosphorylation of Akt (pAkt) regulates multiple physiological and pathological processes including thrombosis and inflammation. In an approach to inhibit the pathological signalling of pAkt by prolyl-hydroxylase-2 (PHD2) we employed -ketoglutarate (KG), a cofactor of PHD2. Octyl-KG supplementation to platelets promoted PHD2 activity through elevated intracellular KG:succinate ratio and reduced aggregation in vitro by suppressing pAkt1(Thr308). Augmented PHD2 activity was confirmed by increased hydroxylated-proline alongside enhanced binding of PHD2 to pAkt in KG-treated platelets. Contrastingly, inhibitors of PHD2 significantly increased pAkt1 in platelets. Octyl-KG followed similar mechanism in monocytes to inhibit cytokine secretion in vitro. Our data also describe a suppressed pAkt1 and reduced activation of platelet and leukocyte obtained from mice supplemented with dietary-KG, unaccompanied by alteration in their counts. Dietary-KG significantly reduced clot formation and leukocyte accumulation in various organs including lung of mice treated with thrombosis-inducing agent carrageenan. Importantly, we observed a significant rescue effect of dietary-KG on inflamed lung of SARS-CoV-2 infected hamsters. KG significantly reduced leukocyte accumulation, clot formation and viral load alongside downmodulation of pAkt in lung of the infected animals. Therefore, our study suggests a safe implementation of dietary-KG in prevention of Akt-driven anomalies including thrombosis and inflammation, highlighting a better pulmonary management in COVID-19.

category: pathology


title: Di(isononyl) cyclohexane-1,2-dicarboxylate (DINCH) 	Alters Transcriptional Profiles, Lipid Metabolism and Behavior in Zebrafish Larvae

abstract: Plasticizers are commonly used in different consumer goods and personal care products to provide flexibility, durability and elasticity to polymers. Due to their reported toxicity, the use of several plasticizers including phthalates has been regulated and/or banned from the market. Di(isononyl) cyclohexane-1,2-dicarboxylate (DINCH) is an alternative plasticizer that was introduced to replace toxic plasticizers. Increasing global demand and lack of information regarding toxicity and safety assessment of DINCH have raised the concern to human and animal health. Hence, in the present study, we investigated the adverse effects of DINCH (at concentrations ranging from 0.01 to 10 M) in early developmental stages of zebrafish using different endpoints such as hatching rate, developmental abnormalities, lipid metabolism, behavior analysis and gene expression. We found that DINCH caused hatching delay in a dose-dependent manner and altered the expression of genes involved in stress response. Lipid staining using Oil Red O stain showed a slight lipid accumulation around the yolk, brain, eye and neck with increasing concentration. Genes associated with lipid metabolism such as fatty acid synthesis, {beta}-oxidation, elongation, lipid transport were significantly altered by DINCH. Behavioral analysis of larvae demonstrated a distinct locomotor activity including distance and acceleration upon exposure to DINCH both in light and dark. Genes involved in cholesterol biosynthesis and homeostasis were also affected by DINCH indicating possible developmental neurotoxicity. The present data shows that DINCH could induce physiological and metabolic toxicity to aquatic organisms. Hence, further analyses and environmental monitoring on DINCH should be conducted to determine its safety and toxicity levels.

category: pharmacology and toxicology


title: Polysaccharide utilization loci in Bacteroides determine population fitness and community-level interactions

abstract: Polysaccharide utilization loci (PULs) in the human gut microbiome have critical roles in shaping human health and ecological dynamics. We develop a CRISPR-FnCpf1-RecT genome-editing tool to study 23 PULs in the highly abundant species B. uniformis (BU). We identify the glycan-degrading functions of multiple PULs and elucidate transcriptional coordination between PULs that enables the population to adapt to the loss of PULs. Exploiting a pooled BU mutant barcoding strategy, we demonstrate that the in vitro fitness and the colonization ability of BU in the murine gut is enhanced by deletion of specific PULs and modulated by glycan availability. We show that BU PULs can mediate complex glycan-dependent interactions with butyrate producers that depend on the mechanism of degradation and the butyrate producer glycan utilizing ability. In sum, PULs are major determinants of community dynamics and butyrate production and can provide a selective advantage or disadvantage depending on the nutritional landscape.

category: synthetic biology


title: Stratum Lacunosum-moleculare Interneurons of the Hippocampus Coordinate Memory Encoding and Retrieval

abstract: Encoding and retrieval of memory are two processes serving distinct biological purposes but operating in highly overlapping brain circuits. It is unclear how the two processes are coordinated in the same brain regions, especially in the hippocampal CA1 region where the two processes converge at the cellular level. Here we find that the neuron-derived neurotrophic factor (NDNF)-positive interneurons at stratum lacunosum-moleculare (SLM) in CA1 play opposite roles in memory encoding and retrieval. These interneurons show high activities in learning and low activities in recall. Increasing their activity facilitates learning but impairs recall. They inhibit the entorhinal- but dis-inhibit the CA3- inputs to CA1 pyramidal cells and thereby either suppress or elevate CA1 pyramidal cells activity depending on animals behavioral states. Thus, by coordinating entorhinal- and CA3- dual inputs to CA1, these SLM interneurons are key to switching the hippocampus between encoding and retrieval modes.

category: neuroscience


title: Silkworm model for Bacillus anthracis infection and virulence determination

abstract: Bacillus anthracis is an obligate pathogen and a causative agent of anthrax. Its major virulence factors are plasmid-coded; however, recent studies have revealed chromosome-encoded virulence factors, indicating that the current understanding of its virulence mechanism is elusive and needs further investigation. In this study, we established a silkworm (Bombyx mori) infection model of B. anthracis Sterne. We showed that silkworms were killed by B. anthracis and cured of the infection when administered with antibiotics. We quantitatively determined the lethal dose of the bacteria that kills 50% larvae and effective doses of antibiotics that cure 50% infected larvae. Furthermore, we demonstrated that B. anthracis mutants with disruption in virulence genes such as pagA, lef, and atxA had attenuated silkworm-killing ability and reduced colonization in silkworm hemolymph. The silkworm infection model established in this study can be utilized in large-scale infection experiments to identify novel virulence determinants and develop novel therapeutic options against B. anthracis infections.

category: microbiology


title: Rhizospheric and endophytic Pseudomonas aeruginosa in edible vegetable plants share molecular and metabolic traits with clinical isolates

abstract: Pseudomonas aeruginosa, a leading opportunistic pathogen causing hospital-acquired infections is predominantly present in agricultural settings. There are minimal attempts to examine the molecular and functional attributes shared by agricultural and clinical strains of P. aeruginosa. This study aims to investigate the presence of P. aeruginosa in edible vegetable plants (including salad vegetables) and analyze the evolutionary and metabolic relatedness of the agricultural and clinical strains. Eighteen rhizospheric and endophytic P. aeruginosa strains were isolated from cucumber, tomato, eggplant, and chili directly from the farms. The identity of these strains was confirmed using biochemical, and molecular markers and their genetic and metabolic traits were compared with clinical isolates. DNA fingerprinting analyses and 16S rDNA-based phylogenetic tree revealed that the plant- and human-associated strains are evolutionarily related. Both agricultural and clinical isolates possessed plant-beneficial properties, including mineral solubilization (phosphorous, potassium, and zinc), ammonification, and the ability to release extracellular siderophore and indole-3 acetic acid. These findings suggest that rhizospheric and endophytic P. aeruginosa strains are genetically and functionally analogous to the clinical isolates. This study highlights the edible plants as a potential source for human and animal transmission of P. aeruginosa.


title: Quantifying live bacterial densities using non-invasive optical measurements of E. coli

abstract: Profiling the growth of bacterial cultures over time can be a tedious and error-prone process. Here, we present the development and evaluation of the use of the ODity platform to optically measure bacterial cell densities non-invasively. The digital growth data for E. coli MG1655 was calibrated against colony forming units (CFU/mL) obtained by plating on solid media. Diauxic-like shifts of liquid E. coli MG1655 cultures grown at 37{degrees}C in LB media were observed at densities as low as 2.9 x 107 {+/-} 1.2 CFU/mL. The shift occurred at a significantly higher cell density (6.0 x 107 {+/-} 1.2 CFU/mL) when the bacteria were cultured at 31{degrees}C. These shifts were only short lived, 15.2 {+/-} 1.5 and 20.8 {+/-} 1.8 min at 37{degrees}C and 31{degrees}C, respectively, with the previous growth rate restored thereafter. We measured minimum doubling times of 17.0 {+/-} 1.1 and 24.8 {+/-} 0.9 min at 37{degrees}C and 31{degrees}C, respectively. These results demonstrate that the growth and growth rate of bacterial cultures can be accurately determined non-invasively using the ODity device.


title: Cysteine dependence in Lactobacillus iners constitutes a novel therapeutic target to modify the vaginal microbiota

abstract: Vaginal microbiota composition affects several important reproductive health outcomes. Lactobacillus crispatus-dominant bacterial communities have favorable associations whereas anaerobe-dominant communities deficient of lactobacilli are linked to poor outcomes, including bacterial vaginosis (BV). Lactobacillus iners, the most abundant vaginal species worldwide, has adverse associations compared to L. crispatus, but standard metronidazole treatment for BV promotes L. iners-dominance, likely contributing to post-treatment relapse. L. iners is under-studied because it fails to grow in standard Lactobacillus media in vitro. Here we trace this in vitro phenotype to a species-specific cysteine requirement associated with limitations in cysteine-related transport mechanisms and show that vaginal cysteine concentrations correlate with Lactobacillus abundance in vivo. We demonstrate that cystine uptake inhibitors selectively impede L. iners growth and that combining an inhibitor with metronidazole thus promotes L. crispatus dominance of defined BV-like communities. These findings identify a novel target for therapeutic vaginal microbiota modulation to improve reproductive health.


title: Cell entry inhibitor with sulfonated colloid gold as new potent broad-spectrum virucides

abstract: We recently developed nontoxic, broad-spectrum virucidal gold nanoparticles, less than 10nm sized, modified with sulfonic acids (mesilate) that mimic heparan sulfates.Camostat, a serine protease inhibitor can introduce gold nanoparticles to the influenza virus via ionic bonds.In this study, we examined the ability of a novel sulfonated colloid gold to inhibit the virus in vivo. Thus, camostat-colloid gold is a promising candidate for the development of antiviral drugs to prevent and treat influenza infection.


title: Determinants of haemosporidian single and co-infections risks in western paleartic birds

abstract: Co-infections with multiple pathogens are common in the wild and may act as a strong selective pressure on both host and parasite evolution. Yet, contrary to single infection, the factors that shape co-infection risk are largely under-investigated. Here, we explored the extent to which bird ecology and phylogeny impact single and co-infection probabilities by haemosporidian parasites using large datasets from museum collections and a Bayesian phylogenetic modelling framework. While both phylogeny and species attributes (e.g. size of the geographic range, life-history strategy, migration) were relevant predictors of co-infection risk, these factors were less pertinent in predicting the probability of being single infected. Our study suggests that co-infection risk is under a stronger deterministic control than single-infection risk. These results underscore the combined influence of host evolutionary history and species attributes in determining single and co-infection pattern providing new avenues regarding our ability to predict infection risk in the wild.

category: ecology


title: Ageing red deer alter their spatial behaviour and become less social

abstract: Social relationships are important to many aspects of animals lives, and an individuals connections may change over the course of their lifespan. Currently, it is unclear whether social connectedness declines within individuals as they age, and what the underlying mechanisms might be, so the role of age in structuring animal social systems remains unresolved, particularly in non-primates. Here, we describe senescent declines in social connectedness using 43 years of data in a wild, individually monitored population of a long-lived mammal (European red deer, Cervus elaphus). Applying a series of spatial and social network analyses, we demonstrate that these declines occur due to within-individual changes in social behaviour, with correlated changes in spatial behaviour (smaller home ranges and movements to lower-density, lower-quality areas). These findings demonstrate that within-individual socio-spatial behavioural changes can lead older animals in fission-fusion societies to become less socially connected, shedding light on the ecological and evolutionary processes structuring wild animal populations.


title: The origin of asexual brine shrimps

abstract: Determining how and how often asexual lineages emerge within sexual species is central to our understanding of sex-asex transitions and the long-term maintenance of sex. Asexuality can arise "by transmission" from an existing asexual lineage to a new one, through different types of crosses. The occurrence of these crosses, cryptic sex, variation in ploidy and recombination within asexuals greatly complicates the study of sex-asex transitions, as they preclude the use of standard phylogenetic methods and genetic distance metrics. In this study we show how to overcome these challenges by developing new approaches to investigate the origin of the various asexual lineages of the brine shrimp Artemia parthenogenetica. We use a large sample of asexuals, including all known polyploids, and their sexual relatives. We combine flow cytometry with mitochondrial and nuclear DNA data. We develop new genetic distance measures and methods to compare various scenarios describing the origin of the different lineages. We find that all diploid and polyploid A. parthenogenetica likely arose within the last 80,000 years through successive and nested hybridization events that involved backcrosses with different sexual species. All A. parthenogenetica have the same common ancestor and therefore likely carry the same asexuality gene(s) and reproduce by automixis. These findings radically change our view of sex-asex transitions in this group, and show the importance of considering asexuality "by transmission" scenarios. The methods developed are applicable to many other asexual taxa.

category: evolutionary biology


title: Comparative therapeutic strategies for preventing aortic rupture in a mouse model of vascular Ehlers Danlos syndrome

abstract: We created a knock-in Col3a1+/G182R mouse model with spontaneous mortality caused by thoracic aortic rupture that recapitulates a rare vascular genetic disease of type III collagen, the vascular Ehlers-Danlos syndrome (vEDS). Investigation of this model showed lower survival rate in males caused by aortic rupture, thin non-inflammatory arteries and altered arterial collagen. Transcriptomic analysis of aortas showed upregulation of genes related to inflammation and cell stress response. Compared to water, survival rate of Col3a1+/G182R mice was not affected by beta-blockers (propranolol or celiprolol). Two other vasodilating anti-hypertensive agents (hydralazine, amlodipine) gave opposite results on aortic rupture and mortality rate. There was a spectacular beneficial effect of losartan, reversed by the cessation of its administration, and a marked deleterious effect of exogenous angiotensin II. These results suggest that blockade of the renin angiotensin system should be tested as a first-line medical therapy in patients with vEDS.

category: genetics


title: Analysis of the Genetic Structure and Diversity of Upland Cotton Groups in Different Planting Areas Based on SNP Markers

abstract: Genetic diversity, kinship and population genetic structure analyses of Gossypium hirsutum germplasm can provide a better understanding of the origin and evolution of G. hirsutum biodiversity. In this study, 1313331 SNP molecular markers were used to construct a phylogenetic tree of each sample using MEGAX, to perform population structure analysis by ADMIXTURE software and principal component analysis (PCA) by EIGENSOFT software, and to estimate relatedness using SPAGeDi. ADMIXTURE software divided the experimental cotton population into 16 subgroups, and the Gossypium hirsutum samples could be roughly clustered according to source place, but there were some overlapping characteristics among samples. The experimental cotton population was divided into six groups according to source to calculate the genetic diversity index (H), and the obtained value (0.306) was close to that for germplasm collected by others in China. Cluster 4 had a relatively high genetic diversity level (0.390). The degrees of genetic differentiation within the experimental cotton population groups were low (the population differentiation indexes ranged from 0.02368 to 0.10664). The genetic distance among cotton accessions varied from 0.000332651 to 0.562664014, with an average of 0.25240429. The results of this study may provide a basis for mining elite alleles and using them for subsequent association analysis.

category: genomics


title: SpotClean adjusts for spot swapping in spatial transcriptomics data

abstract: Spatial transcriptomics (ST) is a powerful and widely-used approach for profiling genome-wide gene expression across a tissue with emerging applications in molecular medicine and tumor diagnostics. Recent spatial transcriptomics experiments utilize slides containing thousands of spots with spot-specific barcodes that bind mRNA. Ideally, unique molecular identifiers at a spot measure spot-specific expression, but this is often not the case owing to bleed from nearby spots, an artifact we refer to as spot swapping. We propose SpotClean to adjust for spot swapping and, in doing so, to increase the sensitivity and precision with which downstream analyses are conducted.


title: CODEC enables 'single duplex' sequencing

abstract: Detecting mutations as rare as a single molecule is crucial in many fields such as cancer diagnostics and aging research but remains challenging. Third generation sequencers can read a double-stranded DNA molecule (a  single duplex) in whole to identify true mutations on both strands apart from false mutations on either strand but with limited accuracy and throughput. Although next generation sequencing (NGS) can track dissociated strands with Duplex Sequencing, the need to sequence each strand independently severely diminishes its throughput. Here, we developed a hybrid method called Concatenating Original Duplex for Error Correction (CODEC) that combines the massively parallel nature of NGS with the single-molecule capability of third generation sequencing. CODEC physically links both strands to enable NGS to sequence a single duplex with a single read pair. By comparing CODEC and Duplex Sequencing, we showed that CODEC achieved a similar error rate (10-6) with 100 times fewer reads and conferred  single duplex resolution to most major NGS workflows.

category: bioengineering


title: Delimitation of the Tick-Borne Flaviviruses. Resolving the Tick-Borne Encephalitis and Louping-Ill Virus Paraphyletic Taxa

abstract: The tick-borne flavivirus (TBFV) group contains at least 12 members where five of them are important pathogens of humans inducing diseases with varying severity (from mild fever forms to acute encephalitis). The taxonomy structure of TBFV is not fully clarified at present. In particular, there is a number of paraphyletic issues of tick-borne encephalitis virus (TBEV) and louping-ill virus (LIV). In this study, we aimed to apply different bioinformatic approaches to analyze all available complete genome amino acid sequences to delineate TBFV members at the species level. Results showed that the European subtype of TBEV (TBEV-E) is a distinct species unit. LIV, in turn, should be separated into two species. Additional analysis of the diversity of TBEV and LIV antigenic determinants also demonstrate that TBEV-E and LIV are significantly different from other TBEV subtypes. The analysis of available literature provided data on other virus phenotypic particularities that supported our hypothesis. So, within the TBEV+LIV paraphyletic group, we offer to assign four species to get a more accurate understanding of the TBFV interspecies structure according to the modern monophyletic conception.


title: Multicellular growth and sporulation in filamentous actinobacteria require the conserved cell division protein SepX

abstract: Filamentous actinobacteria such as Streptomyces undergo two distinct modes of cell division, leading to partitioning of growing hyphae into multicellular compartments via cross-walls, and to septation and release of unicellular spores. Specific determinants for cross-wall formation and the importance of hyphal compartmentalization for Streptomyces development are largely unknown. Here we show that SepX, an actinobacterial-specific protein, is crucial for both cell division modes in Streptomyces venezuelae. Importantly, we find that sepX-deficient mutants grow without cross-walls and that this substantially impairs the fitness of colonies and the coordinated progression through the developmental life cycle. Protein interaction studies and live-cell imaging suggest that SepX contributes to the stabilization of the divisome, a mechanism that also requires the dynamin-like protein DynB. Thus, our work identifies an important determinant for cell division in Streptomyces that is required for cellular development and sporulation.


title: Hyphal compartmentalization and sporulation in Streptomyces require the conserved cell division protein SepX


title: Harnessing methylotrophs as a bacterial platform to reduce adverse effects of the use of the heavy lanthanide gadolinium in magnetic resonance imaging

abstract: Gadolinium is a key component of magnetic resonance imaging contrast agents that are critical tools for enhanced detection and diagnosis of tissue and vascular abnormalities. Untargeted post-injection deposition of gadolinium in vivo, and association with diseases like nephrogenic systemic fibrosis, has alerted regulatory agencies to re-evaluate their widespread use and generated calls for safer gadolinium-based contrast agents (GBCAs). Increasing anthropogenic gadolinium in surface water has also raised concerns of potential bioaccumulation in plants and animals. Methylotrophic bacteria can acquire, transport, store and use light lanthanides as part of a cofactor complex with pyrroloquinoline quinone (PQQ), an essential component of XoxF-type methanol dehydrogenases (MDHs), a critical enzyme for methylotrophic growth with methanol. We report robust gadolinium-dependent methanol growth of a genetic variant of Methylorubrum extorquens AM1, named evo-HLn, for "evolved for heavy lanthanides". Genetic adaptation of evo-HLn resulted in increased xox1 promoter and XoxF MDH activities, transport and storage of Gd3+, and augmented biosynthesis of PQQ. Gadolinium-grown cells exhibited a shorter T1 relaxation time compared to cells with lanthanum or no lanthanide when analyzed by MRI. In addition, evo-HLn was able to grow on methanol using the GBCA Gd-DTPA as the sole gadolinium source, showing the potential of this strain for the development of novel GBCAs and gadolinium recovery from medical waste and/or wastewater.


title: Alpha-synuclein induces epigenomic dysregulation of glutamate signaling and locomotor pathways

abstract:
BackgroundMutations and multiplications in the gene encoding for alpha-synuclein are associated with Parkinsons disease (PD). However, not all individuals with alpha-synuclein variants develop PD, suggesting that additional factors are involved. We hypothesized that increased alpha-synuclein might alter epigenetic regulation of PD pathways.

ObjectivesTo identify genome-wide DNA methylation and hydroxymethylation changes induced by overexpression of two alpha-synuclein variants in human dopaminergic neurons, and to relate these to the corresponding transcriptome.

MethodsWe assessed DNA methylation and hydroxymethylation at >850,000 CpGs using the EPIC BeadChip in LUHMES cells differentiated to dopaminergic neurons. Control LUHMES neurons, LUHMES neurons overexpressing wild type alpha-synuclein, and LUHMES neurons overexpressing A30P alpha-synuclein were compared. We used SMITE network analysis to identify functionally related genes with altered DNA methylation, DNA hydroxymethylation, and/or gene expression, incorporating LUHMES H3K4me1 ChIP-seq to delineate enhancers in addition to the default promoter and gene body regions.

ResultsUsing stringent statistical thresholds, we found that increased expression of wild type or A30P mutant alpha-synuclein induced DNA methylation changes at thousands of CpGs and DNA hydroxymethylation changes at hundreds of CpGs. Differentially methylated sites in both genotypes were enriched for several processes including movement-associated pathways and glutamate signaling. For glutamate and other signaling pathways (i.e. PDGF, insulin), this differential DNA methylation was also associated with transcriptional changes.

ConclusionsOur results indicated that alpha-synuclein altered the DNA methylome of dopaminergic neurons, influencing regulation of pathways involved in development, signaling, and metabolism. This supports a role for alpha-synuclein in the epigenetic etiology of PD.


title: Evidence for two distinct thalamocortical circuits in retrosplenial cortex

abstract:
Retrosplenial cortex (RSC) lies at the interface between perceptual and memory networks in the brain and mediates between these, although it is not yet known how. It has two distinct subregions, granular (gRSC) and dysgranular (dRSC). The present study investigated how these subregions differ with respect to their electrophysiology and connections, as a step towards understanding their functions. gRSC is more closely connected to the hippocampal system, in which theta-band local field potential oscillations are prominent. We therefore compared theta-rhythmic single-unit activity between the two RSC subregions and found, mostly in gRSC, a subpopulation of non-directional cells with spiking activity strongly entrained by theta oscillations, suggesting a stronger coupling of gRSC to the hippocampal system. We then used retrograde tracers to examine whether differences in neural coding between RSC subregions might reflect differential inputs from the anterior thalamus, which is a prominent source of RSC afferents. We found that gRSC and dRSC differ in their afferents from two AV subfields: dorsomedial (AVDM) and ventrolateral (AVVL). AVVL targets both gRSC and dRSC, while AVDM provides a selective projection to gRSC. These combined results suggest the existence of two distinct but interacting RSC subcircuits: one connecting AVDM to gRSC that may comprise part of the cognitive hippocampal system, and the other connecting AVVL to both RSC regions that may link hippocampal and perceptual regions. We suggest that these subcircuits are distinct to allow for differential weighting during integration of converging sensory and cognitive computations: an integration that may take place in thalamus, RSC or both.

HighlightsO_LIThe two retrosplenial cortex subregions, gRSC and dRSC, differ in their temporal firing characteristics and relation to theta oscillations.
C_LIO_LIThere are differential afferents from the anteroventral thalamic nucleus to gRSC and dRSC, with the dorsomedial subnucleus projecting selectively to gRSC.
C_LIO_LIThe anteroventral thalamus-retrosplenial cortex circuitry thus comprises two functionally and anatomically distinct but connected circuits, differentially connected to the hippocampal system, that may support the integration of cognitive and perceptual information.
C_LI


title: Contributions of distinct auditory cortical inhibitory neuron types to the detection of sounds in background noise

abstract:
The ability to separate background noise from relevant acoustic signals is essential for appropriate sound-driven behavior in natural environments. Examples of this separation are apparent in the auditory system, where neural responses to behaviorally relevant stimuli become increasingly noise-invariant along the ascending auditory pathway. However, the mechanisms that underlie this reduction in responses to background noise are not well understood. To address this gap in knowledge, we first evaluated the effects of auditory cortical inactivation on mice of both sexes trained to perform a simple auditory signal-in-noise detection task, and found that outputs from the auditory cortex are important for the detection of auditory stimuli in noisy environments. Next, we evaluated the contributions of the two most common cortical inhibitory cell types, parvalbumin-expressing (PV+) and somatostatin-expressing (SOM+) interneurons, to the perception of masked auditory stimuli. We found that inactivation of either PV+ or SOM+ cells resulted in a reduction in the ability of mice to determine the presence of auditory stimuli masked by noise. These results indicate that a disruption of auditory cortical network dynamics by either of these two types of inhibitory cells is sufficient to impair the ability to separate acoustic signals from noise.

Significance StatementAppropriate behavior in a natural environment relies on the ability to separate background noise from relevant signals. We found that auditory cortical inhibitory neurons play a causal role in separating environmental noise from behaviorally relevant auditory signals. These results advance our understanding of the computations performed by the auditory system to decompose and analyze acoustic stimuli in the presence of noise.


title: TRAF2, an innate immune sensor, reciprocally regulates mitophagy and inflammation to maintain cardiac myocyte homeostasis.

abstract: Mitochondrial damage triggers cell death signaling with catastrophic consequences in long-lived and irreplaceable cells, such as cardiac myocytes. Sensing of leaked mitochondrial DNA upon mitochondrial damage is also a potent trigger of inflammation. Whether the innate immune response pathways monitor mitochondrial damage in mitochondria-rich cardiac myocytes to prevent inflammation and cell death, remains unknown. TRAF2, an adaptor protein downstream of innate immune receptors, localizes to the mitochondria in the unstressed heart, with increased mitochondrial targeting in cardiomyopathic human hearts and after cardiac ischemia-reperfusion injury in mice. Inducible cardiomyocyte-specific deletion of TRAF2 in young adult mice impairs mitophagy with rapid decline in mitochondrial quality, upregulates TLR9 expression in cardiac myocytes, and results in inflammation and cell death manifesting as a fulminant cardiomyopathy. Preventing TLR9-mediated mitochondrial DNA sensing and resultant inflammation provides a short-term reprieve from cardiomyopathy, but persistence of damaged mitochondria results in long-term recrudescence. Restoration of wild-type TRAF2, but not the E3 ubiquitin ligase deficient mutant, improves mitochondrial quality and rescues cardiomyopathy to restore homeostasis. Thus, the innate immune response acts via TRAF2 as the first line of defense against mitochondrial damage by orchestrating homeostatic mitophagy to dampen myocardial inflammation and prevent cell death.

category: cell biology


title: A comprehensive temporal patterning gene network in Drosophila medulla neuroblasts revealed by single-cell RNA sequencing

abstract: During development, neural stem cells are temporally patterned to sequentially generate a variety of neural types before exiting the cell cycle. Temporal patterning is well-studied in Drosophila, where neural stem cells called neuroblasts sequentially express cascades of Temporal Transcription Factors (TTFs) to control the birth-order dependent neural specification. However, currently known TTFs were mostly identified through candidate approaches and may not be complete. In addition, many fundamental questions remain concerning the TTF cascade initiation, progression, and termination. It is also not known why temporal progression only happens in neuroblasts but not in their differentiated progeny. In this work, we performed single-cell RNA sequencing of Drosophila medulla neuroblasts of all ages to study the temporal patterning process with single-cell resolution. Our scRNA-seq data revealed that sets of genes involved in different biological processes show high to low or low to high gradients as neuroblasts age. We also identified a list of novel TTFs, and experimentally characterized their roles in the temporal progression and neural fate specification. Our study revealed a comprehensive temporal gene network that patterns medulla neuroblasts from start to end. Furthermore, we found that the progression and termination of this temporal cascade also require transcription factors differentially expressed along the differentiation axis (neuroblasts -> -> neurons). Lola proteins function as a speed modulator of temporal progression in neuroblasts; while Nerfin-1, a factor required to suppress de-differentiation in post-mitotic neurons, acts at the final temporal stage together with the last TTF of the cascade, to promote the switch to gliogenesis and the cell cycle exit. Our comprehensive study of the medulla neuroblast temporal cascade illustrated mechanisms that might be conserved in the temporal patterning of neural stem cells.

category: developmental biology


title: Ultrasound mediated cellular deflection results in cellular depolarization

abstract: Ultrasound has been used to manipulate cells in both humans and animal models. While intramembrane cavitation and lipid clustering have been suggested as likely mechanisms, they lack experimental evidence. Here we use high-speed digital holographic microscopy (to 100-kHz order) to visualize the cellular membrane dynamics. We show that neuronal and fibroblast membranes deflect about 150 nm upon ultrasound stimulation. Next, we develop a biomechanical model that predicts changes in membrane voltage after ultrasound exposure. Finally, we validate our model predictions using whole-cell patch clamp electrophysiology on primary neurons. Collectively, we show that ultrasound stimulation directly defects the neuronal membrane leading to a change in membrane voltage and subsequent depolarization. Our model is consistent with existing data and provides a mechanism for both ultrasound-evoked neurostimulation and sonogenetic control.


title: Estimation of cancer cell fractions and clone trees from multi-region sequencing of tumors

abstract: AO_SCPLOWBSTRACTC_SCPLOWMulti-region sequencing of one or multiple biopsies of solid tumors from a patient can be used to improve our understanding of the diversity of subclones in the patients tumor and shed light on the evolutionary history of the disease. Due to the large number of possible evolutionary relationships between clones and the fundamental uncertainty of the mutational composition of subclones, elucidating the most probable evolutionary relationships poses statistical and computational challenges. We developed a Bayesian hierarchical model called PICTograph to model uncertainty in the assignment of mutations to subclones and an approach to reduce the space of possible graphical models that postulate their evolutionary origin. Compared to available methods, our approach provided more consistent and accurate estimates of cancer cell fractions and better tree topology reconstruction over a range of simulated clonal diversity. Application of PICTograph to whole exome sequencing data of individuals with pancreatic cancer precursor lesions confirmed known early occurring mutations and indicated substantial molecular diversity, including multiple distinct subclones (range 6 - 12) and intra-sample mixing of subclones. As the complete evolutionary history for some patients was not identifiable, we used ensemble-based visualizations to distinguish between highly probable evolutionary relationships recovered in multiple models from uncertain relationships occurring in a small subset of models. These analyses indicate that PICTograph provides a useful approximation to evolutionary inference, particularly when the evolutionary course of a patients cancer is complex.


title: Improved SNV discovery in barcode-stratified scRNA-seq alignments

abstract:
Single cell SNV analysis is an emerging and promising strategy to connect cell-level genetic variation to cell phenotypes. At the present, SNV detection from 10x Genomics scRNA-seq data is typically performed on the pooled sequencing reads across all cells in a sample. Here, we assess the gain of information of SNV assessments from individual cell scRNA-seq data, where the alignments are split by barcode prior to the variant call. For our analyses we use publicly available sequencing data on the human breast cancer cell line MCF7 cell line generated at consequent time-points during anti-cancer treatment. We analyzed SNV calls by three popular variant callers - GATK, Strelka2 and Mu-tect2, in combination with a method for cell-level tabulation of the sequencing read counts bearing SNV alleles - SCReadCounts. Our analysis shows that variant calls on individual cell alignments identify at least two-fold higher number of SNVs as compared to the pooled scRNA-seq. We demonstrate that scSNVs exclusively called in the single cell alignments (scSNVs) are substantially enriched in novel genetic variants and in coding functional annotations, in particular, stop-codon and missense substitutions. Furthermore, we find that the expression of some scSNVs correlates with the expression of their harbouring gene (cis-scReQTLs).

Overall, our study indicates an immense potential of SNV calls from individual cell scRNA-seq data and emphasizes on the need of cell-level variant detection approaches and tools. Given the growing accumulation of scRNA-seq datasets, cell-level variant assessments are likely to significantly contribute to the understanding of the cellular heterogeneity and the relationship between genetics variants and functional phenotypes. In addition, cell-level variant assessments from scRNA-seq can be highly informative in cancer where they can help elucidate somatic mutations evolution and functionality.


title: UK B.1.1.7 variant exhibits increased respiratory replication and shedding in nonhuman primates

abstract:
The continuing emergence of SARS-CoV-2 variants calls for regular assessment to identify differences in viral replication, shedding and associated disease. In this study, African green monkeys were infected intranasally with either a contemporary D614G or the UK B.1.1.7 variant. Both variants caused mild respiratory disease with no significant differences in clinical presentation. Significantly higher levels of viral RNA and infectious virus were found in upper and lower respiratory tract samples and tissues from B.1.1.7 infected animals. Interestingly, D614G infected animals showed significantly higher levels of viral RNA and infectious virus in rectal swabs and gastrointestinal tract tissues. Our results indicate that B.1.1.7 infection in African green monkeys is associated with increased respiratory replication and shedding but no disease enhancement similar to human B.1.1.7 cases.

One-Sentence SummaryUK B.1.1.7 infection of African green monkeys exhibits increased respiratory replication and shedding but no disease enhancement


title: Heterogeneity in effective size across the genome: effects on the Inverse Instantaneous Coalescence Rate (IICR) and implications for demographic inference under linked selection.

abstract: The relative contribution of selection and neutrality in shaping species genetic diversity is one of the most central and controversial questions in evolutionary theory. Genomic data provide growing evidence that linked selection, i.e. the modification of genetic diversity at neutral sites through linkage with selected sites, might be pervasive over the genome. Several studies proposed that linked selection could be modelled as first approximation by a local reduction (e.g. purifying selection, selective sweeps) or increase (e.g. balancing selection) of effective population size (Ne). At the genome-wide scale, this leads to variations of Ne from one region to another, reflecting the heterogeneity of selective constraints and recombination rates between regions. We investigate here the consequences of such genomic variations of Ne on the genome-wide distribution of coalescence times. The underlying motivation concerns the impact of linked selection on demographic inference, because the distribution of coalescence times is at the heart of several important demographic inference approaches. Using the concept of Inverse Instantaneous Coalescence Rate, we demonstrate that in a panmictic population, linked selection always results in a spurious apparent decrease of Ne along time. Balancing selection has a particularly large effect, even when it concerns a very small part of the genome. We also study more general models including genuine population size changes, population structure or transient selection and find that the effect of linked selection can be significantly reduced by that of population structure. The models and conclusions presented here are also relevant to the study of other biological processes generating apparent variations of Ne along the genome.


title: Encapsulated allografts preclude host sensitization and promote ovarian endocrine function in ovariectomized young rhesus monkeys and sensitized mice

abstract: Transplantation of allogeneic donor ovarian tissue holds great potential for female cancer survivors who often experience premature ovarian insufficiency. To avoid complications associated with immune suppression and to protect transplanted ovarian allografts from immune-mediated injury, we have developed an immuno-isolating hydrogel-based capsule that supports the function of ovarian allografts without triggering an immune response. Encapsulated ovarian allografts implanted in naive ovariectomized BALB/c mice responded to the circulating gonadotropins without direct revascularization and maintained function for 4 months, as evident by regular estrous cycles and presence of antral follicles in the retrieved grafts. Repeated implantations of encapsulated mouse ovarian allografts did not sensitize naive BALB/c mice in contrast to non-encapsulated controls, which was confirmed with undetectable levels of allo-antibody. Further, encapsulated allografts implanted in hosts previously sensitized by implantation of non-encapsulated allografts restored estrous cycles similarly to our results in naive recipients. Next, we tested the translational potential and efficiency of the immune-isolating capsule in a rhesus monkey model by implanting encapsulated ovarian auto- and allografts in young ovariectomized animals. The encapsulated ovarian grafts survived and restored basal levels of urinary estrone conjugate and pregnanediol 3-glucuronide during the approximate 4-5 month observation period. We demonstrate, for the first time, that encapsulation of ovarian allografts prevents sensitization and protects the allograft from rejection in young rhesus monkeys and in sensitized mice.


title: Metabolomic analysis of diverse mice reveals hepatic arginase-1 as source of plasma arginase in Plasmodium chabaudi infection

abstract:
Infections disrupt host metabolism, but the factors that dictate the nature and magnitude of metabolic change are incompletely characterized. To determine how host metabolism changes in relation to disease severity in murine malaria, we performed plasma metabolomics on eight Plasmodium chabaudi-infected mouse strains with diverse disease phenotypes. We identified plasma metabolic biomarkers for both the nature and severity of different malarial pathologies. A subset of metabolic changes, including plasma arginine depletion, match the plasma metabolomes of human malaria patients, suggesting new connections between pathology and metabolism in human malaria. In our malarial mice, liver damage, which releases hepatic arginase-1 (Arg1) into circulation, correlated with plasma arginine depletion. We confirmed that hepatic Arg1 was the primary source of increased plasma arginase activity in our model, which motivates further investigation of liver damage in human malaria patients. More broadly, our approach shows how leveraging phenotypic diversity can identify and validate relationships between metabolism and the pathophysiology of infectious disease.

ImportanceMalaria is a severe and sometimes fatal infectious disease endemic to tropical and subtropical regions. Effective vaccines against malaria-causing Plasmodium parasites remain elusive, and malaria treatments often fail to prevent severe disease. Small molecules that target host metabolism have recently emerged as candidates for therapeutics in malaria and other diseases. However, our limited understanding of how metabolites affect pathophysiology limits our ability to develop new metabolite therapies. By providing a rich dataset of metabolite-pathology correlations, and by validating one of those correlations, our work is an important step toward harnessing metabolism to mitigate disease. Specifically, we showed that liver damage in P. chabaudi-infected mice releases hepatic arginase-1 into circulation, where it may deplete plasma arginine, a candidate malaria therapeutic that mitigates vascular stress. Our data suggest that liver damage may confound efforts to increase levels of arginine in human malaria patients.


title: Anti-SARS-CoV-2 potential of Cissampelos pareira L. identified by Connectivity map-based analysis and in vitro studies

abstract:
BackgroundViral infections have a history of abrupt and severe eruptions through the years in the form of pandemics. And yet, definitive therapies or preventive measures are not present.

PurposeHerbal medicines have been a source of various antiviral compounds. An accelerated repurposing potential of antiviral herbs can provide usable drugs and identify druggable targets. In this study, we dissect the anti-coronavirus activity of Cissampelos pareira L (Cipa). using an integrative approach.

MethodsWe analysed the signature similarities between predicted antiviral agents and Cipa using the connectivity map (https://clue.io/). Next, we tested the anti-SARS-COV-2 activity of Cipa in vitro. A three-way comparative analysis of Cipa transcriptome, COVID-19 BALF transcriptome and CMAP signatures of small compounds was also performed.

ResultsSeveral predicted antivirals showed a high positive connectivity score with Cipa such as apcidin, emetine, homoharringtonine etc. We also observed 98% inhibition of SARS-COV-2 replication in infected Vero cell cultures with the whole extract. Some of its prominent pure constituents e.g pareirarine, cissamine, magnoflorine exhibited 40-80% inhibition. Comparison of genes between BALF and Cipa showed an enrichment of biological processes like transcription regulation and response to lipids, to be downregulated in Cipa while being upregulated in COVID-19. CMAP also showed that Triciribine, torin-1 and VU-0365114-2 had positive connectivity with BALF 1 and 2, and negative connectivity with Cipa.

category: systems biology


title: Newly synthesized RNA Sequencing Characterizes Transcription Dynamics in Three Pluripotent States

abstract: Unique transcriptomes define naive, primed and paused pluripotent states in mouse embryonic stem cells. Here we perform transient transcriptome sequencing (TT-seq) to de novo define and quantify coding and non-coding transcription units (TUs) in different pluripotent states. We observe a global reduction of RNA synthesis, total RNA amount and turnover rates in ground state naive cells (2i) and paused pluripotency (mTORi). We demonstrate that elongation velocity can be reliably estimated from TT-seq nascent RNA and RNA polymerase II occupancy and observe a transcriptome-wide attenuation of elongation velocity in the two inhibitor-induced states. We also discover a relationship between elongation velocity and termination read-through distance. Our analysis suggests that steady-state transcriptomes in mouse ES cells are controlled predominantly on the level of RNA synthesis, and that signaling pathways governing different pluripotent states immediately control key parameters of transcription.


title: Tools, strains, and strategies to effectively conduct anaerobic and aerobic transcriptional reporter screens and assays in Staphylococcus aureus

abstract:
Transcriptional reporters are reliable and time-tested tools to study gene regulation. In Staphylococcus aureus, {beta}-galactosidase (lacZ)-based genetic screens are not widely used because of the necessity of selectable markers for strain construction and the production of staphyloxanthin pigment which obfuscates results. We describe a series of vectors that allow for markerless insertion of codon-optimized lacZ-based transcriptional reporters. The vectors encode for different ribosomal binding sites allowing for tailored lacZ expression. A {Delta}crtM::kanR deletion insertion mutant was constructed that prevents the synthesis of staphyloxanthin, thereby permitting blue-white screening without the interference of carotenoid production. We demonstrate the utility of these vectors to monitor aerobic and anaerobic transcriptional activity. For the latter, we describe the use of a ferrocyanide-ferricyanide redox system (Fe(CN)63-/4-) permitting blue-white screening in the absence of oxygen. We also describe additional reporter systems and methods for monitoring transcriptional activity during anaerobic culture including a FAD-binding fluorescent protein (EcFbFP), alpha-hemolysin (hla), or lipase (geh). The systems and methods described are compatible with vectors utilized to create and screen high-density transposon mutant libraries.

ImportanceStaphylococcus aureus is a human pathogen and a leading cause of infectious disease-related illness and death worldwide. For S. aureus to successfully colonize and invade host tissues, it must tightly control the expression of genes encoding for virulence factors. Oxygen tension varies greatly at infection sites and many abscesses are devoid of oxygen. In this study, we have developed novel tools and methods to study how and when S. aureus alters transcription of genes. A key advantage to these methods and tools is that they can be utilized in the presence and absence of oxygen. A better understanding of anaerobic gene expression in S. aureus will provide important insight into the regulation of genes in low oxygen environments.


title: Monoclonal antibodies against cell wall chitooligomers as accessory tools for the control of cryptococcosis

abstract: Therapeutic strategies against systemic mycoses can involve antifungal resistance and significant toxicity. Thus, novel therapeutic approaches to fight fungal infections are urgent. Monoclonal antibodies (mAbs) are promising tools to fight systemic mycoses. In this study, mAbs of the IgM isotype were developed against chitin oligomers. Chitooligomers derive from chitin, an essential component of the fungal cell wall and a promising therapeutic target, as it is not synthesized by humans or animals. Surface plasmon resonance (SPR) assays and cell-binding tests showed that the mAbs recognizing chitooligomers have high affinity and specificity for the chitin derivatives. In vitro tests showed that the chitooligomer mAbs increased the fungicidal capacity of amphotericin B against Cryptococcus neoformans. The chitooligomer-binding mAbs interfered with two essential properties related to cryptococcal pathogenesis: biofilm formation and melanin production. In a murine model of C. neoformans infection, the combined administration of the chitooligomer-binding mAb and subinhibitory doses of amphotericin B promoted disease control. The data obtained in this study support the hypothesis that chitooligomer antibodies are of great potential as accessory tools in the control of cryptococcosis.


title: Glycolytic inhibitor 2-Deoxy-D-glucose attenuates SARS-CoV-2 multiplication in host cells and weakens the infective potential of progeny virions

abstract: The COVID-19 pandemic is an ongoing public health emergency of international concern. While a lot of efforts are being invested in vaccinating the population, there is also an emergent requirement to find potential therapeutics to effectively counter this fast mutating SARS-CoV-2 virus-induced pathogenicity. Virus-infected host cells switch their metabolism to a more glycolytic phenotype. This switch induced by the virus is needed for faster production of ATP and higher levels of anabolic intermediates, required for new virion synthesis and packaging. In this study, we used 2-Deoxy-D-glucose (2-DG) to target and inhibit the metabolic reprogramming induced by SARS-CoV-2 infection. Our results showed that virus infection induces glucose influx and glycolysis resulting in selective high accumulation of the fluorescent glucose/2-DG analogue, 2-NBDG in these cells. Subsequently, 2-DG inhibits glycolysis in infected cells thereby reducing the virus multiplication and alleviates the cells from virus induced cytopathic effect (CPE) and cell death. Herein, we demonstrate that the crucial Nglycosites (N331 and N343) of RBD in spike protein of progeny virions produced from 2-DG treated cells were found unglycosylated and defective with compromised infectivity potential. In line with earlier reported observations, our study also showed that 2-DG mediated metabolic inhibiton can attenuate SARS-COV-2 multiplication. In addition, mechanistic study revealed that the inhibition of SARS-COV-2 multiplication is attributed to 2-DG induced un-glycosylation of spike protein. Our findings strengthen the notion that 2-DG effectively inhibits SARS-CoV-2 multiplication. Therefore, based on its previous human trials in different types of Cancer and Herpes patients, it could be a potential molecule to study in COVID-19 patients.


title: Canada's human footprint reveals large intact areas juxtaposed against areas under immense anthropogenic pressure

abstract: Efforts are underway in Canada to set aside terrestrial lands for conservation, thereby protecting them from anthropogenic pressures. Here we produce the first Canadian human footprint map to identify intact and modified lands and ecosystems. Our results showed strong spatial variation in pressures across the country, with just 18% of Canada experiencing measurable human pressure. However, some ecosystems are experiencing very high pressure, such as the Great Lakes Plains and Prairies national ecological areas which have over 75% and 56% of their areas, respectively, with a high human footprint. In contrast, the Arctic and Northern Mountains have less than 0.02% and 0.2% under high human footprint. A validation of the final map resulted in a Cohen Kappa statistic of 0.911, signifying an  almost perfect agreement between the human footprint and the validation data set. By increasing the number and accuracy of mapped pressures, our map demonstrates much more widespread pressures in Canada than were indicated by previous global mapping efforts, demonstrating the value in specific national data applications. Ecological areas with immense anthropogenic pressure, highlight challenges that may arise when planning for ecologically representative protected areas.


title: A TPR scaffold couples signal detection to OdhI phosphorylation in metabolic control by the protein kinase PknG

abstract:
Signal transduction is essential for bacteria to adapt to changing environmental conditions. Among many forms of post-translational modifications, reversible protein phosphorylation has evolved as a ubiquitous molecular mechanism of protein regulation in response to specific stimuli. The Ser/Thr protein kinase PknG modulates the fate of intracellular glutamate by controlling the phosphorylation status of the 2-oxoglutarate dehydrogenase regulator OdhI, a function that is conserved among diverse actinobacteria. PknG has a modular organization characterized by the presence of regulatory domains surrounding the catalytic domain. Here we present an investigation through in vivo experiments as well as biochemical and structural methods of the molecular bases of the regulation of PknG from C. glutamicum (CgPknG), in the light of previous knowledge available for the kinase from M. tuberculosis (MtbPknG). We found that OdhI phosphorylation by CgPknG is regulated by a conserved mechanism that depends on a C-terminal domain composed of tetratricopeptide repeats (TPR) essential for metabolic homeostasis. Furthermore, we identified a conserved structural motif that physically connects the TPR domain and a flexible N-terminal extension of the kinase that is involved in docking interactions with OdhI. Based on our results and previous reports, we propose a model in which the TPR domain of PknG couples signal detection to the specific phosphorylation of OdhI. Overall, the available data indicate that conserved PknG domains in distant actinobacteria retain their roles in kinase regulation in response to nutrient availability.

IMPORTANCEBacteria control the metabolic processes by which they obtain nutrients and energy in order to adapt to the environment. In this way, the metabolic characteristics of a microorganism determine its ecological role and its usefulness in industrial processes. Here, we use genetic, biochemical, and structural approaches to study a key component in a system that regulates glutamate production in C. glutamicum, a species that is used for the industrial production of amino acids. We elucidated molecular mechanisms involved in metabolic control in C. glutamicum, which are conserved in related pathogenic bacteria. The findings have broader significance for diverse actinobacteria, including microorganisms that cause disease as well as environmental species used to produce billions of dollars of amino acids and antibiotics every year.

category: biochemistry


title: Supervised learning with word embeddings derived from PubMed captures latent knowledge about protein kinases and cancer

abstract: Inhibiting protein kinases (PKs) that cause cancers has been an important topic in cancer therapy for years. So far, almost 8% of more than 530 PKs have been targeted by FDA-approved medications and around 150 protein kinase inhibitors (PKIs) have been tested in clinical trials. We present an approach based on natural language processing and machine learning to the relations between PKs and cancers, predicting PKs whose inhibition would be efficacious to treat a certain cancer. Our approach represents PKs and cancers as semantically meaningful 100-dimensional vectors based on co-occurrence patterns in PubMed abstracts. We use information about phase I-IV trials in ClinicalTrials.gov to construct a training set for random forest classification. In historical data, associations between PKs and specific cancers could be predicted years in advance with good accuracy. Our model may be a tool to predict the relevance of inhibiting PKs with specific cancers.


title: Interpretable modeling of genotype-phenotype landscapes with state-of-the-art predictive power

abstract: Large-scale measurements linking genetic background to biological function have driven a need for models that can incorporate these data for reliable predictions and insight into the underlying biochemical system. Recent modeling efforts, however, prioritize predictive accuracy at the expense of model interpretability. Here, we present LANTERN (https://github.com/usnistgov/lantern), a hierarchical Bayesian model that distills genotype-phenotype landscape (GPL) measurements into a low-dimensional feature-space that represents the fundamental biological mechanisms of the system while also enabling straightforward, explainable predictions. Across a benchmark of large-scale datasets, LANTERN equals or outperforms all alternative approaches, including deep neural networks. LANTERN furthermore extracts useful insights into the landscape including its inherent dimensionality, a latent space of additive mutational effects, and novel metrics of landscape structure. LANTERN facilitates straightforward discovery of fundamental mechanisms in GPLs, while also reliably extrapolating to unexplored regions of genotypic-space.


title: Disentangling reward processes underlying payoff maximization from individual differences in gain frequency bias and reinforcement learning

abstract:
Choice selection strategies and decision making are typically investigated using multiple-choice gambling paradigms that require participants to maximize reward payoff. However, research shows that performance in such paradigms suffers from individual biases towards the frequency of gains to choose smaller local gains over larger longer term gain, also referred to as melioration. Here, we developed a simple two-choice reward task, implemented in 186 healthy human adult subjects across the adult lifespan to understand the behavioral, computational, and neural bases of payoff maximization versus melioration. The observed reward choice behavior on this task was best explained by a reinforcement learning model of differential future reward prediction. Simultaneously recorded and source-localized electroencephalography (EEG) showed that diminished theta-band activations in the right rostral anterior cingulate cortex (rACC) correspond to greater reward payoff maximization, specifically during the presentation of cumulative reward information at the end of each task trial. Notably, these activations (greater rACC theta) predicted depressed mood symptoms, thereby showcasing a reward processing marker of potential clinical utility.

Significance StatementThis study presents cognitive, computational and neural (EEG-based) analyses of a rapid reward-based decision-making task. The research has the following three highlights. 1) It teases apart two core aspects of reward processing, i.e. long term expected value maximization versus immediate gain frequency melioration based choice behavior. 2) It models reinforcement learning based behavioral differences between individuals showing that observed performance is best explained by differential extents of reward prediction. 3) It investigates neural correlates in 186 healthy human subjects across the adult lifespan, revealing specific theta band cortical source activations in right rostral anterior cingulate as correlates for maximization that further predict depressed mood across subjects.


title: Rostral Anterior Cingulate Activations inversely relate to Reward Payoff Maximation & predict Depressed Mood


title: Simulation scalability of large brain neuronal networks thanks to time asynchrony

abstract: We present a new algorithm to efficiently simulate random models of large neural networks satisfying the property of time asynchrony. The model parameters (average firing rate, number of neurons, synaptic connection probability, and postsynaptic duration) are of the order of magnitude of a small mammalian brain, or of human brain areas. Through the use of activity tracking and procedural connectivity (dynamical regeneration of synapses), both computational and memory complexities of this algorithm are proved to be theoretically linear with the number of neurons. These results are experimentally validated by sequential simulations of millions of neurons and billions of synapses running in few minutes using a single thread of an equivalent desktop computer.


title: Scalability of large neural network simulations via activity tracking with time asynchrony and procedural connectivity


title: OFF-periods reduce the complexity of neocortical activity during sleep

abstract: Complexity of electroencephalographic signals decreases during slow-wave sleep (SWS), however, the neural mechanism for this decrease remains elusive. Here, we show that this complexity reduction is caused by synchronous neuronal OFF-periods. We analyse in-vivo recordings from neocortical neuronal populations, finding that OFF-periods in SWS trap cortical dynamics, making the population activity more recurrent, deterministic, and less chaotic than during REM sleep or Wakefulness. Importantly, when we exclude OFF-periods, SWS becomes indistinguishable from Wakefulness or REM sleep. In fact, we show that spiking activity for all states has a universal scaling compatible with critical phenomena. We complement these results by a critical branching model that replicates our experimental findings, where we show that forcing OFF-periods to a percentage of neurons suffices to generate a decrease in complexity that replicates SWS.


title: An automated method for precise axon reconstruction from recordings of high-density micro-electrode arrays

abstract:
Neurons communicate with each other by sending action potentials through their axons. The velocity of axonal signal propagation describes how fast electrical action potentials can travel, and can be affected in a human brain by several pathologies, including multiple sclerosis, traumatic brain injury and channelopathies. High-density microelectrode arrays (HD-MEAs) provide unprecedented spatio-temporal resolution to extracellularly record neural electrical activity. The high density of the recording electrodes enables to image the activity of individual neurons down to subcellular resolution, which includes the propagation of axonal signals. However, axon recon-struction, to date, mainly relies on a manual approach to select the electrodes and channels that seemingly record the signals along a specific axon, while an automated approach to track multiple axonal branches in extracellular action-potential recordings is still missing.

In this article, we propose a fully automated approach to reconstruct axons from extracellular electrical-potential landscapes, so-called "electrical footprints" of neurons. After an initial electrode and channel selection, the proposed method first constructs a graph, based on the voltage signal amplitudes and latencies. Then, the graph is interrogated to extract possible axonal branches. Finally, the axonal branches are pruned and axonal action-potential propagation velocities are computed.

We first validate our method using simulated data from detailed reconstructions of neurons, showing that our approach is capable of accurately reconstructing axonal branches. We then apply the reconstruction algorithm to experimental recordings of HD-MEAs and show that it can be used to determine axonal morphologies and signal-propagation velocities at high throughput.

We introduce a fully automated method to reconstruct axonal branches and estimate axonal action-potential propagation velocities using HD-MEA recordings. Our method yields highly reliable and reproducible velocity estimations, which constitute an important electrophysiological feature of neuronal preparations.


title: A Framework for Brain Atlases: Lessons from seizure Dynamics

abstract: Brain maps, or atlases, are essential tools for studying brain function and organization. The abundance of available atlases used across the neuroscience literature, however, creates an implicit challenge that may alter the hypotheses and predictions we make about neurological function and pathophysiology. Here, we demonstrate how parcellation scale, shape, anatomical coverage, and other atlas features may impact our prediction of the brains function from its underlying structure. We show how network topology, structure-function correlation (SFC), and the power to test specific hypotheses about epilepsy pathophysiology may change as a result of atlas choice and atlas features. Through the lens of our disease system, we propose a general framework and algorithm for atlas selection. This framework aims to maximize the descriptive, explanatory, and predictive validity of an atlas. Broadly, our framework strives to provide empirical guidance to neuroscience research utilizing the various atlases published over the last century.


title: Plasmodium falciparum Acetyl-CoA Synthetase is essential for parasite intraerythrocytic development and chromatin modification

abstract: The malaria parasite Plasmodium falciparum possesses a unique Acetyl-CoA Synthetase (PfACS) which provides acetyl moieties for different metabolic and regulatory cellular pathways. We characterized PfACS and studied its role focusing on epigenetic modifications using the var gene family as reporter genes. For this, mutant lines to modulate plasmodial ACS expression by degron-mediated protein degradation or ribozyme induced transcript decay were created. Additionally, an ACS inhibitor was tested for its effectiveness and specificity in interfering with PfACS. The knockdown of PfACS or its inhibition led to impaired parasite growth. Decreased levels of PfACS also led to differential histone acetylation patterns, altered variant gene expression and concomitantly decreased cytoadherence of infected red blood cells containing knocked-down parasites. Further, ChIP analysis revealed the presence of PfACS in many loci in ring stage parasites, underscoring its involvement in the regulation of chromatin. Due to its significant differences to human ACS, PfACS seems an interesting target for drug development.


title: The activity and functions of subarctic soil microbial communities vary across vegetation types

abstract: Increased microbial activity in high-latitude soils due to climate change might lead to higher greenhouse gas (GHG) emissions. However, mechanisms of microbial GHG production and consumption in tundra soils are not thoroughly understood. We analyzed 116 soil metatranscriptomes from 73 sites in the Finnish sub-Arctic to investigate how the diversity and functional potential of bacterial and archaeal communities vary across vegetation types and soil layers. Soils differed in physicochemical conditions, with meadow soils being characterized by higher pH and low soil organic matter (SOM) and carbon/nitrogen ratio whereas dwarf shrub-dominated ecosystems with high SOM and low pH. Actinobacteria, Acidobacteria, Alphaproteobacteria, and Planctomycetes predominated all communities but there were significant differences on genus level between vegetation types, as plant polymer degrading groups were more active in shrub-dominated soils compared to meadows. Given that climate change scenarios predict expansion in dwarf shrubs at high latitudes, our results indicate that the rate of carbon turnover in tundra soils may increase in the future. Additionally, transcripts of methanotrophs were detected in the mineral layer of all soils, potentially moderating methane fluxes from deeper layers. In all, this study provides new insights into possible shifts in tundra microbial diversity and activity with climate change.


title: The activity and functions of soil microbial communities in the Finnish sub-Arctic vary across vegetation types


title: Evolution of Selective RNA Processing and Stabilization operons in cellulosome-harboring Clostridium spp.

abstract: In selective RNA processing and stabilization (SRPS) operons, the stoichiometry of encoded proteins is determined by their respective 3-end stem-loops (SLs), yet the evolution of this mechanism remains elusive. In cellulosomal operons of Clostridium spp., we show that the SLs and their associated genes form a monogamy companionship during the operon evolution. Based on {Delta}G of such SLs, we propose CoSLOE (Composite SL-based Operon Evolution) model with evolutionary ratio (ER) >1 or <1 for positive or negative selection of SRPS operons. In the composite SL-{Delta}G-based tree (CoSL-tree) of cellulosomal operons, when traversing from leafs to the root nodes, ERs reveal diversifying/positive selection towards a less efficient cellulosomal system, consistent with glycoside-hydrolase gene variation both in-operon and genome-wide. A similar pattern is followed by the ATPase operon and the majority of orthologous SRPS operons genome-wide, suggesting conservation among operons in such selection. Thus SRPS operons via their transcript-stabilizing non-coding elements are highlighting a link between operon stoichiometry and operon evolution.


title: Captivity-induced behaviour and spatial learning abilities in an enigmatic, aquifer-dwelling blind eel, Rakthamichthys digressus

abstract: We investigated the impact of captive life on behaviour and learning abilities in an enigmatic, aquifer-dwelling blind eel, Rakthamichthys digressus. Of eight major behavioural traits related to exploration and activity in a novel arena, four were significantly altered by life in captivity. While the startle response upon introduction into the arena and overall swimming away from the walls increased after captivity, inactivity exhibited immediately after the startle and the reaction to an external disturbance decreased. We also observed behavioural syndromes between  startle responses and  horizontal wall following, and between  overall activity and  vertical wall following; however, these behavioural syndromes were not altered by maintenance in captivity. Interestingly, this blind-eel failed to learn a simple spatial task in a Y-maze apparatus. Captive-associated behavioural changes in R. digressus may influence their survival after reintroduction into natural habitats, and such changes must be taken into account while developing protocols for ex-situ conservation and subsequent release.

category: animal behavior and cognition


title: A PMMA bone cement composite that functions as a drug-refillable depot for localized, multi-window chemotherapeutic treatment of bone cancer

abstract: Standard chemotherapy for primary and secondary bone tumors typically involves systemic administration of chemotherapeutic drugs, such as doxorubicin (DOX). However, non-targeted delivery increases dose requirements, and results in off-target toxicity and suboptimal chemotherapeutic efficacy. When chemotherapy is ineffective, substantial resection of tissue and/or total amputation become necessary - a debilitating outcome for any patient. In this work, we developed a proof-of-concept, non-biodegradable, mechanically robust, and refillable composite system for chemotherapeutic (i.e. DOX) delivery comprised of poly(methyl methacrylate) (PMMA) bone cement and insoluble polymeric {gamma}-cyclodextrin ({gamma}-CD) microparticles. The porosity and compressive strength of DOX-filled PMMA composites were characterized. DOX filling capacity, elution kinetics, cytotoxicity against primary osteosarcoma and lung cancer cells, and refilling capacity of composites were evaluated. PMMA composites containing up to 15wt% {gamma}-CD microparticles provided consistent, therapeutically-relevant release of DOX with ~100% of the initial DOX released after 100 days. Over the same period, only ~6% of DOX was liberated from PMMA with free DOX. Following prolonged curing, PMMA composites with up to 15wt% {gamma}-CD surpassed compressive strength requirements outlined by international standards for acrylic bone cements. Compared to DOX-filled PMMA, DOX-filled PMMA/{gamma}-CD composites provided long-term release with decreased burst effect, correlating to long-term cytotoxicity against cancer cells. Refillable properties demonstrated by the PMMA composite system may find utility for treating local recurrences, limiting chemoresistance, and altering drug combinations to provide customized treatment regimens. Overall, findings suggest that PMMA composites have the potential to serve as a platform for the delivery of combinatorial chemotherapeutics to treat bone tumors.


title: Further Theoretical Considerations for Next-Generation Proteomics

abstract: In a recent Journal of Proteome Research paper, I described some general properties and constraints of a hypothetical next generation of proteomics technology based on single-molecule peptide sequencing. This work prompted many interesting questions, both from the reviewers of the initial manuscript and later from readers and colleagues. This follow-up paper addresses some of questions by clarifying the original results, considering alternative metrics, and a number of new simulations. Specifically, the discriminative power of individual amino acids is revisited, simulating additional proteolytic agents. These simulations show allowing missed cleavages generally increases the discriminative power of the amino acids in the proteolytic motif. Additional simulations show the effect of non-ideal conditions modelled on the number of proteins lacking proteotypic reads is very small, and that the average number of proteotypic reads per protein follow the same rule on the performance of the optimal choice of labeled amino acids as the number of distinguishable proteins in NeXtProt. The goal of this paper is to expand prior results and continue the scientific discussion on the possibilities of future proteomics technologies.


title: Workflows for rapid functional annotation of diverse arthropod genomes

abstract:
Genome sequencing of a diverse array of arthropod genomes is already underway and these genomes will be used to study human health, agriculture, biodiversity and ecology. These new genomes are intended to serve as community resources and provide the foundational information that is required to apply  omics technologies to a more diverse set of species. However, biologists require genome annotation to use these genomes and derive a better understanding of complex biological systems. Genome annotation incorporates two related but distinct processes: demarcating genes and other elements present in genome sequences (structural annotation); and associating function with genetic elements (functional annotation). While there are well established and freely available workflows for structural annotation of gene identification in newly assembled genomes, workflows for providing the functional annotation required to support functional genomics studies are less well understood. Genome-scale functional annotation is required for functional modeling (enrichment, networks, etc.) and a first-pass genome-wide functional annotation effort can rapidly identify under-represented gene sets for focused community annotation efforts. We present an open source, open access and containerized pipeline for genome-scale functional annotation of insect proteomes and apply it to a diverse range of arthropod species. We show that the performance of the predictions is consistent across a set of arthropod genomes with varying assembly and annotation quality.Complete instructions for running each component of the functional annotation pipeline on the command line, a high performance computing cluster and the CyVerse Discovery Environment can be found at the readthedocs site (https://agbase-docs.readthedocs.io/en/latest/agbase/workflow.html).

Simple summaryGenomic technologies are accumulating information about genes at a faster rate than ever before, and sequencing initiatives like the Earth Biogenome Project, i5k and Ag100Pest are expected to increase this rate of acquisition. However, if genomic sequencing is to be used for improvement of human health, agriculture and our understanding of biological systems, it is necessary to identify genes and understand how they contribute to biological outcomes. While there are several well-established workflows for assembling genomic sequences and identifying genes, understanding gene function is essential to create actionable knowledge. Moreover this functional annotation process must be easily accessible and provide information at a genomic scale to keep up with new sequence data. We report a well defined workflow for rapid functional annotation of whole proteomes to produce Gene Ontology and pathways information. We test this workflow on a diverse set of arthropod genomes and compare it to common arthropod reference genomes. The workflow we described is freely and publicly available via a web interface on CyVerse or as biocontainers that can be deployed scalably on local computing systems.


title: FiberSim: a flexible open-source model of myofilament-level contraction

abstract:
FiberSim is a flexible open-source model of myofilament-level contraction. The code uses a spatially explicit technique, meaning that it tracks the position and status of each contractile molecule within the lattice framework. This allows the model to simulate some of the mechanical effects modulated by myosin binding protein-C, as well as dose-dependence of myotropes and the effects of varying isoform expression levels. This paper provides a short introduction to FiberSim and presents simulations of tension-pCa curves with and without regulation of thick filament transitions by myosin-binding protein C. The software was designed to be flexible (the user can define their own model and/or protocol) and computationally efficient (simulations can be performed on a regular laptop). We hope that other investigators will use FiberSim to explore myofilament level mechanisms and to accelerate research focusing on the contractile properties of sarcomeres.

Statement of significanceMyotropes, such as omecamtiv mecarbil and mavacamten, are new therapeutics that bind directly to sarcomeric proteins. Their clinical development has reenergized interest in mechanistic understanding of sarcomere level function. FiberSim is an open-source spatially-explicit computer model that simulates myofilament level mechanics and can predict how modulating the function of a sarcomeric protein will impact contractility. The software was specifically designed to be flexible and user-friendly and may help to further accelerate myofilament-research.

category: biophysics


title: TMEM120 is a coenzyme A-binding membrane protein with structural similarities to ELOVL fatty acid elongase

abstract: TMEM120A, also named as TACAN, is a novel membrane protein highly conserved in vertebrates and was recently proposed to be a mechanosensitive channel involved in sensing mechanical pain. Here we present the single particle cryo-EM structure of human TMEM120A which forms a tightly packed dimer with extensive interactions mediate by the N-terminal coiled coil domain (CCD), the C-terminal transmembrane domain (TMD), and the re-entrant loop between the two domains. The TMD of each TMEM120A subunit contains six transmembrane helices (TMs) and has no clear structural feature of a channel protein. Instead, the six TMs form an -barrel with a deep pocket where a coenzyme A (CoA) molecule is bound. Intriguingly, some structural features of TMEM120A resemble those of elongase for very long-chain fatty acid (ELOVL) despite low sequence homology between them, pointing to the possibility that TEME120A may function as an enzyme for fatty acid metabolism, rather than a mechanosensitive channel.


title: The extra-embryonic space is a geometric constraint regulating cell arrangement in nematodes

abstract: In multicellular systems, cells communicate with adjacent cells to decide their positions and fates. Cellular arrangement in space is thus important for development. Orientation of cell division, cell-cell interaction (i.e., attraction and repulsion), and geometrical constraints are the three major factors that define cell arrangement. Here we found that the amount and location of extra-embryonic space (ES), the empty space within the eggshell not occupied by embryonic cells, are critical to define cell arrangement in the 4-cell stage embryo of nematodes. This discovery was motivated by observations of a T-reversed-type arrangement, which was not explained by a model assuming simplified shapes of the eggshell, in our previous experiments. In this study, we incorporated the precise shape of the C. elegans eggshell in our newly developed multicellular morphology model based on the phase-field method. The new model succeeded in reproducing the T-reverse arrangement, demonstrating the importance of the precise shape of the eggshell. Further analyses revealed that the amount and location of ES is critical to develop various cell arrangements. Overall, our analyses characterized the roles of new geometrical contributors to cell arrangements, which should be considered for any multicellular system.


title: Shaping liposomes by cell-free expressed bacterial microtubules

abstract: Genetic control over a cytoskeletal network inside lipid vesicles offers a potential route to controlled shape changes and DNA segregation in synthetic cell biology. Bacterial microtubules (bMTs) are protein filaments found in bacteria of the genus Prosthecobacter. They are formed by the tubulins BtubA and BtubB which polymerize in the presence of GTP. Here, we show that the tubulins BtubA/B can be functionally expressed from DNA templates in a reconstituted transcription-translation system, thus providing a cytosol-like environment to study their biochemical and biophysical properties. We found that bMTs spontaneously interact with lipid membranes and display treadmilling. When compartmentalized inside liposomes, de novo synthesized BtubA/B tubulins self-organize into cytoskeletal structures of different morphologies. Moreover, bMTs can exert a pushing force on the membrane and deform liposomes, a phenomenon that can be reversed by light-activated disassembly of the filaments. Our work establishes bMTs as a new building block in synthetic biology. In the context of creating a synthetic cell, bMTs could help shape the lipid compartment, establish polarity or directional transport, and assist the division machinery.


title: Radiosynthesis and preclinical evaluation of a carbon-11 labeled PDE7 inhibitor for PET neuroimaging

abstract:
BackgroundDysfunction of cyclic nucleotide phosphodiesterase 7 (PDE7) has been associated with excess intracellular cAMP concentrations, fueling pathogenic processes that are implicated in neurodegenerative disorders. The aim of this study was to develop a suitable PDE7-targeted positron emission tomography (PET) probe that allows non-invasive mapping of PDE7 in the mammalian brain.

MethodsBased on a spiro cyclohexane-1,4-quinazolinone scaffold with known inhibitory properties towards PDE7, we designed and synthesized a methoxy analog that was suitable for carbon-11 labeling. Radiosynthesis was conducted with the respective desmethyl precursor using [11C]MeI. The resulting PET probe, codenamed [11C]26, was evaluated by cell uptake studies, ex vivo biodistribution and radiometabolite studies, as well as in vivo PET experiments in rodents and nonhuman primates (NHP).

ResultsTarget compound 26 and the corresponding phenolic precursor were synthesized in 2-3 steps with overall yields of 49.5% and 12.4%, respectively. An inhibitory constant (IC50) of 31 nM towards PDE7 was obtained and no significant interaction with other PDE isoforms were observed. [11C]26 was synthesized in high molar activities (170 - 220 GBq/{micro}mol) with radiochemical yields of 34{+/-}7%. In vitro cell uptake of [11C]26 was 6-7 folds higher in PDE7 overexpressing cells, as compared to the controls, whereas an in vitro specificity of up to 90% was measured. Ex vivo metabolite studies revealed a high fraction of intact parent in the rat brain (98% at 5 min and 75% at 30 min post injection). Considerable brain penetration was further corroborated by ex vivo biodistribution and PET imaging studies - the latter showing heterogenic brain uptake. While marginal specific binding was observed by PET studies in rodents, a moderate, but dose-dependent, blockade was observed in the NHP brain following pretreatment with non-radioactive 26.

ConclusionIn this work, we report on the preclinical evaluation of [11C]26 (codename [11C]P7-2104), a PDE7-targeted PET ligand that is based on a spiroquinazolinone scaffold. [11C]26 displayed promising in vitro performance characteristics, a moderate degree of specific binding in PET studies with NHP. Accordingly, [11C]26 will serve as a valuable lead compound for the development of a new arsenal of PDE7-targeted probes with potentially improved in vivo specificity.


title: Feedback alignment with weight normalization can provide a biologically plausible mechanism for learning

abstract: The mechanism by which plasticity in millions of synapses in the brain is orchestrated to achieve behavioral and cognitive goals is a fundamental question in neuroscience. In this regard, insights from learning methods in artificial neural networks (ANNs) and in particular the idea of backpropagation (BP) seem inspiring. However, the implementation of BP requires exact matching of forward and backward weights, which is unrealistic given the known connectivity pattern in the brain (known as "weight transport problem"). Notably, it is recently shown that under certain conditions, error BackPropagation Through Random backward Weights (BP-TRW), can lead to partial alignment of forward and backward weights overtime (feedback alignment or FA) and result in surprisingly good accuracies in simple classification tasks using shallow ANNs. In this work, we took a closer look at FA to find out why it occurs when using BP-TRW and explored ways to boost it for deep ANNs. We first show that the gradual alignment of forward and backward weights arises from the successive application of BP-TRW update rule on forward weights regardless of learning or loss function if error signals and outputs of neurons satisfy certain conditions such as when they are autocorrelated. Moreover, we show that FA in deeper networks can be improved significantly by applying a biologically-inspired weight normalization (WN) to the input weights of each neuron. In addition, WN can improve the performance of both BP and BP-TRW when class labels are changed across time, an under-explored phenomenon in ANNs which is crucial for flexible learning in the brain in everyday life. With WN, BP-TRW test accuracy can almost match that of BP following class label changes. Altogether, our results portray a clearer picture of the FA mechanism and provide evidence for how learning can occur using BP-like mechanisms while abiding by biological limits on synaptic weights.


title: Multimodal Single-Cell Characterization of the Human Granulocyte Lineage

abstract: High throughput single cell transcriptomics (scRNA-seq) has been successfully applied to characterize immune cell heterogeneity across a diverse range of settings; however, analysis of human granulocytes remains a significant challenge due to their low gene expression transcript detection. Consequently, granulocytes are typically either absent or highly under-represented and inaccurately enumerated in most human scRNA-seq datasets. Here, we apply multi-modal CITE-seq profiling to characterize granulocytes in human whole blood and bone marrow, and we show that these populations can be accurately detected and analyzed using the antibody-based modality, and that their frequencies and phenotype align well with antibody-based characterization of the same samples using CyTOF. These analyses also clearly highlight extremely low gene transcript detection across the entire granulocyte lineage including the earliest neutrophil progenitor populations when using the 10X Genomics platform. By contrast, when performing parallel analyses of the same samples using the BD Rhapsody platform, we recovered a much higher proportion of granulocyte gene transcripts, enabling true multi-modal characterization of human granulocyte heterogeneity.

category: immunology


title: Protein kinase J regulates Rv0642c expression and sensitivity to rifampicin of mycobacteria

abstract: Mycobacterium tuberculosis (M.tb) persists for long-duration inside the human host in both active and latent form by modulating the immune response. The mechanisms employed by M.tb to survive inside the host and evade the host immune system need to be explored in greater depth for the rational design of novel treatment strategies. The phosphorylation and methylation of biomolecules need to be addressed in mycobacteria because it has an important role in infection establishment and persistence. In the present study, we elaxborate on the role of PknJ in the slow growth of BCG and its association with mmaA4 protein during extracellular and intracellular growth. The pknJ knock-out (KO) BCG has been used to decode the functional significance in mycobacteria. The mmaA4 expression and methyltransferase activity is decreased in knocked-out BCG strain (pknJ-/-) during extracellular growth, while mmaA4 expression and methyltransferase activity is increased during intracellular growth of mycobacteria. The knocked-out BCG strain is highly sensitive to the rifampicin antibiotics during extracellular growth in compared to control. A significant association of pknJ and mmaA4 was found in our studies during the growth and intracellular persistence of mycobacteria.